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The Point
These things have one common denominator: first-year media communication major
Van Soest has gone viral on Instagram and TikTok for recording people’s reactions as he knocks on their doors to create engaging videos in Point Loma Nazarene University’s freshman dorm halls
He’s brought in a local mariachi band
dressed up as Santa and even served breakfast in front of students’ doors
It all started when Van Soest’s roommate sent him a video of people knocking on doors and posing in front of them to see the occupant’s reaction
Since Van Soest had prior experience making TikTok videos
his roommate urged him to make a similar video in their dorm hall
but the views increased once Van Soest recorded his “human pyramid” video
“[I] knocked on people’s doors
and made the pyramid bigger and bigger,” Van Soest said
and there’s so much we can do with this
creative ideas and ways of pranking people at their dorms.”
His most popular video has 18.5 million views on Instagram and 62.2 million on TikTok
Van Soest knocked on Angelica Moser’s door
Moser and her roommates couldn’t stop laughing as Van Soest walked around their room
“We thought it was the funniest thing ever,” Moser said
“It just brought so much excitement and light to our room
… You can tell that he’s doing it to serve the Lord
bring people together and just bring joy and excitement to Hendricks.”
they might come across Van Soest’s videos and move on after getting a good laugh
his videos have become more than just funny content for social media
They’ve built an undeniable sense of community
“I think Hendricks as a dorm is closer because of these videos,” Van Soest said
“I can’t even take credit for all the videos
because a lot of my friends give me the ideas and I wouldn’t be able to film without them
It’s been really fun to see the community that’s formed.”
a first-year business administration major
lives across the hall from Van Soest and met him during Welcome Week
Palm has participated in many of Van Soest’s dorm pranks and echoed that these videos have brought community to Hendricks
“I’ve met so many people [on every floor],” Palm said
“But not only have [these videos] brought Hendricks together
and I met 10 to 15 people just from making videos there.”
While Van Soest naturally brought in views from people in his dorm hall
the millions of views he’s received on social media have been organic
the virality of his videos has been random
“I can’t say I fully know or can fully take credit for any of the success because a lot of it is luck,” Van Soest said
“I think some videos deserve to go viral that don’t
and some videos don’t necessarily deserve to go viral that do.”
Van Soest said that college content is a niche subject that doesn’t often get covered on social media
which could also contribute to his success
But Van Soest emphasized that if you want to make videos like his
it’s more important to have fun with it than to get caught up in the numbers
“If you’re not having fun with it
you shouldn’t be doing it,” Van Soest said
“The goal in mind shouldn’t be to chase the high numbers
That doesn’t bring any sort of fulfillment
So if you enjoy the videos and the content you’re making
you’ll end up really enjoying the process
the better you’ll get at it and the more fun it’ll become.”
he said he’s feeling out where he wants to take his social media pages and is seeing what the Lord leads him to do next
Van Soest’s viral dorm hall videos can be found on Instagram @westonvansoest
Mary’s in Duluth. She was born August 19
the daughter of Neal and Florence (Rystad) Van Soest. She was a graduate of Biwabik High School and went on to receive her bachelor’s degree in Medical Record Administration from St
Scholastica. Judith was united in marriage to Roger Paul Linden on June 18
Judy worked in Medical Records Administration at the Swedish Hospital in Minneapolis
and Olmstead Community Hospital in Rochester
MN before working as a Medical Transcriptionist at the East Range Clinic in Virginia
MN. She retired in 2010. She was a member of the First Covenant Church in Virginia where she was the church pianist for many years. She enjoyed spending time with her family and friends
Judy is survived by her husband of 58 years
MN; siblings: Dorothy (Susan) Van Soest of Seattle
and Richard (Lynette) Van Soest of Coon Rapids
and Thomas Linden; and numerous extended family and friends
She was preceded in death by her parents
Visitation will be from 1:00 p.m
at the Biwabik Covenant Church. In leu of flowers
please consider a donation to the Mesabi Humane Society.
Family services are provided by Bauman-Cron
a Bauman Family Funeral Home in Virginia. To express condolences online please see: www.baumanfuneralhome.com
Mesabi Humane Society2305 Southern Drive, Virginia MN 55792Tel: 1-218-741-7425Email: info@mesabihumanesociety.orgWeb: https://www.mesabihumanesociety.org/donate
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NORTH AMERICAN PREMIERE After living in darkness for nearly 40 years
becomes one of the first people in the world to receive an experimental bionic eye implant
as the groundbreaking scientific advancement offers hope alongside perplexing technical limitations
as director Landon Van Soest presents impressionistic visualizations of Nichols’ enhanced sight
inviting questions about the nature of our perception
The first screening will be followed by a Q&A with director Landon Van Soest
and producers Jo Budzilowicz and Stacey Kleiger
All in-person screening venues provide sound amplification headphones upon request with venue management
IFC Center can also provide a T-Coil loop for compatible devices
Due to a recent equipment failure at Village East Cinemas, the room where this film screens is currently not accessible to patrons using wheelchairs. We apologize for the inconvenience. However the film is available to view at home as part of DOC NYC’s online festival. Please reach out to info@docnyc.net with any questions
Don't forget to add SUBMISSIONS to the email subject line
Both the podcast Bach & Co and all the episodes of Bach ad Infinitum in the week from 9 September to 13 September feature Peter Kooij
Peter Kooij is an internationally renowned bass singer and has sung on many CDs with Bach cantatas
Peter Kooij has been the bass singer with Philippe Herreweghe for many years
he sings on Masaaki Suzuki’s cantata series
with Rudolf Lutz and has contributed to many other CD recordings
His partner is the well-known contralto Margot Oitzinger and they live together with their family in Soest
Peter Kooij will soon be 70 and he thought this was a great opportunity to release all the Bach solo cantatas for bass and alto on a double CD together with his wife
That was of course reason enough to invite them both to our podcast Bach & Co
This invitation led to a very nice conversation with them
When Govert Jan Bach encounters Peter Kooij as a soloist in a cantata
he always announces him as: “Our Peter Kooij from Soest”
This world-famous star lives only a 20-minute bike ride from where we record Bach & Co
that he flew to Japan 5 times a year for 20 years for Masaaki Suzuki’s cantata series
He prefers to stay closer to home and he tries to coordinate his agenda as much as possible with that of his wife
Margot will soon be leaving for concerts in Taiwan
The entire conversation with Peter and Margot can be listened to in the Bach & Co podcast
We have declared the Bach ad Infinitum week that starts on September 9 as ‘Peter Kooij’ week
with a cantata by him or his wife Margot every day
Now she is also “OUR Margot from Soest”
The podcast Bach & Co is available from Monday 9 September on the podcast platforms
one of the most influential animal scientists of his generation
Van Soest’s pioneering work permanently changed the chemical and in vitro analysis of feeds and the understanding of herbivore nutrition
His novel and revolutionary methods of analysis created the standard for how researchers measure the nutritional value of ruminating animal forages worldwide
“Van Soest was a one-of-a-kind scientist and professor
but he was also a unique individual with a myriad of interests – a modern Renaissance man,” said David Mertens
president of Mertens Innovation Research LLC and a former graduate student and friend of Van Soest
“His curiosity knew few bounds and he had the passion and intellect to pursue whatever interested him.”
Army as a biochemist at the Walter Reed Army Institute of Research in Silver Spring
Van Soest joined the Dairy Cattle Research Branch of the Animal Husbandry Research Division at the U.S
Department of Agriculture-Agriculture Research Service in Beltsville
He joined Cornell’s Department of Animal Science in 1968
and retired as a professor emeritus in 1995
He continued to publish papers and informally teach graduate students into his 90th year
One major area of Van Soest’s research involved developing a nutritionally relevant fiber analysis
Van Soest’s detergent analysis gave researchers a way to precisely separate the chemical components of feeds into three categories: parts that were digestible to all animals; indigestible without fermentation; and completely indigestible
This work offered new ways to measure chemical compositions and the exact energy output of feeds
Van Soest’s detergent analysis has been expanded and adopted worldwide
replacing a less accurate system that had been used since the 1860s. Further
his work in fiber chemistry was adopted by human nutritionists and researchers interested in the effects of dietary fiber on gut health and obesity.
Peter Van Soest was also a consummate teacher and mentor – he loved to teach at every opportunity,” Mertens said
“Innumerable students benefitted from his knowledge and ability to explain complicated concepts in ways that made them easier to understand.”
One of his other significant contributions was as a co-inventor of the Cornell Net Carbohydrate and Protein System
a cattle nutrition model currently used to formulate diets for over 70% of dairy cattle in North America and used in over forty other countries.
“Peter’s ability to translate his methods and understanding of biological systems into a functional quantitative framework fully characterizes his ability to synthesize data and provide solutions which are still applicable today,” said Mike Van Amburgh Ph.D
professor of animal science and also a former student
Van Soest’s numerous honors included awards from the American Dairy Science Association and the American Society of Animal Science
election as a fellow of the American Institute of Nutrition
and an honorary doctorate from the University of Milan in 1992
An accomplished musician and music historian
Van Soest is survived by his ex-wife Marian Van Soest
Gifts may be made to Ithaca Hospicare in Peter’s name and a graduate student scholarship is being developed in his name
Get Cornell news delivered right to your inbox
A large fire raged in a restaurant in Soest on Monday morning
including apartments located above the restaurant
Veiligheidsregio Utrecht
the office that covers the region’s emergency services
first reported blaze at restaurant Bauers on Burgemeester Grothestraat at 2:41 a.m
“Several fire engines have been dispatched to Grothestraat in Soest
where smoke is coming from the roof,” the Veilighedsregio said
“It is not yet known what exactly is on fire
the Veiligheidsregio reported that it had escalated the situation to GRIP 1
which means that all the emergency services present now work under one commanding officer
The smoke was spreading through the neighborhood and the emergency services urged locals to keep their doors and windows closed
Firefighters got the blaze under control at around 6:33 a.m
Evacuated locals will have to wait for the fire brigade to check their homes before they can return
The Veiligheidsregio warned road users in the area to beware of slippery conditions
Extinguishing water combined with freezing temperatures resulted in icy roads
A gritter truck was at the scene to spread salt against the iciness
The GLORY Kickboxing women’s super bantamweight Tiffany van Soest recently announced that she will be retiring after her next match against Sarah Moussaddak at GLORY 88
Tim Wheaton of Combat Press spoke with the decorated world champion in which Tiffany van Soest explained:
“I’m happy with what I’ve done and I want to live my life and pursue a career in helping people
the American athlete van Soest will look to defend her GLORY Kickboxing throne against the French-Moroccan Sarah Moussaddak
The two had met previously with van Soest earning a victory in late 2022
GLORY 88 will be broadcast live from Paris
The 34-year-old “Time Bomb” Tiffany van Soest has put together an incredible career in combat sports
she would earn national titles in the sport
she would transition to Muay Thai and kickboxing where she would earn impressive titles in organizations such as Lion FIghts
Tiffany van Soest, 34, made the retirement announcement earlier today at a press conference
“Following the fight in Paris on Sep
This is going to be my last fight … When I started my journey in fighting
I don’t know if it was a deeper inner-knowing
but the number 35 was always in my mind to stop … I owe it to myself and everybody who has been a part of my journey
to go out on top and go out with honor and in the best way possible.”
GLORY 88GLORY KickboxingSarah MoussaddakTiffany Van Soest
Metrics details
Over 100 countries have set or are considering net-zero emissions or neutrality targets
most of the information on emissions neutrality (such as timing) is provided for the global level
we look at national-level neutrality-years based on globally cost-effective 1.5 °C and 2 °C scenarios from integrated assessment models
These results indicate that domestic net zero greenhouse gas and CO2 emissions in Brazil and the USA are reached a decade earlier than the global average
and in India and Indonesia later than global average
These results depend on choices like the accounting of land-use emissions
The results also show that carbon storage and afforestation capacity
and transport sector emissions affect the variance in projected phase-out years across countries
We further compare these results to an alternative approach
using equity-based rules to establish target years
These results can inform policymakers on net-zero targets
directly relevant for national policy-making and international negotiations
Although IAMs have developed to represent individual countries and current climate policies in more detail
IAMs are not the only tools for analyses such as presented here—national energy system models
These tools are already applied jointly to develop national-level pathways that account for national circumstances but still meet the global goals of the Paris Agreement
The results that we present here should be complemented with an assessment of feasible reductions at the national level
considerations of equity and national model results
Individual models are indicated by symbols
whereas the bars show the minimum–maximum range (enlarged circles: model median)
individual models show a phase-out after 2100 in the extrapolated data (indicated by an asterisk) or no phase-out at all (#)
Diamonds plotted at the 2030 mark indicate a change between the 2 °C and 1.5 °C scenario in terms of a country reaching net zero earlier than
Vertical dotted lines indicate the global average phase-out year
India and Turkey show a later than global average phase-out in the 2 °C scenario (only India and Japan in the 1.5 °C scenario)
The other countries have a phase-out year comparable to the global average
Comparing the phase-out years for CO2 emissions with those for only fossil CO2 shows that countries in which land use is a source of emission (e.g.
Indonesia) will see a later phase-out of CO2 than of fossil CO2 only
whereas in countries in which land-use forms a sink (e.g
this means that Brazil and the United States typically have a phase-out year earlier than the global average
whereas India is projected to reach net-zero emissions later than the global average (in four out of six scenario–source combinations)
China and the EU are relatively similar to the global average (namely in four out of six scenario–source combinations and later than global average in the remaining two)
The remaining five countries show a mixed picture: results vary across sources of emissions and temperature targets
Supplementary Table 9 shows additional information on the emissions projections
to support thinking about linking longer-term
net-zero emissions goals to shorter-term action such as formulated in Nationally Determined Contributions (NDCs)
GHG emissions are projected to peak in 2020 in many countries that have not yet seen peak emissions and be reduced by between 12% (India) and 36% (Japan
Canada and Indonesia) by 2030 relative to 2015 levels
these reductions amount to 52% (Brazil) to 72% (USA)
and up to 90% (USA) under the 1.5 °C scenario
A number of technical issues has a strong influence on the reported phase-out year at the national level. We explore four that are highly debated but not yet in the context of neutrality targets, i.e., the use of inventory data for LULUCF-related emissions, the allocation of negative emissions, the GWPs and equity considerations (respectively, Fig. 2a–d).
India and Turkey are not shown for the 2 °C scenario
because the equity range included 2100 (which may actually mean somewhere after 2100)
while the cost-optimal median phase-out year was calculated as being beyond 2100 in these two cases
Individual models are indicated by symbols
whereas the error bars show the minimum–maximum range from models (enlarged circle: median)
Extrapolated emissions data were used to calculate the phase-out year difference
so as to not introduce a bias when calculating differences in phase-out years
Vertical lines at 0 indicate no difference between the default and sensitivity cases
resulting in a later phase-out when using inventory data
whereas the cost-optimal scenarios included LULUCF
This difference could lead to earlier phase-out years in this study (on a global level: 10–20 years)
The comparison showed that when taking a different equity approach
many of the countries studied here would have to phase out GHG emissions earlier than under a cost-optimal allocation
notably developed countries such as Canada and the EU
Brazil would be allowed to phase out emissions later
as well as other countries with lower per-capita emissions or developing economies
This implies that countries with later equity-based phase-out years could receive support from countries with earlier equity-based phase-out years
to help them meet their earlier domestic targets
See Supplementary Table 2 for details of how the variables were calculated (units are displayed in the lower left corner of each panel)
Six turned out to be the optimal number of variables (see “Methods”)
The model would then be (uncertainty range indicates two times SE):
Where CCSshare stands for CO2 uptake from CCS as share of net total GHG emissions in 2050
Afforestation refers to CO2 uptake from afforestation and reforestation in 2050
transport share is the share of transportation emissions in total CO2 emissions in 2015
nonCO2share is the share of non-CO2 emissions in total GHG emissions in 2015
GDPcap is the gross domestic product (GDP) per capita in 2015
and forestshare is the share of forests in total land cover in 2015
A more parsimonious (simpler) model would contain only the variables with p-value smaller than 0.05
That model has slightly lower explanatory power
but the benefit is having further reduced the number of explanatory variables
The formula for the final model then becomes:
The signs can be explained as follows: the larger the CCS capacity and afforestation
the more potential for negative emissions contributing to faster reductions and an earlier phase-out year
The higher the current share of non-CO2 emissions
the more difficult to decarbonize so the later the phase-out
the stronger the growth in emissions; thus
A higher GDP per capita could also imply greater capacity or willingness to mitigate emissions
The share of transport emissions showing a negative correlation is less straightforward
It seems to imply that this sector is relatively easy to decarbonize
but not for freight and also not for international aviation
countries with a relatively large share of transport emissions often also have a relatively high GDP and smaller baseline emissions growth
A large transport share could also imply slower growth of this sector and smaller shares of other
All of these factors would contribute to earlier phase-out
Emissions in the phase-out year of GHG (year indicated per model—focusing on the same two models as in the previous section
by greenhouse gas (colours) and country (panels)
focusing on a country with an average phase-out year (b China)
and two with an early projected phase-out of GHG emissions (a Brazil and d USA)
Positive numbers denote remaining emissions of CH4
whereas negative numbers denote negative emissions in energy supply and in Agriculture
Forestry and Other Land Use (IPCC Category 3)
CO2 from energy supply includes CO2 emissions from fuel combustion and fugitive emissions from fuels: electricity and heat production and distribution (IPCC category 1A1a)
including pipeline transportation (IPCC category 1A3ei)
fugitive emissions from fuels (IPCC category 1B) and emissions from carbon dioxide transport and storage (IPCC category 1C)
Negative emissions in this sector result from the use of (BE)CCS
It should be noted that Brazil presents an exception for many indicators
as it has a relatively large share of non-CO2 emissions but an early phase-out
This can be explained by the breakdown of emissions in the phase-out year
which shows that a large potential for negative emissions can compensate for those remaining emissions
Other countries with an early phase-out (USA) generally also have a relatively large potential for negative emissions
Countries with a late phase-out (India and
China and the EU) have relatively large remaining emissions of both CO2 and non-CO2 GHGs
We analysed when major emitting countries are projected to reach CO2 and GHG emissions neutrality using 1.5 °C and 2 °C scenarios from IAMs
We also looked into the question how this depends on definitions and the reasons behind differences between countries
the United States (CO2 and all GHGs) and Japan (GHG only) are projected to have an earlier phase-out year than the global average
India and Indonesia typically have a late phase-out year
the phase-out year is typically near the global average
the global average is different for CO2 and all GHGs
The model spread is fairly large for Brazil and India
and is smaller for the United States and the EU
Definition factors (such as harmonization of data in the base year and the allocation of negative emissions) play a role in the phase-out year and this works out differently for different countries
These findings highlight the importance of clear definitions and political agreement on issues such as the use of land-use data and allocation of negative emissions
When harmonizing the model projections towards the countries’ reported net land-use emissions estimates in their GHG inventories
net-zero GHG emissions are projected to be reached earlier in all countries
The difference between inventory data and the model output for net land-use emissions is caused by a systematic difference in definition of anthropogenic land sources and sinks
The differences between these data sources are relatively large for China
When allocating negative emissions from biomass with CCS (BECCS) to the biomass-producing country instead of the carbon-storing country
with a large model range for Brazil and India)
Updating GWPs from IPCC AR4 to IPCC AR5 values does not significantly affect phase-out years
Applying equity approaches rather than a cost-optimal allocation of mitigation effort would imply earlier phase-out years for many of the countries studied here
but later phase-out years for Brazil and other countries with lower per-capita emissions or developing economies (e.g.
The multiple linear regression showed that factors affecting negative emissions (e.g.
afforestation and CCS) explain the lion’s share of the variance in phase-out years
Mitigation potential and especially the potential for negative emissions are dominant factors
determining when a country can reach net-zero emissions
as well as the current shares of transport emissions
have the strongest relationship with phase-out years (negative for the former three
In addition to showing a relatively large potential for negative emissions
countries with a projected early phase-out (Brazil and the United States) generally have relatively low emission levels of CO2 from the energy demand sectors
a low current share of non-CO2 emissions (except Brazil) and low population density
That potential for negative emissions is high enough in Brazil to compensate for its relatively high levels of non-CO2 emissions
Countries with late phase-out (India and Indonesia
and to a smaller extent also China and the EU) show the reverse pattern and have relatively large remaining emissions of both CO2 and non-CO2 GHGs
which suggest a significant share of mitigation costs could be covered by accounting for air quality and other co-benefits
making additional domestic mitigation more attractive
Canada’s foreseen 2050 net-zero emissions goal does not specify whether it would apply to all GHG or CO2 only
but both would need to be phased out slightly earlier than 2050 to be in line with the models’ cost-optimal 1.5 °C scenarios (for 2 °C
2050 net-zero emissions would suffice according to these models)
the specification of target coverage is important
Our findings show that to meet these targets
countries should pay special attention to enhancing the capacity to realize negative emissions
clearly specify the land-use emissions accounting and related data (especially important for Canada and the USA)
agree on the accounting of negative emissions from BECCS (important for Brazil and Japan) and clarify their approach to equity and the use of ITMOs (all countries)
could shed light on governments’ capacity to implement net-zero targets
Our results can inform the national target setting
as they present an advancement in knowledge on national-level results from IAM scenarios
The results notably address the Talanoa Dialogue questions of Where do we want to go
They can also inform international negotiations related to Article 6 and methodological choices
such as LUC data and accounting for negative emissions from BECCS
non-state actors can help their governments define realistic and potentially more ambitious targets
we used standardized variables given their different units
We only used the projections by the POLES and IMAGE models for the multiple linear regression
because these are the only two models that cover all ten countries
that data subset had an equal number of records for each country (i.e.
while still representing more than one model for robustness
The two budgets represent limiting global warming to below 2 °C during the twenty-first century and below 1.5 °C in 2100 with more than 66% probability
cost-optimal mitigation was assumed to start in 2020 (i.e.
emission reductions where and when they are cheapest to achieve)
it was assumed that only existing policies were implemented (historical data up to 2020 was not yet available when these scenarios were developed between 2016 and 2018)
Non-CO2 emissions were taxed with the same carbon price as that of CO2 in the cost-optimal scenarios
Emission pathways for the ten countries were linearly extrapolated to 2200 based on the 2050–2100 trajectory
to estimate the phase-out years beyond 2100 where needed
We used the CO2-equivalent emissions based on GWPs from IPCC AR4 (time horizon of 100 years) as default and show the effect of using those from AR5
The text of the Paris Agreement leaves the choice of metric open and refers to the common metrics assessed by the IPCC
Model results can be found in the open access CD-LINKS scenario explorer https://data.ene.iiasa.ac.at/cd-links/. Policy-relevant data are available in the Global Stocktake tool https://themasites.pbl.nl/o/global-stocktake-indicators/
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and Efforts to Eradicate Poverty (eds Masson-Delmotte
Zero emission targets as long-term global goals for climate protection
Emissions: world has four times the work or one-third of the time
Mitigation Pathways Compatible with 1.5 °C in the Context of Sustainable Development in Global Warming of 1.5 °C
Paris agreement zero emissions goal is not always consistent with 2 °C and 1.5 °C temperature targets
Implications of possible interpretations of ‘greenhouse gas balance’ in the Paris Agreement
New use of global warming potentials to compare cumulative and short-lived climate pollutants
CD-LINKS project. CD-LINKS Scenario Database, https://db1.ene.iiasa.ac.at/CDLINKSDB (IIASA
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Comparing transformation pathways across major economies
The key role of forests in meeting climate targets requires science for credible mitigation
Reconciling global-model estimates and country reporting of anthropogenic forest CO2 sinks
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Stable climate metrics for emissions of short and long-lived species-combining steps and pulses
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Download references
The authors received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement no
The work is largely based on scenarios from integrated assessment modelling studies published in the CD-LINKS database (financed by European Union’s Horizon 2020 research and innovation programme under grant agreement number 642147)
We are very grateful to the model teams who submitted the scenarios that were used here
Christoph Bertram (REMIND) and Laurent Drouet (WITCH)
Kaj-Ivar van der Wijst and Hans Visser (PBL) for their invaluable contributions
PBL Netherlands Environmental Assessment Agency
Copernicus Institute of Sustainable Development
contributed to the analysis and article review
The authors declare no competing interests
Peer review information Nature Communications thanks Yann Robiou de Pont and Katsumasa Tanaka for their contribution to the peer review of this work
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations
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Partly cloudy skies. Low 33F. Winds NNE at 5 to 10 mph.
A celebration of life for Hugh Gardner Soest is scheduled for 11 a.m. Friday at First Baptist Church.
A celebration of life for Hugh Gardner Soest is scheduled for 11 a.m. Friday at First Baptist Church. In lieu of flowers, donations may be made to First Baptist Church, Turning Point of Jackson Hole or the Jackson Hole Historical Society and Museum.
Soest, a Jackson Hole resident for 72 years, died peacefully Oct. 12 at home with family by his side. He was 96.
As a young child Hugh attended The Rectory School, where he enjoyed choir and learned to play the organ. Grades 6 through 12 he attended Choate School and there began a love for ice hockey, which he played well into adulthood. Claiming he was a “child left behind” (with a distaste for traditional classroom methods), he completed remedial courses at Salisbury, the University of Connecticut and was ultimately admitted to the University of Wyoming in 1943.
Denied military service for medical reasons, Hugh contributed to the wartime cause by hunting rabbits, whose fur was used on military clothing. He also built and manned the Fletcher Park and Britania forest fire lookouts near Laramie. He obtained a pilot license while at UW, but when confronted with the choice of flying or his future wife, he chose Donna.
As an advocate for agriculture, Hugh was involved in the Teton County Planning Commission, Wyoming State Farm Bureau and Ducks Unlimited. He kept his exploration interests active via Rotary International, various student exchange programs and as a board member of Up With People.
He is survived by his wife of 72 years, Donna Marie Soest; daughters Susan Johansson and Sandra Rodeck (Larry); grandchildren Christine Johansson, Malcolm (Amy), Cameron, and Sara Rodeck; and great-grandchildren, Noah and Olivia Rodeck.
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Tourists come to admire its half-timbered houses, unique churches
Just a five-minute drive beyond those walls lies a fenced-off area of grey buildings standing in a row
The complex's forlornness belies the many historic stories it
A small plaque on one building identifies it as a barrack for Belgian troops over four decades of the Cold War. There is no similar sign indicating the historic gem nestled in this same building's eaves: A chapel with floor-to-ceiling murals painted by French officers held as prisoners of war (POW) during WWII. Today, the chapel is a protected memorial to victims of the Nazi regime
the city of Soest has wound its way towards installing a permanent museum next to the chapel
Yet there has been strong disagreement over what exactly the museum should commemorate
as the story of the French prisoners is only one of a series of important events that took place on the site and make up the 20th-century history of the city
A key player in the museum push has been a non-profit historical organization known as the Geschichtswerkstatt Französische Kapelle (GFK)
the GFK's volunteers have cared for the chapel
researched the lives of French officer POWs and other camp inhabitants
who include descendants of people held there
who was invited by the GFK to visit: Both his father and his uncle had been imprisoned in the Soest complex during WWII when it was a POW camp for officers
"My father practically never talked about what he lived through in that camp," Laurent told DW
"Our visit in May to the officer camp was particularly moving for us because we learned many things we were totally ignorant of," Laurent said
He was also touched by the GFK's fraternity and how it cared for the memory of the French prisoners
The experiences of French officer POWs are not widely known
many prisoners rarely spoke of their experiences after returning home
and WWII researchers have focused on other victims who suffered greater persecution under the Nazis.
Read more: Belarus: An unknown story of the Holocaust brings forgotten camp 'back into Europe's conscience'
The GFK initially wanted a museum to focus exclusively on the chapel and French prisoners
It would have been the first of its kind in Germany
but the city decided it wanted to incorporate the history of the Belgian troops stationed there during the Cold War
a leading expert at a regional museum agency
was asked by the city of Soest to provide recommendations
She ended up suggesting a far more holistic approach
Focusing on the French prisoners alone would have been insufficient
"All the layers of a historical place over time must be represented," she said
Doing otherwise would be "like going into the cellar and shining a flashlight at just one element
Then you have not experienced the whole room," Gilhaus added
"For me it was about illuminating the whole room."
the whole room — the former military complex the chapel building is part of — has many elements: The barracks were built in 1938 under the Nazis
many Soviet officer POWs were also imprisoned there alongside French
Yet unlike their western European counterparts
the Soviets were not protected by the 1929 Geneva Convention on war prisoners
They faced much worse conditions including abuse
the former barracks first housed some of the 300,000 freed forced laborers that had passed through Soest
followed by ethnic Germans driven out of Central and Eastern Europe and into Germany
Read more: Berlin Airlift 70 years on: When occupiers became protectors
Gilhaus said the museum must start with the complex's construction and trace its use and inhabitants all the way up through the present
all the while connecting what happened within the fenced-in site to the history of the city beyond it
Explaining the different experiences of French and Soviet officers is particularly important
as the latter's degrading treatment must be understood as a hallmark of Nazi racial ideology
"Some are worthy of receiving better treatment
and others are located all the way at the bottom of an ethnic hierarchy and received minimal support in all of life's essentials
Such an ethnic hierarchy is a typical characteristic of Nazism," she explained.
Gilhaus' proposals caused the GFK chairperson pushing for a narrow focus to step down
all groups of victims must be included," he told DW
Liedmann believes the French prisoners and chapel are highlighted sufficiently thanks to the museum's installation next door
we leave this place and create new connections" whether with foreign visitors or even the city of Soest
Liedmann believes forging a connection to Soest is crucial, as the city lacks awareness of its 20th-century history. "We have very little culture of remembrance in the city," he said
A museum would be in part "about making the people of Soest aware that this [the complex] is part of their history
and not some phantom that randomly landed in Soest."
Gilhaus agrees it's important for the city to have a space that lays out its history during the Nazi era
"We have a whole series of museums in Soest," she said
But "there is no place today that really deals with the history of National Socialism."
Read more: Holocaust remembrance in Germany: A changing culture
Connecting people through educational exchange
will not be possible without external funding
the chairperson of the city government's committee for culture
Her committee will decide on the future museum's size and breadth on September 12
A bigger space is essential for the broader approach but will take more resources
The GFK is anxiously awaiting the decision
Liedmann has made it clear that the GFK will only stay onboard if the resulting proposal provides enough space and resources for the group to carry on with the face-to-face educational work it has been doing for years with chapel visitors
Communicating history "can only be done in exchange with people" and especially young people
adding that such work contributes to living together peacefully
"And what could Europe possibly need more?"
How to Fix Asymmetrical Chromatography Peaks
a developer of advanced low-flow fluid delivery technologies for drug discovery and other life science applications
announces that Remco van Soest has been promoted to the Vice President level within the company
continuing to be responsible for Product Management
He has been with the company for nearly three years
first as product manager and then as director of product management for the NanoLC™ product line for proteomics research and biomarker discovery
van Soest will oversee management and strategy development for all three Eksigent product lines
including the ExpressLC™ systems and the soon-to-be-released chip-based platform
“van Soest’s ability to understand customer’s needs in the proteomics arena has allowed Eksigent to continue our leadership in the drug discovery and biomarker sector with the NanoLC product line,” commented Bill Masuda
“We will now leverage his exceptional product vision to our entire product line
His close relationships with industry-leading researchers will allow Eksigent to continue to provide users with leading-edge technology.”
Waters Corporation has successfully acquired Wyatt Technology
a leading provider of light scattering and field-flow fractionation instruments
Reference standards are essential in chromatography to ensure the correct identification of the sample components and the accuracy of quantitation
Techcomp Instruments Ltd (TIL) is pleased to announce that on 15 April
2021 Scion Instruments NL BV agreed to acquire the Teledyne Tekmar Headspace product..
The world is changing as regulators and governments wake up to threat of climate change
Alternative energy sources are constantly being appraised as fossil-..
SETAC Europe
MSB 2025
ChemUK 2025
ASMS Conference
HPLC 2025
Jeremy Levine '07 and Landon Van Soest '04 began working together while students at Ithaca College
Now they are an Emmy Award-winning documentary film-making team
a feature documentary about how efforts to aid Africa may be undermining the very communities they aim to serve
The film was awarded the Witness Award for Human Rights at its 2009 Silverdocs Film Festival premiere
the 2011 Overseas Press Club Carl Spielvogel Award for international reporting
and a 2007 fellowship from the Sundance Institute
Good Fortune was broadcast on the award-winning PBS series
a feature documentary about vigilantes along the U.S.–Mexico border was screened in dozens of film festivals around the world
and recognized with several awards for production
They have also produced for several on-going television series
including PBS NewsHour and Doomsday Preppers
the highest rated series ever on National Geographic Channel
Jeremy and Landon co-founded the Brooklyn Filmmakers Collective
a non-profit organization that provides support to independent filmmakers in New York City
They launched Transient Pictures in the fall of 2005
which has produced a wide range of original content for broadcasters
and arts organizations such as the Sundance Channel
“One of the most amazing things about my time at Ithaca College was how quickly I was out in the world making films
I found myself in the Sonora desert filming the nascent vigilante movement along the U.S.–Mexico border
which went on to screen at film festivals and on television around the world.”
“There is something really incredible about this process—of being trusted to start creating really early on
Combined with robust theory classes and a really supportive and nurturing faculty
Metrics details
Reactive Oxygen Species (ROS) derived from mitochondrial respiration are frequently cited as a major source of chromosomal DNA mutations that contribute to cancer development and aging
experimental evidence showing that ROS released by mitochondria can directly damage nuclear DNA is largely lacking
we investigated the effects of H2O2 released by mitochondria or produced at the nucleosomes using a titratable chemogenetic approach
This enabled us to precisely investigate to what extent DNA damage occurs downstream of near- and supraphysiological amounts of localized H2O2
Nuclear H2O2 gives rise to DNA damage and mutations and a subsequent p53 dependent cell cycle arrest
Mitochondrial H2O2 release shows none of these effects
even at levels that are orders of magnitude higher than what mitochondria normally produce
We conclude that H2O2 released from mitochondria is unlikely to directly damage nuclear genomic DNA
limiting its contribution to oncogenic transformation and aging
but it is not clear how this compares to physiological continuous production of O2•− and subsequently H2O2 by mitochondria in terms of achieved intracellular steady state and peak concentrations
it remains unclear to what extent respiration-derived ROS contribute directly to DNA damage
We set out to systematically analyze the localization-dependent cellular response to H2O2
making use of the chemogenetic production of H2O2 by ectopic expression of DAAO
we fused DAAO to the cytosol-facing end of the targeting sequence of the outer mitochondrial membrane protein TOMM20 and stably expressed it in RPE1-hTERT cells
RPE1 cells expressing nucleosome-targeted DAAO serve as a positive control for what happens when H2O2 reaches the nuclear DNA
Careful titration of a substrate for DAAO (D-Ala) enables the continuous production of near and supra-physiological H2O2 levels
we show that H2O2 released from mitochondria does not induce direct damage to the nuclear DNA when produced at levels likely achievable by respiration
H2O2 generation in close proximity to the nuclear DNA causes DNA mutations
DNA strand breaks and subsequent activation of the DNA damage response (DDR)
resulting in cell cycle arrest with many characteristics of senescence
similar to what has been described for cells exposed to high levels of exogenous ROS
Based on these observations we conclude that mitochondrial respiration-derived ROS is probably not a major factor in the induction of chromosomal DNA mutations
in both non-transformed human RPE1-hTERT and MCF7 breast cancer cells
a Schematic representation of RPE1hTERT cells expressing H2B-mScarlet-I-DAAO or TOM20-mScarlet-I-DAAO (further described as DAAOH2B and DAAOTOM20 respectively) to induce localized intracellular H2O2 production
b Immunofluorescence confocal images of cell lines depicted in (a) showing colocalization of DAAOH2B and DNA (Hoechst)
and colocalization of DAAOTOM20 with mitochondria (MitoTracker Green)
c Oxygen consumption rate (OCR) upon D-Ala administration as a readout for enzymatic activity of ectopically expressed DAAO
The increase in OCR shows that both lines produce roughly similar amounts of H2O2 upon addition of D-Ala
Data is normalized to OCR upon Oligomycin addition
at the last timepoint before D-Ala injection
The experiment was repeated at least 3 times with reproducible results
Data are presented as mean values of 5 separate wells -/+ SD
Source data are provided in the Source Data file
d Normalized OCR values from (c) plotted versus [D-ala]
The 6th timepoint after each D-ala injection was used for this graph
mitochondrial H2O2 release as modeled by the RPE1-hTERT-DAAOTOM20 cell line is already far above what can be expected to occur under (patho)physiological conditions
peroxiredoxins can become inactivated through overoxidation (PRDXSO2/3)
no overoxidation of PRDX was detected upon DAAO activation at viable levels
This indicates that continues intracellular H2O2 production evokes a very different cellular redox response as compared to a bolus of exogenous H2O2
a–d H2O2 measurements using the ratiometric HyPer7 sensor to determine diffusion of DAAO-produced H2O2
Treating cells with 400 µM H2O2 was used to determine 100% HyPer7 oxidation
becomes already oxidized at the lowest concentration (2.5 mM) of D-Ala used in RPE1-hTERT-DAAOTOM20 cells (a) In contrast
nuclear NLS-HyPer7 oxidation is not detected at 0–10 mM D-Ala
b NLS-HyPer7 oxidation is already detected at 2.5 mM D-Ala in RPE1-hTERT-DAAOH2B cells (c) while NES-HyPer7 oxidation is only detected starting from 10 mM D-Ala
d Data are presented as mean values -/+ SD
e Representative image of crystal violet staining of DAAO lines after 24 h L/D-Ala treatment
Both RPE1-hTERT-DAAOH2B and RPE1-hTERT-DAAOTOM20 cells do not survive continuous D-Ala treatments above 10 mM D-Ala
b Western Blot for components of the DNA damage response in WT or p53KO RPE1-hTERT-DAAOH2B and RPE1-hTERT-DAAOTOM20 cells treated for 48 h with L- or D-alanine
Treatment for 15 min with 200 µM of exogenous H2O2 serves as a positive control
Nuclear H2O2 production results in activation of the DNA damage response (pCHK1
γH2AX) and subsequent p53 stabilization and p21 expression
H2O2 produced at mitochondria by DAAOTOM20 does not activate the DNA damage response
Similar results have been obtained at least 3 times
d Representative images and quantification of alkaline comet assay using RPE1-hTERT-DAAOH2B and RPE1-hTERT-DAAOTOM20 cells treated for 2 h with L- or D-Ala
H2O2 production at mitochondria does not result in DNA strand breaks at levels that are compatible with survival (≤10 mM D-Ala)
H2O2 production close to the DNA does give rise to DNA strand breaks
Quantifications display all data points of 3 biological repeats along with the median
For each repeat ~50 comets/condition were analyzed
One-way Anova analysis + Dunnett’s multiple comparisons test is performed on the mean of the 3 biological repeats per cell line
e Schematic of workflow for obtaining DNA samples for whole genome sequencing (WGS) and mutational analysis
f Quantification of the amount of single base substitutions (SBS) acquired in RPE1-hTERT-DAAOH2B;p53KO and RPE1-hTERT-DAAOTOM20;p53KO cells treated with L-Ala or D-Ala
n = 3 biological independent clones per condtion; Data are presented as mean values -/+ SD
A one-sided unpaired t-test was performed for each line to compare L-Ala to D-Ala treatment (*p ≤ 0.05
high levels of H2O2 production at the outer mitochondrial membrane does not induce DNA damage as measured by activation of the DDR
even in the absence of the nuclear envelope and when mitochondria are closer to the DNA
indicating that also in this breast cancer cell line mitochondrial H2O2 release does not directly induce nuclear DNA damage
these experiments show that H2O2 produced in close proximity to the DNA can indeed trigger widespread DNA damage and mutation
DNA breaks and a subsequent DNA damage response
H2O2 released by mitochondria is not able to induce these phenotypes at levels compatible with cellular survival
a Quantification of the BrdU-incorporation assay
N = 3 biological replicates ~10.000 cells were analyzed per condition in each replicate
Statistical significance was determined by one-way ANOVA + Dunnett’s multiple comparison test (****p ≤ 0.0001)
but comparing RPE1-hTERT-DAAOH2B WT cells with p53KO cells
N = 3 biological replicates ~ 10.000 cells were analyzed in each replicate per condition
c Cell cycle profile analysis of RPE1-hTERT-DAAOH2B and RPE1-hTERT-DAAOTOM20 cells
showing the percentage of cells with a 4 N DNA content
Dots represent 3 biological replicates ~ 10.000 cells were analyzed in each replicate per condition
Statistical significance was determined by one-way ANOVA test + Dunnett’s multiple comparison test (**p ≤ 0.01
e Quantification of cell cycle profiles of nocodazole treated cells
Statistical significance was determined by one-way ANOVA test + Dunnett’s multiple comparison test (*p ≤ 0.05
f Quantification of cell cycle profiles of RPE1-hTERT-DAAOH2B cells containing the FUCCI cell cycle indicator
Dots represent 3 biological replicates ~ 10.000 cells were analyzed per replicate per condition
Statistical significance was determined by a two-sided unpaired t-test (**p ≤ 0.01
a Senescence associated-β-galactosidase staining of RPE1-hTERT-DAAOH2B and RPE1-hTERT-DAAOTOM20 cells
RPE1-hTERT-DAAOH2B cells become positive for the staining upon D-Ala addition
Scale bar = 100 µm b Quantification of Senescence Associated-β-galactosidase positivity of RPE1-hTERT-DAAOH2B cells treated wilt L-Ala or D-Ala
Statistical significance was determined by a two-sided unpaired t-test (***p ≤ 0.001
c Representative images of DAPI staining of RPE1-hTERT-DAAOH2B and RPE1-hTERT-DAAOTOM20 cells
RPE1-hTERT-DAAOH2B cells undergo chromatin restructuring in response to D-Ala treatment
demonstrating a phenotype reminiscent of senescence-associated heterochromatin foci (SAHFs)
Phenotype has been observed in at least three independent experiments
d Quantification of IF stainings for senescence markers p21 and Lamin B1
Induction of nuclear H2O2 results in a decrease in nuclear Lamin B1 signal and an increase in nuclear p21 intensity
For each repeat 3 images were analyzed per condition
Two-way Anova + Dunett’s multiple comparison test was performed (**p ≤ 0.01
e Quantification of cell viability by PI exclusion in RPE1-hTERT-DAAOTOM20 cells
which can be rescued by ferroptosis inhibitors ferrostatin and liproxstatin
Two-way Anova + Dunett’s multiple comparison test was performed (****p ≤ 0.0001)
These observations indicate that H2O2 produced by DAAO at the cytosolic side of the mitochondrial membrane does not lead to loss of mitochondrial function
and is therefore likely not the cause for the loss of viability upon higher levels of D-Ala
Continuous elevated H2O2 levels in close vicinity to the DNA can cause mutations and DNA strand breaks
resulting in activation of the DNA damage response and ultimately senescence
H2O2 released from mitochondria is not able to reach the nuclear DNA at physiological levels
and therefore does not directly contribute to nuclear DNA damage
Artificially high levels of mitochondrial H2O2 release
but in parallel causes oxidative cell death
ROS derived from oxygen-dependent mitochondrial ATP production is frequently implicated in cancer initiation
Unambiguous evidence for the direct oxidation and subsequent mutation of chromosomal DNA by ROS released from mitochondria at (patho) physiological levels
these levels of mitochondrial H2O2 production and release are unlikely to occur as a result of mitochondrial respiration and cells are not able to maintain redox homeostasis and die
which suggests that these two scenarios do not occur to a significant extent in our model system
These and other observations indicate that more work is needed to fully grasp the role of mitochondrial ROS in chromosomal DNA stability
all of which could influence factors like H2O2 diffusion range and sensitivity to ferroptosis
It is therefore not unthinkable that mitochondria-derived ROS does play a role in the further accumulation of mutations during later stages of tumor progression
Our work indicates that ROS-inducing agents might indeed be useful to induce cell cycle arrest and senescence
but that these should be targeted to the nucleus and will likely only be effective in p53-expressing tumors
Oxidative cell death induced by cytosolic H2O2
like in RPE1-hTERT-DAAOTOM20 cells seems to occur largely independent of p53
The source of the oxidant that induces these culture-induced mutations remains elusive
but our data presented here suggest that mitochondrial respiration is not directly involved
our sequencing data show that several hundred background single base substitutions can be found when comparing cells at the start of control L-Ala treatment with monoclonals grown out after 4 rounds of treatment and 3 days recovery
which may be due to oxidative modification
It may be interesting to see whether and how this number changes in case the experiment would be carried out under a more physiological oxygen tension
but since DAAO activity also depends on oxygen it will be difficult to estimate the rates of H2O2 production under these circumstances
An open question remains at what subcellular localization these signaling events are initiated
whether the localization determines the downstream phenotype and how these signals are transduced
Senescence markers have been shown to be heterogeneous and may depend on the type of senescence induction
Further studies are needed to better classify whether the observed arrest in our RPE1-hTERT-DAAOH2B cells also display other markers like a Senescence Associated Secretory Phenotype
The oxidative cell death induced in our RPE1-hTERT-DAAOTOM20 could be partially blocked by ferroptosis inhibitors Ferrostatin and Liproxstatin
We therefore think it is premature to state the observed oxidative cell death at artificially high mitochondrial H2O2 release is truly and only ferroptotic cell death
We do think that our model system may be useful in future work for elucidating mechanisms of ferroptosis
It can be used to study from what subcellular localizations ferroptosis can be triggered
whether there is involvement of mitochondrial lipid peroxidation and whether cells survive or die by other means when ferroptosis is blocked
our work shows that not only the dose but also the localization and kinetics of H2O2 production can have dramatic effects on phenotypic outcome
the release of ROS by mitochondria is not a significant source of direct DNA oxidation or indirect DNA mutation and is therefore not likely to contribute considerably to cancer initiation and aging
Retinal pigment epithelial cells (hTERT RPE-1
CRL-4000) were cultured in DMEM-F12 (Gibco
100 Units Penicillin-Streptomycin (Sigma-Aldrich
P4458) and 2 mM L-glutamine (Sigma-Aldrich
Cells were grown at 37 °C under 6% CO2 atmosphere
HTB-22) and human epithelial kidney cells (HEK293T
CRL-3216) used for lentiviral production were cultured in DMEM-High Glucose (Sigma-Aldrich
100 Units Penicllin-Streptomycin and 2 mM L-glutamine
A7377) were dissolved in PBS to create a 1 M stock solution and frozen in aliquots
Addition of alanine was always precluded by refreshing medium
For trapping cells in mitosis to visualize a G0/1 arrest
SML0583) or 200 nM Liproxstatin (Sigma-Aldrich
SML1414) was added 1 h before treatment with L- or D-Ala
Cells were selected using 5 µg/ml puromycin (Santa Cruz) and 10 µg/ml blasticidin (InvivoGen) for RPE1-hTERT cells and 1 µg/ml puromycin for MCF7 cells
Single cells were expanded in 96-wells plates and expanded to create monoclonal lines
only cells with high HyPer7 expression were sorted
PCR reactions for cloning were performed using Q5 High-Fidelity DNA polymerase (New England Biolabs
DNA and plasmid isolation from agarose gels
PCR reactions or bacterial culture was performed using QIAquick Gel extraction kit (Qiagen
27104) and Maxiprep purification kit (LabNed
RPE1-hTERT-DAAOH2B and RPE1-hTERT-DAAOTOM20 were transiently transfected using PEI Max
After 48 h GFP positive cells were single-cell sorted into a 96-wells plate using FACS
P53 knock-out status of clones was confirmed by testing sensitivity for the p53 stabilizer Nutlin-3a (10 µM for 16 h
10004372) and Western Blotting for p53 protein
Antibodies used for Western Blot were pChk1 S345 (Cell Signaling
lot# 1000538-1 1:2000) and PRDXSO2/3 (Abcam
Antibodies used for IF were LaminB1 (Merck
Mean signal of segmented nuclei was measured
a mito stress test was performed by injection of 2 µM oligomycin A (Cayman Chemical
A8674) and 1 µM Rotenone (Sigma-Aldrich R8875)
cells in each well were lysed using a buffer containing 0.1% TX-100
Plates were put at −20 °C overnight and protein concentration per well was measured by use of a Pierce BCA protein assay kit (Thermo Fischer Scientific
23225) according to manufacturer’s protocol
OCR measurements were then normalized by protein concentration
Red-Green Color-blind friendly false color images were prepared with mKO2-CDT1 (G0/1) displayed in Blue and mAG1-Geminin (S/G2) in Green
Protein lysates were obtained by washing cells with PBS and subsequently lysed and scraped in 1 x Sample Buffer (2% SDS
Samples were first heated for 5 min at 95 °C before running SDS-PAGE and standard blotting procedure
Samples for γH2AX staining were transferred onto a 0.2 µm nitrocellulose membrane (Cytvia Amersham
whereas other samples were transferred onto a polyvinylidene difluoride (PVDF) membrane (Millipore
Samples were blocked for 1 h at 4 °C in TBS-Tween (1% v/v)
Primary antibody incubation was performed overnight or over weekend at 4 °C in TBS-T containing 1% BSA
Secondary antibodies conjugated with horseradish peroxidase (1:10000
610-4302 and 611-1302) in TBS-Tween were incubated for 1 h at room temperature
Blots were imaged on Image Quant LAS (GE HealthCare) with ImageQuant LAS4000 (version 1.3) software
Contrast was adjusted for clarity by linear image processing in Adobe Photoshop if required
Uncropped Western Blots can be found in the associated Source Data file
cells were incubated for 5 min with prewarmed PBS containing 12.5 mg/ml N-Ethylmaleimide (Millipore
E3876) at 37 °C prior to harvesting to fix the cysteine-dependent redox state
Cells were then washed with PBS and lysed in buffer containing 50 mM Tris-HCl (pH 7.5)
L2023) and 100 mM iodoacetamide (Sigma-Aldrich
Samples were divided over two tubes and a reducing and a non-reducing sample was prepared by adding SDS sample buffer with or without beta-mercaptoethanol
Cell viability was assessed by propidium iodide (PI
Medium and subsequent wash with PBS were collected
resuspended and added to the collected medium + PBS (15 ml falcon tubes)
Beckmann tabletop centrifuge with swingout rotor) and put on ice
and the pelleted cells were resuspended in PBS containing 20 µg/ml propium iodide
After centrifugation and removal of supernatant
200 µl of PBS was added to prevent cells from aggregating during fixation
Cells were fixed by dropwise addition of 5 ml of ice-cold 70% ethanol while vortexing
Samples were left at 4 °C overnight or longer
fixed cells were pelleted (1500 RPM 5 min) and resuspended in PBS containing 20 µg/ml propium iodide and 200 µg/ml RNase (Millipore
In case of cells containing the FUCCI cell cycle reporter
pelleted (1200 RPM 5 min) and resuspended in PBS containing 2 µg/ml of the membrane permeable DNA staining Hoechst33342 (Thermo Fischer Scientific
cells were cultured in medium containing 10 µM BrdU (Invitrogen
Cells were collected and fixed in the same fashion as for DNA profiles
after centrifugation the cell pellet was resuspended in 0.1 M HCL solution containing 0.5 mg/ml of the protease pepsin (Sigma-Aldrich
1.07185) and left for 20 min at room temperature
After washing with PBS containing 0.1% BSA and 0.5% Tween
cells were pelleted and treated with 2 M HCL for 12 min at 37 °C to denature the DNA
after which double the volume of borate buffer (0.1 M H3BO3 set at pH 8.5 with 0.1 M Na2B4O7) was added
the cell pellet was incubated with PBS/BSA/Tween containing BrdU-FITC antibody (1:50
Cells were centrifuged and resuspended in PBS/BSA containing 20 µg/ml PI and 200 µg/ml RNase
To analyze mitochondrial membrane potential
cells were cultured in medium containing 100 nM tetramethylrodamine (TMRM
pelleted (1200 RPM 5 min) and resuspended in 250 µl PBS
As a positive control for loss of membrane potential
10 µM FCCP was added for at least 15 min prior to measuring
Flow cytometry samples were measured on BD FACSCelesta Cell analyzer (BD biosciences) and analyzed with BD FacsDiva Software. Gating strategies for Flow Cytometry experiments can be found in Supplementary Fig. 16
cells were treated with L- or D-Alanine for 24 h
Cells were then washed 2x with PBS to remove dead cells and fixed with ice-cold (−20 °C) methanol for 5 min
Methanol was removed and replaced with crystal violet solution (0.5% w/v in 25% v/v methanol in H2O) for 10 min
Crystal violet was removed and wells are washed with water and airdried overnight
Plates were imaged on an Epson 3170 photo scanner (Epson)
the staining was dissolved in 10% acetic acid for 30 min
transferred to a 96-wells plate and measured in a plate reader by spectrophotometry at 595 nm
Cells were treated for 2 days with L- or D-Ala and kept in culture for an additional 5 days
Then cells were washed twice with PBS and fixed for 15 min with 3.7% formalin (37% formalin diluted in PBS (SAFC
Cells were washed twice with PBS and then incubated with freshly made SA-β-gal buffer containing 25 mM Na2HPO4 (Supelco
5 mM Potassium Ferricyanide (Sigma-Aldrich
5 mM Potassium Ferrocyanide (Sigma-Aldrich
60279) and 0.1% X-Gal (from a 4% X-Gal solution in dimethylformamide
Buffer was passed through a 0.2 micron syringe filter before X-Gal was added
Cells were rinsed 2x with PBS and kept under 70% ethanol
Cells were imaged on an EVOS-M5000 microscope (Thermo Fisher)
Alkaline comet assays were performed using the CometAssay single cell gel electrophoresis assay kit (R&D systems 4250-050-K) according to the manufacturers’ instructions
Cells were treated for 2 h with L/D-Ala after which they were trypsinized and counted to make 105 cells/ml dilutions
DNA alkaline unwinding for 1 h and electrophoresis for 1 h
DNA staining was performed using Midori Green Advance for 30 min at room temperature (1:5000 in TE buffer pH7.5
Comets were imaged on an EVOS-M5000 microscope (Thermo Fisher) using the GFP filter block
Comets were manually analyzed using FIJI (version 2.9.0/1.54b) software
Samples were blinded before quantification
The area and mean pixel value of the comet head and the complete comet were measured
the mean pixel value of the local background was determined
The mean pixel value of the background was subtracted of the mean pixel value of the comet head and total comet
These values were then multiplied with the area of the comet head or total comet to create the integrated densities corrected for background signal
The integrated density of the comet tail value was calculated by subtracting the integrated density of the comet head from the integrated density of the total comet
The percentage DNA in comet tail was then calculated by dividing the integrated density of the comet tail value with the integrated density of the total comet
Statististical analyses were performed using GraphPad Prism (version 10.1.1)
type of test and exact p-values are indicated in the figure legends
All findings reported here were observed multiple independent times
No statistical methods were used to predetermine sample size
For experiments where analysis was performed manually (e.g
investigators were blinded to the sample’s treatment conditions during analysis
Further information on research design is available in the Nature Portfolio Reporting Summary linked to this article
Iron ion-dependent modification of bases in DNA by the superoxide radical-generating system hypoxanthine/xanthine oxidase
Damage to the bases in DNA induced by hydrogen peroxide and ferric ion chelates
Base excision repair pathways of bacteria: new promise for an old problem
Base excision repair in archaea: back to the future in DNA repair
Reactive oxygen species (ROS) as pleiotropic physiological signalling agents
Oxidatively generated base damage to cellular DNA
How mitochondria produce reactive oxygen species
Repair and genetic consequences of endogenous DNA base damage in mammalian cells
Deconvoluting the context-dependent role for autophagy in cancer
Mitochondria and mitochondrial ROS in cancer: novel targets for anticancer therapy
Spatial and temporal control of mitochondrial H2 O2 release in intact human cells
Ultrasensitive genetically encoded indicator for hydrogen peroxide identifies roles for the oxidant in cell migration and mitochondrial function
Monitoring cytosolic H(2)O(2) fluctuations arising from altered plasma membrane gradients or from mitochondrial activity
Khan, K. et al. Mitochondria-derived reactive oxygen species are the likely primary trigger of mitochondrial retrograde signaling in arabidopsis. Curr. Biol. https://doi.org/10.1016/j.cub.2023.12.005 (2024)
Mitochondrial reactive oxygen species are scavenged by cockayne syndrome B protein in human fibroblasts without nuclear DNA damage
The mutational impact of culturing human pluripotent and adult stem cells
DNA damage and oxidant stress activate p53 through differential upstream signaling pathways
A ROS-dependent mechanism promotes CDK2 phosphorylation to drive progression through S phase
Chemogenetic generation of hydrogen peroxide in the heart induces severe cardiac dysfunction
Oxygen-consumption based quantification of chemogenetic H(2)O(2) production in live human cells
Complexities of the chemogenetic toolkit: differential mDAAO activation by d-amino substrates and subcellular targeting
The sites and topology of mitochondrial superoxide production
Diffusion and transport of reactive species across cell membranes
Superoxide dismutases: dual roles in controlling ROS damage and regulating ROS signaling
Redox regulation of RAD51 Cys319 and homologous recombination by peroxiredoxin 1
Towards a comprehensive view of 8-oxo-7,8-dihydro-2’-deoxyguanosine: highlighting the intertwined roles of DNA damage and epigenetics in genomic instability
The molecular architecture of cell cycle arrest
Persistent repair intermediates induce senescence
Hypersensitivity to DNA damage in antephase as a safeguard for genome stability
A FOXO-dependent replication checkpoint restricts proliferation of damaged cells
Visualizing spatiotemporal dynamics of multicellular cell-cycle progression
Rb-mediated heterochromatin formation and silencing of E2F target genes during cellular senescence
Signatures of mutational processes in human cancer
Intracellular localization of 8-oxo-dGTPase in human cells
with special reference to the role of the enzyme in mitochondria
Mechanisms of mitochondrial iron-sulfur protein biogenesis
Micromolar intracellular hydrogen peroxide disrupts metabolism by damaging iron-sulfur enzymes
Eukaryotic DNA polymerases require an iron-sulfur cluster for the formation of active complexes
Emerging critical roles of Fe-S clusters in DNA replication and repair
Author correction: age-associated mitochondrial DNA mutations cause metabolic remodeling that contributes to accelerated intestinal tumorigenesis
All-optical spatiotemporal mapping of ROS dynamics across mitochondrial microdomains in situ
Chemoptogenetic damage to mitochondria causes rapid telomere dysfunction
The role of ROS in tumour development and progression
The hallmarks of cancer from a redox perspective
ROS homeostasis and metabolism: a dangerous liason in cancer cells
Metabolic interactions in the tumor microenvironment
Redox-directed cancer therapeutics: molecular mechanisms and opportunities
LSD1: more than demethylation of histone lysine residues
DNA oxidation as triggered by H3K9me2 demethylation drives estrogen-induced gene expression
Functional analysis of Nox4 reveals unique characteristics compared to other NADPH oxidases
Oxidation of F-actin controls the terminal steps of cytokinesis
Mutations induced by 8-hydroxyguanine (8-oxo-7,8-dihydroguanine)
Mutagenic effects of 8-hydroxy-dGTP in live mammalian cells
The repertoire of mutational signatures in human cancer
Real-time monitoring of basal H2O2 levels with peroxiredoxin-based probes
Redox-dependent control of FOXO/DAF-16 by transportin-1
Oxidative stress-induced premature senescence dysregulates VEGF and CFH expression in retinal pigment epithelial cells: implications for age-related macular degeneration
Genome engineering using the CRISPR-Cas9 system
Fiji: an open-source platform for biological-image analysis
Medical image computing and computer assisted intervention
Comprehensive single-cell genome analysis at nucleotide resolution using the PTA analysis toolbox
Download references
We would like to thank Tessa Vreeman for help with constructing lentiviral HyPer7 plasmids
Peter de Keizer for advice on the assessment of senescence
Livio Kleij for technical support with the HyPer7 measurements
Fried Zwartkruis for critical reading of the manuscript and our colleagues at the Center for Molecular Medicine
UMC Utrecht for their valuable input and suggestions
This work was funded by grants from the Dutch Cancer Society (KWF Kankerbestrijding
KWF grant 14798 and KWF UU2014-6902) to TBD
BMTB and RvB are members of the Oncode Institute
which is partly funded by the Dutch Cancer Society (KWF Kankerbestrijding)
Princess Máxima Center for Pediatric Oncology
performed bioinformatic mutational analysis
validated and analysed the monoclonal MCF7-DAAO lines
made the CRISPR/Cas9 knockout p53 KO lines
designed and supervised the sequencing and mutation analysis experiments
provided critical input on the experimental setup and the manuscript
Nature Communications thanks Nadja de Souza-Pinto and the other
reviewer(s) for their contribution to the peer review of this work
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DOI: https://doi.org/10.1038/s41467-024-47008-x
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