Weirdly and even though most of this paper’s authors are from the University of British Columbia (UBC; Canada), only one press release was issued and that was by the lead author’s (Gesa Busch) home institution, the University of Göttingen (Germany).
I’m glad Busch, the other authors, and the work are getting some attention (if not as much as I think they should).
A research team from the University of Göttingen and the University of British Columbia (Canada) has investigated how people in five different countries react to various usages of genome editing in agriculture. The researchers looked at which uses are accepted and how the risks and benefits of the new breeding technologies are rated by people. The results show only minor differences between the countries studied – Germany, Italy, Canada, Austria and the USA. In all countries, making changes to the genome is more likely to be deemed acceptable when used in crops rather than in livestock. The study was published in Agriculture and Human Values.
Relatively new breeding technologies, such as CRISPR [clustered regularly interspaced short palindromic repeats) gene editing, have enabled a range of new opportunities for plant and animal breeding. In the EU, the technology falls under genetic engineering legislation and is therefore subject to rigorous restrictions. However, the use of gene technologies remains controversial. Between June and November 2019, the research team collected views on this topic via online surveys from around 3,700 people from five countries. Five different applications of gene editing were evaluated: three relate to disease resistance in people, plants, or animals; and two relate to achieving either better quality of produce or a larger quantity of product from cattle.
“We were able to observe that the purpose of the gene modification plays a major role in how it is rated,” says first author Dr Gesa Busch from the University of Göttingen. “If the technology is used to make animals resistant to disease, approval is greater than if the technology is used to increase the output from animals.” Overall, however, the respondents reacted very differently to the uses of the new breeding methods. Four different groups can be identified: strong supporters, supporters, neutrals, and opponents of the technology. The opponents (24 per cent) identify high risks and calls for a ban of the technology, regardless of possible benefits. The strong supporters (21 per cent) see few risks and many advantages. The supporters (26 per cent) see many advantages but also risks. Whereas those who were neutral (29 per cent) show no strong opinion on the subject.
This study was made possible through funding from the Free University of Bozen-Bolzano and Genome BC.
I have one quick comment about the methodology. It can be difficult to get a sample that breaks down along demographic lines that is close to or identical to national statistics. That said, it was striking to me that every country was under represented in the ’60 years+ ‘ category. In Canada, it was by 10 percentage points (roughly). For other countries the point spread was significantly wider. In Italy, it was a 30 percentage point spread (roughly).
I found the data in the Supplementary Materials yesterday (July 13, 2021). When I looked this morning, that information was no longer there but you will find what appears to be the questionnaire. I wonder if this removal is temporary or permanent and, if permanent, I wonder why it was removed.
Participants for the Canadian portion of the survey were supplied by Dynata, a US-based market research company. Here’s the company’s Wikipedia entry and its website.
Information about how participants were recruited was also missing this morning (July 14, 2021).
Genome British Columbia (Genome BC)
I was a little surprised when I couldn’t find any information about the program or the project on the Genome BC website as the organization is listed as a funder.
There is a ‘Genomics and Society’ tab (seems promising, eh?) on the homepage where you can find the answer to this question: What is GE³LS Research?,
GE3LS research is interdisciplinary, conducted by researchers across many disciplines within social science and humanities, including economics, environment, law, business, communications, and public policy.
There’s also a GE3LS Research in BC page titled Project Search; I had no luck there either.
It all seems a bit mysterious to me and, just in case anything else disappears off the web, here’s a July 13, 2021 news item about the research on phys.org as backup to what I have here.
I’ve been wondering when there’d be more ‘public engagement’ discussion in Canada about artificial intelligence, human genome editing, robotics, and other technologies which are rapidly changing status from ’emerging technologies’ to ’embedded technologies’.
In October of 2020, Jennifer Doudna and Emmanuelle Charpentier were awarded the Nobel Prize in chemistry for their discovery of an adaptable, easy way to edit genomes, known as CRISPR [clustered regularly interspaced short palindromic repeats], which has transformed the world of genetic engineering.
CRISPR has been used to fight lung cancer and correct the mutation responsible for sickle cell anemia in stem cells. But the technology was also used by a Chinese scientist to secretly and illegally edit the genomes of twin girls — the first-ever heritable mutation of the human germline made with genetic engineering.
“We’ve moved away from an era of science where we understood the risks that came with new technology and where decision stakes were fairly low,” says Dietram Scheufele, a professor of life sciences communication at the University of Wisconsin-Madison.
Today, Scheufele and his colleagues say, we’re in a world where new technologies have very immediate and sometimes unpredictable but significant impacts on society. In a paper published the week of April 26 [2021] in the Proceedings of the National Academy of Sciences [PNAS], the researchers argue that such advanced tech, especially CRISPR, demands more robust and thoughtful public engagement if it is to be harnessed to benefit the public without crossing ethical lines.
The authors say that being thoughtful and transparent about public engagement goals and using evidence from social science can help facilitate the difficult conversations society must have about scientific issues like CRISPR and their societal implications. Effective public engagement, in turn, lays the groundwork for public ownership of advances that do arise from CRISPR.
Life sciences communication Professor Dominique Brossard and graduate student Nicole Krause, along with University of Vienna research assistant Isabelle Freiling, co-authored the report with Scheufele. The paper stems from a 2019 National Academy of Sciences colloquium on CRISPR.
Since 2012, when the CRISPR system was first described, scientists have understood both its genetic engineering potential and the need for public engagement to discuss the possible uses of the technology. Many scientists wanted to avoid rehashing the controversies surrounding genetically modified organisms, which have been harshly criticized as unnatural and unnecessary by some activists despite broad scientific support for their use.
Yet, Krause says, some scientists who supported using CRISPR began by errantly repeating the public engagement methods employed for GMOs, which “assumes that people just need more knowledge, more of an ability to understand the science.” Instead, Krause adds: “Solutions focused on tailoring communications to people’s values would make more sense.”
This values-based public engagement strategy is supported by social science research into how people form and change their opinions around new technologies. Some public engagement methods engage value systems, and encourage thoughtful conversation, more than others.
For example, what researchers term “public involvement” and “public collaboration” are methods of two-way communication involving the joint exchange of information and values and the identification and design of science-based decisions that adhere to those values. That contrasts with “public communication,” which focuses only on the dissemination of scientific information.
Scheufele and his colleagues say that such collaborative approaches could help scientists widen the representation of voices in debates around science to groups who are often overlooked, such as people with disabilities or racial minorities.
“As the scientific community, we don’t have a long track record of effective engagement mechanism with these communities,” says Scheufele. This failure to reach broader groups stems in part from the low participation rates of most science engagement events, which also attract highly selective audiences.
Another challenge is rewarding scientists for public engagement. “There’s very little incentive in academia to do this kind of work,” says Scheufele.
A recent report by Brossard and others found that a majority of land-grant faculty felt that public engagement was very important, but believed it was less important to their colleagues. That divide suggests scientists feel their engagement efforts won’t be rewarded by their peers, says Brossard.
Now, Brossard, Krause, Scheufele and colleagues have a grant from the National Science Foundation to research how to depolarize debates around CRISPR. Previous studies suggest that making people accountable for their positions helps them think more critically about their underlying reasoning. And when social scientists emphasize the complexity inherent in people’s values, it helps people consider controversial issues with more nuance.
But engaging a diverse society with pluralistic value systems in deliberations on the latest technologies will never be easy.
“The policymaking process involves a lot more than just science. Science will inform how we regulate technologies, and so will religious, political, ethical, regulatory and economic considerations,” says Scheufele. “And so the ability to actually do engagement in this much broader setting where we meaningfully contribute and guide the debate with the best available science is a major challenge.”
Apparently the magic is in the lipid nanoparticles. A March 1, 2021 news item on Nanowerk announced research into lipid nanoparticles as a means to deliver CRISPR (clustered regularly interspaced short palindromic repeats) to specific organs (Note: A link has been removed),
The genome editing technology CRISPR has emerged as a powerful new tool that can change the way we treat disease. The challenge when altering the genetics of our cells, however, is how to do it safely, effectively, and specifically targeted to the gene, tissue and organ that needs treatment.
Scientists at Tufts University and the Broad Institute of Harvard [University] and MIT [Massachusetts Institute of Technology] have developed unique nanoparticles comprised of lipids — fat molecules — that can package and deliver gene editing machinery specifically to the liver.
In a study published in the Proceedings of the National Academy of Sciences [PNAS] (“Lipid nanoparticle-mediated codelivery of Cas9 mRNA and single-guide RNA achieves liver-specific in vivo genome editing of Angptl3”), they have shown that they can use the lipid nanoparticles (LNPs) to efficiently deliver the CRISPR machinery into the liver of mice, resulting in specific genome editing and the reduction of blood cholesterol levels by as much as 57% — a reduction that can last for at least several months with just one shot.
The problem of high cholesterol plagues more than 29 million Americans, according to the Centers for Disease Control and Prevention. The condition is complex and can originate from multiple genes as well as nutritional and lifestyle choices, so it is not easy to treat. The Tufts and Broad researchers, however, have modified one gene that could provide a protective effect against elevated cholesterol if it can be shut down by gene editing.
The gene that the researchers focused on codes for the angiopoietin-like 3 enzyme (Angptl3). That enzyme tamps down the activity of other enzymes – lipases – that help break down cholesterol. If researchers can knock out the Angptl3 gene, they can let the lipases do their work and reduce levels of cholesterol in the blood. It turns out that some lucky people have a natural mutation in their Angptl3 gene, leading to consistently low levels of triglycerides and low-density lipoprotein (LDL) cholesterol, commonly called “bad” cholesterol, in their bloodstream without any known clinical downsides.
“If we can replicate that condition by knocking out the angptl3 gene in others, we have a good chance of having a safe and long term solution to high cholesterol,” said Qiaobing Xu, associate professor of biomedical engineering at Tufts’ School of Engineering and corresponding author of the study. “We just have to make sure we deliver the gene editing package specifically to the liver so as not to create unwanted side effects.”
Xu’s team was able to do precisely that in mouse models. After a single injection of lipid nanoparticles packed with mRNA coding for CRISPR-Cas9 and a single-guide RNA targeting Angptl3, they observed a profound reduction in LDL cholesterol by as much as 57% and triglyceride levels by about 29 %, both of which remained at those lowered levels for at least 100 days. The researchers speculate that the effect may last much longer than that, perhaps limited only by the slow turnover of cells in the liver, which can occur over a period of about a year. The reduction of cholesterol and triglycerides is dose dependent, so their levels could be adjusted by injecting fewer or more LNPs in the single shot, the researchers said.
By comparison, an existing, FDA [US Food and Drug Administration]-approved version of CRISPR mRNA-loaded LNPs could only reduce LDL cholesterol by at most 15.7% and triglycerides by 16.3% when it was tested in mice, according to the researchers.
The trick to making a better LNP was in customizing the components – the molecules that come together to form bubbles around the mRNA. The LNPs are made up of long chain lipids that have a charged or polar head that is attracted to water, a carbon chain tail that points toward the middle of the bubble containing the payload, and a chemical linker between them. Also present are polyethylene glycol, and yes, even some cholesterol – which has a normal role in lipid membranes to make them less leaky – to hold their contents better.
The researchers found that the nature and relative ratio of these components appeared to have profound effects on the delivery of mRNA into the liver, so they tested LNPs with many combinations of heads, tails, linkers and ratios among all components for their ability to target liver cells. Because the in vitro potency of an LNP formulation rarely reflects its in vivo performance, they directly evaluated the delivery specificity and efficacy in mice that have a reporter gene in their cells that lights up red when genome editing occurs. Ultimately, they found a CRISPR mRNA-loaded LNP that lit up just the liver in mice, showing that it could specifically and efficiently deliver gene-editing tools into the liver to do their work.
The LNPs were built upon earlier work at Tufts, where Xu and his team developed LNPs with as much as 90% efficiency in delivering mRNA into cells. A unique feature of those nanoparticles was the presence of disulfide bonds between the long lipid chains. Outside the cells, the LNPs form a stable spherical structure that locks in their contents. When they are inside a cell, the environment within breaks the disulfide bonds to disassemble the nanoparticles. The contents are then quickly and efficiently released into the cell. By preventing loss outside the cell, the LNPs can have a much higher yield in delivering their contents.
“CRISPR is one of the most powerful therapeutic tools for the treatment of diseases with a genetic etiology. We have recently seen the first human clinical trail for CRISPR therapy enabled by LNP delivery to be administered systemically to edit genes inside the human body. Our LNP platform developed here holds great potential for clinical translation,” said Min Qiu, post-doctoral researcher in Xu’s lab at Tufts. “We envision that with this LNP platform in hand, we could now make CRISPR a practical and safe approach to treat a broad spectrum of liver diseases or disorders,” said Zachary Glass, graduate student in the Xu lab. Qiu and Glass are co-first authors of the study.
The Mutant Project is both a book (The Mutant Project: Inside the Global Race to Genetically Modify Humans) and an event about gene editing with special reference to the CRISPR (clustered regularly interspaced short palindromic repeats) twins, Lulu and Nana. The event is being held by Toronto’s ArtSci Salon. Here’s more from their March 3, 2021 announcement (received via email),
The Mutant Project
A talk and discussion with Eben Kirksey
Discussants:
Dr. Elizabeth Koester, Postdoctoral fellow, Department of History, UofT [University of Toronto]
Vincent Auffrey, PhD student, IHPST [Institute for the History and Philosophy of Science and Technology], UofT
Fan Zhang, PhD student, IHPST, UofT
This event will be streamed on Zoom and on Youtube
At a conference in Hong Kong in November 2018, Dr. He Jiankui announced that he had created the first genetically modified babies—twin girls named Lulu and Nana—sending shockwaves around the world. A year later, a Chinese court sentenced Dr. He to three years in prison for “illegal medical practice.”
As scientists elsewhere start to catch up with China’s vast genetic research program, gene editing is fueling an innovation economy that threatens to widen racial and economic inequality. Fundamental questions about science, health, and social justice are at stake: Who gets access to gene editing technologies? As countries loosen regulations around the globe, from the U.S. to Indonesia, can we shape research agendas to promote an ethical and fair society?
Join us to welcome Dr. Kirksey, who will discuss key topics from his book “The Mutant Project”.
The talk will be followed by a Q&A
EBEN KIRKSEY is an American anthropologist who finished his latest book as a Member of the Institute for Advanced Study in Princeton, New Jersey. He has been published in Wired, The Atlantic, The Guardian and The Sunday Times. He is sought out as an expert on science in society by the Associated Press, The Wall Street Journal, The New York Times, Democracy Now, Time and the BBC, among other media outlets. He speaks widely at the world’s leading academic institutions including Oxford, Yale, Columbia, UCLA, and the International Summit of Human Genome Editing, plus music festivals, art exhibits, and community events. Professor Kirksey holds a long-term position at Deakin University in Melbourne, Australia. For more information, please visit https://eben-kirksey.space/.
Elizabeth Koester currently holds a SSHRC [Social Science and Humanities Research Council of Canada] Postdoctoral Fellowship in the Department of History at the University of Toronto. After practising law for many years, she undertook graduate studies in the history of medicine at the Institute for the History and Philosophy of Science and Technology at the University of Toronto and was awarded a PhD in 2018. A book based on her dissertation, In the Public Good: Eugenics and Law in Ontario, will be published by McGill-Queen’s University Press and is anticipated for Fall 2021.
Vincent Auffrey is pursuing his PhD at the Institute for the History of Philosophy of Science and Technology (IHPST) at the University of Toronto. His focus is set primarily on the social history of medicine and the history of eugenics in Canada. Secondary interests include the histories of scientific racism and of anatomy, and the interplay between knowledge and power.
Fan Zhang is a PhD student at the History of Philosophy of Science and Technology (IHPST) at the University of Toronto
The ‘pair of scissors’ analogy is probably the most well known of the attempts to describe how the CRISPR (clustered regularly interspaced short palindromic repeats)-Cas9 gene editing system works. It seems a new analogy is about to be added according to a January 19 2021 news item on ScienceDaily (Note: This October 30, 2019 posting features more CRISPR analogies),
In a series of experiments with laboratory-cultured bacteria, Johns Hopkins scientists have found evidence that there is a second role for the widely used gene-cutting system CRISPR-Cas9 — as a genetic dimmer switch for CRISPR-Cas9 genes. Its role of dialing down or dimming CRISPR-Cas9 activity may help scientists develop new ways to genetically engineer cells for research purposes.
Here’s an image illustrating the long form of the tracrRNA or ‘dimmer switch’ alongside the more commonly used short form,
Caption: Left – a schematic of the long form of the tracrRNA used by the CRISPR-Cas9 system in bacteria; Right – the standard guide RNA used by many scientists as part of the gene-cutting CRISPR-Cas9 system. Credit: Joshua Modell, Rachael Workman and Johns Hopkins Medicine
First identified in the genome of gut bacteria in 1987, CRISPR-Cas9 is a naturally occurring but unusual group of genes with a potential for cutting DNA sequences in other types of cells that was realized 25 years later. Its value in genetic engineering — programmable gene alteration in living cells, including human cells — was rapidly appreciated, and its widespread use as a genome “editor” in thousands of laboratories worldwide was recognized in the awarding of the Nobel Prize in Chemistry last year to its American and French co-developers.
CRISPR stands for clustered, regularly interspaced short palindromic repeats. Cas9, which refers to CRISPR-associated protein 9, is the name of the enzyme that makes the DNA slice. Bacteria naturally use CRISPR-Cas9 to cut viral or other potentially harmful DNA and disable the threat, says Joshua Modell, Ph.D., assistant professor of molecular biology and genetics at the Johns Hopkins University School of Medicine. In this role, Modell says, “CRISPR is not only an immune system, it’s an adaptive immune system — one that can remember threats it has previously encountered by holding onto a short piece of their DNA, which is akin to a mug shot.” These mug shots are then copied into “guide RNAs” that tell Cas9 what to cut.
Scientists have long worked to unravel the precise steps of CRISPR-Cas9’s mechanism and how its activity in bacteria is dialed up or down. Looking for genes that ignite or inhibit the CRISPR-Cas9 gene-cutting system for the common, strep-throat causing bacterium Streptococcus pyogenes, the Johns Hopkins scientists found a clue regarding how that aspect of the system works.
Specifically, the scientists found a gene in the CRISPR-Cas9 system that, when deactivated, led to a dramatic increase in the activity of the system in bacteria. The product of this gene appeared to re-program Cas9 to act as a brake, rather than as a “scissor,” to dial down the CRISPR system.
“From an immunity perspective, bacteria need to ramp up CRISPR-Cas9 activity to identify and rid the cell of threats, but they also need to dial it down to avoid autoimmunity — when the immune system mistakenly attacks components of the bacteria themselves,” says graduate student Rachael Workman, a bacteriologist working in Modell’s laboratory.
To further nail down the particulars of the “brake,” the team’s next step was to better understand the product of the deactivated gene (tracrRNA). RNA is a genetic cousin to DNA and is vital to carrying out DNA “instructions” for making proteins. TracrRNAs belong to a unique family of RNAs that do not make proteins. Instead, they act as a kind of scaffold that allows the Cas9 enzyme to carry the guide RNA that contains the mug shot and cut matching DNA sequences in invading viruses.
TracrRNA comes in two sizes: long and short. Most of the modern gene-cutting CRISPR-Cas9 tools use the short form. However, the research team found that the deactivated gene product was the long form of tracrRNA, the function of which has been entirely unknown.
The long and short forms of tracrRNA are similar in structure and have in common the ability to bind to Cas9. The short form tracrRNA also binds to the guide RNA. However, the long form tracrRNA doesn’t need to bind to the guide RNA, because it contains a segment that mimics the guide RNA. “Essentially, long form tracrRNAs have combined the function of the short form tracrRNA and guide RNA,” says Modell.
In addition, the researchers found that while guide RNAs normally seek out viral DNA sequences, long form tracrRNAs target the CRISPR-Cas9 system itself. The long form tracrRNA tends to sit on DNA, rather than cut it. When this happens in a particular area of a gene, it prevents that gene from expressing, — or becoming functional.
To confirm this, the researchers used genetic engineering to alter the length of a certain region in long form tracrRNA to make the tracrRNA appear more like a guide RNA. They found that with the altered long form tracrRNA, Cas9 once again behaved more like a scissor.
Other experiments showed that in lab-grown bacteria with a plentiful amount of long form tracrRNA, levels of all CRISPR-related genes were very low. When the long form tracrRNA was removed from bacteria, however, expression of CRISPR-Cas9 genes increased a hundredfold.
Bacterial cells lacking the long form tracrRNA were cultured in the laboratory for three days and compared with similarly cultured cells containing the long form tracrRNA. By the end of the experiment, bacteria without the long form tracrRNA had completely died off, suggesting that long form tracrRNA normally protects cells from the sickness and death that happen when CRISPR-Cas9 activity is very high.
“We started to get the idea that the long form was repressing but not eliminating its own CRISPR-related activity,” says Workman.
To see if the long form tracrRNA could be re-programmed to repress other bacterial genes, the research team altered the long form tracrRNA’s spacer region to let it sit on a gene that produces green fluorescence. Bacteria with this mutated version of long form tracrRNA glowed less green than bacteria containing the normal long form tracrRNA, suggesting that the long form tracrRNA can be genetically engineered to dial down other bacterial genes.
Another research team, from Emory University, found that in the parasitic bacteria Francisella novicida, Cas9 behaves as a dimmer switch for a gene outside the CRISPR-Cas9 region. The CRISPR-Cas9 system in the Johns Hopkins study is more widely used by scientists as a gene-cutting tool, and the Johns Hopkins team’s findings provide evidence that the dimmer action controls the CRISPR-Cas9 system in addition to other genes.
The researchers also found the genetic components of long form tracrRNA in about 40% of the Streptococcus group of bacteria. Further study of bacterial strains that don’t have the long form tracrRNA, says Workman, will potentially reveal whether their CRISPR-Cas9 systems are intact, and other ways that bacteria may dial back the CRISPR-Cas9 system.
The dimmer capability that the experiments uncovered, says Modell, offers opportunities to design new or better CRISPR-Cas9 tools aimed at regulating gene activity for research purposes. “In a gene editing scenario, a researcher may want to cut a specific gene, in addition to using the long form tracrRNA to inhibit gene activity,” he says.
Here’s a link to and a citation for the paper,
A natural single-guide RNA repurposes Cas9 to autoregulate CRISPR-Cas expression by Rachael E. Workman, Teja Pammi, Binh T.K. Nguyen, Leonardo W. Graeff, Erika Smith, Suzanne M. Sebald, Marie J. Stoltzfus, Chad W. Euler, Joshua W. Modell. Cell DOI:https://doi.org/10.1016/j.cell.2020.12.017 Published Online:J anuary 08, 2021
Before reading further please note, the research discussed in this posting is based on animal testing, which many people find highly disturbing.
CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 (CRISPR-associated protein 9), or more familiarly CRISPR/Cas9, has been been used to edit simian immunodeficiency virus from infected monkeys’ cells according to a December 2, 2020 article by Matthew Rozsa for Salon.com (Note: Links have been removed),
With multiple coronavirus vaccines being produced as we speak, the COVID-19 pandemic appears to have an end in sight, though the HIV pandemic continues after more than 40 years. That might seem like a head-scratcher: why is HIV, a virus we’ve known about for decades, so much harder to cure than a virus discovered just last year? Part of the reason is that HIV, as a retrovirus, is a more complex virus to vaccinate against than SARS-CoV-2 — hence why a vaccine or other cure has eluded scientists for decades.
Now, a surprising new study on a related retrovirus shows incredible promise for the potential to develop a cure for HIV, or human immunodeficiency virus. In an article published in the scientific journal Nature Communications, scientists revealed that they had used CRISPR – a genetic technology that can alter DNA and whose developers won the 2020 Nobel Prize in Chemistry [specifically, Jennifer Doudna and Emanuelle Charpentier received the Nobel for developing CRISPR-cas9 or CRISPR/Cas9 not CRISPR alone) — to successfully edit SIV (simian immunodeficiency virus), a virus similar to HIV, out of the genomes of non-human primates. Specifically, the scientists were able to edit out the SIV genome from rhesus macaque monkeys’ infected cells.
For anyone who’s interested in how CRISPR was developed and the many contributions which have led to the current state-of-the-art for CRISPR gene editing, see the History subsection of Wikipedia’s CRISPR entry.
“This study used the CRISPR CaS9 system, which has been described as molecular scissors,” Andrew G. MacLean, PhD, wrote to Salon. MacLean is an associate professor at the Tulane National Primate Research Center and the Department of Microbiology and Immunology at Tulane University School of Medicine and was a senior co-investigator of the study. “It uses a highly specific targeting system to cut out a specific portion of DNA that is necessary for HIV to be able to produce more virus.”
He added, “Our collaborators at in the Khalili Lab at Temple University have developed a method of ‘packaging’ this within a single so-called vector. A vector is a non-disease causing virus that is used as a carrier for the CRISPR CaS9 scissors to get it into the tissues of interest.”
The experiments with SIV are considered to be a gateway to understanding HIV, as HIV is believed to have evolved from SIV, and is genetically similar.
“The rhesus macaque model of HIV/AIDS is the most valuable model to test efficacy of new interventions or approaches for preventing or treating HIV infection, prior to human clinical trials,” Binhua Ling, PhD, associate professor at the Southwest National Primate Research Center, Texas Biomedical Research Institute, wrote to Salon. “This first proof-of-principal [emphasis mine] study on the rhesus macaque model indicates that this virus-vehicle-delivered-CRISPR system can reach many tissue sites of the body, and is able to effectively delete virus DNA in infected cells. This paves the way for applying the same technology to the human body, which could lead to a cure for HIV infection.”
Tricia H. Burdo, PhD, another senior co-investigator on the new study who works at the Lewis Katz School of Medicine at Temple University, explained to Salon by email that “HIV is in a class of viruses (retroviruses) that inserts itself into the DNA of the host, so you can really think of this now as a genetic disease” — in other words, the kind of thing that would be ripe for CRISPR’s scissors-like ability to remove errant or unwanted genetic material. Burdo notes that the CRISPR technology discussed in the article “cuts out this foreign viral gene.”
…
The study was conducted on eight Rhesus macaque monkeys. That is a very small number to start with and not all of the monkeys received the CRISPR/Cas9 treatment. From the ‘Animals used in the study and ethical statement‘ subsection of the study, “Animals were sacrificed for tissue collection 3 weeks after … .” Leaving aside how anyone may feel about ‘sacrificing …’, three weeks is not a long time for observation.
If you want to read the whole study, here’s a link and a citation,
CRISPR based editing of SIV proviral DNA in ART treated non-human primates by Pietro Mancuso, Chen Chen, Rafal Kaminski, Jennifer Gordon, Shuren Liao, Jake A. Robinson, Mandy D. Smith, Hong Liu, Ilker K. Sariyer, Rahsan Sariyer, Tiffany A. Peterson, Martina Donadoni, Jaclyn B. Williams, Summer Siddiqui, Bruce A. Bunnell, Binhua Ling, Andrew G. MacLean, Tricia H. Burdo & Kamel Khalili. Nature Communications volume 11, Article number: 6065 (2020) DOI: https://doi.org/10.1038/s41467-020-19821-7 Published: 27 November 2020
This paper is open access.
As Rozsa notes in his December 2, 2020 article, the Joint United Nations Programme on HIV/AIDS estimates that 32.7 million [24.8 million–42.2 million] people have died from AIDS-related illnesses since the start (1981?) of the epidemic to the end of 2019.
“Plus ça change, plus c’est la même chose (the more things change, the more things stay the same), is an old French expression that came to mind when I stumbled across two stories about genetic manipulation of food-producing plants.
The first story involves CRISPR (clustered regularly interspersed short palindromic repeats) gene editing and the second involves more ancient ways to manipulate plant genetics.
Getting ‘CRISPR’d’ plant cells to grow into plants
Plants often don’t grow from cells after researchers alter their genomes. Using a new technology, a team coaxed wheat (above) and other crops to more readily produce genome-edited healthy adult plants. Credit: Juan Debernardi
An October 13, 2020 news item on phys.org announces research about getting better results after a plant’s genome has been altered,
Researchers know how to make precise genetic changes within the genomes of crops, but the transformed cells often refuse to grow into plants. One team has devised a new solution.
Scientists who want to improve crops face a dilemma: it can be difficult to grow plants from cells after you’ve tweaked their genomes.
A new tool helps ease this process by coaxing the transformed cells, including those modified with the gene-editing system CRISPR-Cas9, to regenerate new plants. Howard Hughes Medical Institute Research Specialist Juan M. Debernardi and Investigator Jorge Dubcovsky, together with David Tricoli at the University of California, Davis [UC Davis] Plant Transformation Facility, Javier Palatnik from Argentina, and colleagues at the John Innes Center [UK], collaborated on the work. The team reports the technology, developed in wheat and tested in other crops, October 12, 2020, in the journal Nature Biotechnology.
“The problem is that transforming a plant is still an art [emphasis mine],” Dubcovsky says. The success rate is often low – depending on the crop being modified, 100 attempts may yield only a handful of green shoots that can turn into full-grown plants. The rest fail to produce new plants and die. Now, however, “we have reduced this barrier,” says Dubcovsky, a plant geneticist at UC Davis. Using two genes that already control development in many plants, his team dramatically increased the formation of shoots in modified wheat, rice, citrus, and other crops.
Although UC Davis has a pending patent for commercial applications, Dubcovsky says the technique is available to any researcher who wants to use it for research, at no charge. A number of plant breeding companies have also expressed interested in licensing it. “Now people are trying it in multiple crops,” he says.
Humans have worked to improve plants since the dawn of agriculture, selecting wild grasses to produce cultivated maize and wheat, for example. Nowadays, though, CRISPR has given researchers the ability to make changes to the genome with surgical precision. They have used it to create wheat plants with larger grains, generate resistance to fungal infection, design novel tomato plant architectures, and engineer other traits in new plant varieties.
But the process isn’t easy. Scientists start out with plant cells or pieces of tissue, into which they introduce the CRISPR machinery and a small guide to the specific genes they’d like to edit. They must then entice the modified cells into forming a young plant. Most don’t sprout – a problem scientists are still working to understand.
They have tried to find work-arounds, including boosting the expression of certain genes that control early stages of plant development. While this approach has had some success, it can lead to twisted, stunted, sterile plants if not managed properly.Dubcovsky and his colleagues looked at two other growth-promoting genes, GRF and GIF, that work together in young tissues or organs of plants ranging from moss to fruit trees. The team put these genes side-by-side, like a couple holding hands, before adding them to plant cells. “If you go to a dance, you need to find your partner,” Dubcovsky says. “Here, you are tied with a rope to your partner.”
Dubcovsky’s team found that genetically altered wheat, rice, hybrid orange, and other crops produced many more shoots if those experiments included the linked GRF and GIF genes. In experiments with one variety of wheat, the appearance of shoots increased nearly eight-fold. The number of shoots in rice and the hybrid orange, meanwhile, more than doubled and quadrupled, respectively. What’s more, these shoots grew into healthy plants capable of reproducing on their own, with none of the defects that can result when scientists boost other development-controlling genes. That’s because one of the genes is naturally degraded in adult tissues, Dubcovsky says.
Caroline Roper, a plant pathologist at University of California, Riverside who was not involved in the work, plans to use the new technology to study citrus greening, a bacterial disease that kills trees and renders oranges hard and bitter.
To understand how citrus trees can protect themselves, she needs to see how removing certain genes alters their susceptibility to the bacterium — information that could lead to ways to fight the disease. With conventional techniques, it could take at least two years to generate the gene-edited plants she needs. She hopes Dubcovsky’s tool will shorten that timeline.
“Time is of the essence. The growers, they wanted an answer yesterday, because they’re at the brink of having to abandon cultivating citrus,” she says.
For anyone who noticed the reference to citrus greening in the last paragraphs of this news release, I have more information aboutthe disease and efforts to it in an August 6, 2020 posting.
As for the latest in gene editing and regeneration, here’s a link to and a citation for the paper,
A GRF–GIF chimeric protein improves the regeneration efficiency of transgenic plants by Juan M. Debernardi, David M. Tricoli, Maria F. Ercoli, Sadiye Hayta, Pamela Ronald, Javier F. Palatnik & Jorge Dubcovsky. Nature Biotechnology volume 38, pages 1274–1279(2020) DOI: https://doi.org/10.1038/s41587-020-0703-0 First Published Online: 12 October 2020 Journal Issue Date: November 2020
This paper is behind a paywall.
Ancient farming techniques for engineering crops
I stumbled on this story by Gabriela Serrato Marks for Massive Science almost three years late (it’s a Dec. 5, 2017 article),
There are more than 50 strains of maize, called landraces, grown in Mexico. A landrace is similar to a dog breed: Corgis and Huskies are both dogs, but they were bred to have different traits. Maize domestication worked the same way.
Some landraces of maize can grow in really dry conditions; others grow best in wetter soils. Early maize farmers selectively bred maize landraces that were well-adapted to the conditions on their land, a practice that still continues today in rural areas of Mexico.
If you think this sounds like an early version of genetic engineering, you’d be correct. But nowadays, modern agriculture is moving away from locally adapted strains and traditional farming techniques and toward active gene manipulation. The goal of both traditional landrace development and modern genetic modification has been to create productive, valuable crops, so these two techniques are not necessarily at odds.
But as more farmers converge on similar strains of (potentially genetically modified) seeds instead of developing locally adapted landraces, there are two potential risks: one is losing the cultural legacy of traditional agricultural techniques that have been passed on in families for centuries or even millennia, and another is decreasing crop resilience even as climate variability is increasing.
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Mexico is the main importer of US-grown corn, but that imported corn is primarily used to feed livestock. The corn that people eat or use to make tortillas is grown almost entirely in Mexico, which is where landraces come in.
It is a common practice to grow multiple landraces with different traits as an insurance policy against poor growth conditions. The wide range of landraces contains a huge amount of genetic diversity, making it less likely that one adverse event, such as a drought or pest infestation, will wipe out an entire crop. If farmers only grow one type of corn, the whole crop is vulnerable to the same event.
Landraces are also different from most commercially available hybrid strains of corn because they are open pollinating, which means that farmers can save seeds and replant them the next year, saving money and preserving the strain. If a landrace is not grown anymore, its contribution to maize’s genetic diversity is permanently lost.
This diversity was cultivated over generations from maize’s wild cousin, teosinte, by 60 groups of indigenous people in Mexico. Teosinte looks like a skinny, hairier version of maize. It still grows wild in some parts of Central America, but its close relatives have been found, domesticated, at archaeological sites in the region over 9,000 years old. These early maize cobs could easily fit in the palm of your hand – not big enough to be a staple crop that early farmers could depend upon for sustenance. Genetically, they were more similar to wild teosinte than to modern maize.
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[] archaeologists also found that the cobs in Honduras, which is outside the natural range of teosinte, were larger than cobs of the same age from the original domestication region in southern Mexico. The scientists think that people in Honduras were able to develop more productive maize landraces because their crops were isolated from wild teosinte.
The size and shape of the ancient cobs from Honduras show that early farmers engineered the maize crop [emphasis mine] to make it more productive. They developed unique landraces that were well adapted to local conditions and successfully cultivated enough maize to support their communities. In many ways, they were early geneticists. [emphasis mine] …
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We have a lot to learn from the indigenous farmers who were growing maize 4,000 years ago. Their history provides examples of both environmentally sound genetic modification and effective adaptation to climate variability. [emphases mine] …
It seems lignin is a bit of a problem. Its presence in a tree makes processing the wood into various products more difficult. (Of course, some people appreciate trees for other reasons both practical [carbon sequestration?] and/or aesthetic.)
In any event, scientists have been working on ways to reduce the amount of lignin in poplar trees since at least 2014 (see my April 7, 2014 posting titled ‘Good lignin, bad lignin: Florida researchers use plant waste to create lignin nanotubes while researchers in British Columbia develop trees with less lignin’; scroll down about 40% of the way for the ‘less lignin’ story).
(I don’t believe the 2014 research was accomplished with the CRISPR (clustered regularly interspaced short palindromic repeats)-Cas9 technique as it had only been developed in 2012.)
The latest in the quest to reduce the amount of lignin of poplar trees comes from a Belgian/US team, from an Oct. 6, 2020 news item on ScienceDaily,
Researchers led by prof. Wout Boerjan (VIB-UGent [Ghent University] Center for Plant Systems Biology) have discovered a way to stably finetune the amount of lignin in poplar by applying CRISPR/Cas9 technology. Lignin is one of the main structural substances in plants and it makes processing wood into, for example, paper difficult. This study is an important breakthrough in the development of wood resources for the production of paper with a lower carbon footprint, biofuels, and other bio-based materials. Their work, in collaboration with VIVES University College (Roeselare, Belgium) and University of Wisconsin (USA) appears in Nature Communications.
Picture Tailoring lignin and growth by creating CCR2 allelic variants (From left to right: wild type, CCR2(-/-), CCR2(-/*) line 206, CCR2(-/*) line 12) Courtesy: VIB (Flanders Institute of Biotechnology)
Today’s fossil-based economy results in a net increase of CO2 in the Earth’s atmosphere and is a major cause of global climate change. To counter this, a shift towards a circular and bio-based economy is essential. Woody biomass can play a crucial role in such a bio-based economy by serving as a renewable and carbon-neutral resource for the production of many chemicals. Unfortunately, the presence of lignin hinders the processing of wood into bio-based products.
Prof. Wout Boerjan (VIB-UGent): “A few years ago, we performed a field trial with poplars that were engineered to make wood containing less lignin. Most plants showed large improvements in processing efficiency for many possible applications. The downside, however, was that the reduction in lignin accomplished with the technology we used then – RNA interference – was unstable and the trees grew less tall.”
New tools
Undeterred, the researchers went looking for a solution. They employed the recent CRISPR/Cas9 technology in poplar to lower the lignin amount in a stable way, without causing a biomass yield penalty. In other words, the trees grew just as well and as tall as those without genetic changes.
Dr. Barbara De Meester (VIB-UGent): “Poplar is a diploid species, meaning every gene is present in two copies. Using CRISPR/Cas9, we introduced specific changes in both copies of a gene that is crucial for the biosynthesis of lignin. We inactivated one copy of the gene, and only partially inactivated the other. The resulting poplar line had a stable 10% reduction in lignin amount while it grew normally in the greenhouse. Wood from the engineered trees had an up to 41% increase in processing efficiency”.
Dr. Ruben Vanholme (VIB-UGent): “The mutations that we have introduced through CRISPR/Cas9 are similar to those that spontaneously arise in nature. The advantage of the CRISPR/Cas9 method is that the beneficial mutations can be directly introduced into the DNA of highly productive tree varieties in only a fraction of the time it would take by a classical breeding strategy.”
The applications of this method are not only restricted to lignin but might also be useful to engineer other traits in crops, providing a versatile new breeding tool to improve agricultural productivity.
It’s possible there’s a more dramatic development in the field of contemporary gene-editing but it’s indisputable that CRISPR (clustered regularly interspaced short palindromic repeats) -cas9 (CRISPR-associated 9 [protein]) ranks very highly indeed.
The technique, first discovered (or developed) in 2012, has brought recognition in the form of the 2020 Nobel Prize for Chemistry to CRISPR’s two discoverers, Emanuelle Charpentier and Jennifer Doudna.
The Nobel Prize in chemistry went to two researchers Wednesday [October 7, 2020] for a gene-editing tool that has revolutionized science by providing a way to alter DNA, the code of life—technology already being used to try to cure a host of diseases and raise better crops and livestock.
Emmanuelle Charpentier of France and Jennifer A. Doudna of the United States won for developing CRISPR-cas9, a very simple technique for cutting a gene at a specific spot, allowing scientists to operate on flaws that are the root cause of many diseases.
“There is enormous power in this genetic tool,” said Claes Gustafsson, chair of the Nobel Committee for Chemistry.
More than 100 clinical trials are underway to study using CRISPR to treat diseases, and “many are very promising,” according to Victor Dzau, president of the [US] National Academy of Medicine.
“My greatest hope is that it’s used for good, to uncover new mysteries in biology and to benefit humankind,” said Doudna, who is affiliated with the University of California, Berkeley, and is paid by the Howard Hughes Medical Institute, which also supports The Associated Press’ Health and Science Department.
The prize-winning work has opened the door to some thorny ethical issues: When editing is done after birth, the alterations are confined to that person. Scientists fear CRISPR will be misused to make “designer babies” by altering eggs, embryos or sperm—changes that can be passed on to future generations.
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Unusually for phys.org, this October 7, 2020 news item is not a simple press/news release reproduced in its entirety but a good overview of the researchers’ accomplishments and a discussion of some of the issues associated with CRISPR along with the press release at the end.
An October 7, 2020 article by Michael Grothaus for Fast Company provides a business perspective (Note: A link has been removed),
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Needless to say, research by the two scientists awarded the Nobel Prize in Chemistry today has the potential to change the course of humanity. And with that potential comes lots of VC money and companies vying for patents on techniques and therapies derived from Charpentier’s and Doudna’s research.
One such company is Doudna’s Editas Medicine [according to my search, the only company associated with Doudna is Mammoth Biosciences, which she co-founded], while others include Caribou Biosciences, Intellia Therapeutics, and Casebia Therapeutics. Given the world-changing applications—and the amount of revenue such CRISPR therapies could bring in—it’s no wonder that such rivalry is often heated (and in some cases has led to lawsuits over the technology and its patents).
As Doudna explained in her book, A Crack in Creation: Gene Editing and the Unthinkable Power to Control Evolution, cowritten by Samuel H. Sternberg …, “… —but we could also have woolly mammoths, winged lizards, and unicorns.” And as for that last part, she made clear, “No, I am not kidding.”
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Everybody makes mistakes and the reference to Editas Medicine is the only error I spotted. You can find out more about Mammoth Biosciences here and while Dr. Doudna’s comment, “My greatest hope is that it’s used for good, to uncover new mysteries in biology and to benefit humankind,” is laudable it would seem she wishes to profit from the discovery. Mammoth Biosciences is a for-profit company as can be seen at the end of the Mammoth Biosciences’ October 7, 2020 congratulatory news release,
About Mammoth Biosciences
Mammoth Biosciences is harnessing the diversity of nature to power the next-generation of CRISPR products. Through the discovery and development of novel CRISPR systems, the company is enabling the full potential of its platform to read and write the code of life. By leveraging its internal research and development and exclusive licensing to patents related to Cas12, Cas13, Cas14 and Casɸ, Mammoth Biosciences can provide enhanced diagnostics and genome editing for life science research, healthcare, agriculture, biodefense and more. Based in San Francisco, Mammoth Biosciences is co-founded by CRISPR pioneer Jennifer Doudna and Trevor Martin, Janice Chen, and Lucas Harrington. The firm is backed by top institutional investors [emphasis mine] including Decheng, Mayfield, NFX, and 8VC, and leading individual investors including Brook Byers, Tim Cook, and Jeff Huber.
Prize amount: 10 million Swedish kronor, to be shared equally between the Laureates.
In Canadian money that amount is $1,492,115.03 (as of Oct. 9, 2020 12:40 PDT when I checked a currency converter).
Ordinarily there’d be a mildly caustic comment from me about business opportunities and medical research but this is a time for congratulations to both Dr. Emanuelle Charpentier and Dr. Jennifer Doudna.
Heidi Ledford’s June 25, 2020 article (Note: Links have been removed) for Nature focuses on three studies (not yet peer-reviewed) that viewed together suggest CRISPR (clustered regularly interspaced short palindromic repeats) gene-editing is less like using a pair of scissors to cut out unwanted mutations and more like using a catalyst (a chemical agent which increases chemical reactions) and getting unanticipated and unwatned reactions. Except, it’s an unpredictable catalyst.
A suite of experiments that use the gene-editing tool CRISPR–Cas9 to modify human embryos have revealed how the process can make large, unwanted changes to the genome at or near the target site. [emphasis mine]
The studies were published this month on the preprint server bioRxiv, and have not yet been peer-reviewed1,2,3. But taken together, they give scientists a good look at what some say is an underappreciated risk of CRISPR–Cas9 editing. Previous experiments have revealed that the tool can make ‘off target’ gene mutations far from the target site, but the nearby changes identified in the latest studies can be missed by standard assessment methods.
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These safety concerns are likely to inform the ongoing debate over whether scientists should edit human embryos to prevent genetic diseases — a process that is controversial because it creates a permanent change to the genome that can be passed down for generations. “If human embryo editing for reproductive purposes or germline editing were space flight, the new data are the equivalent of having the rocket explode at the launch pad before take-off,” says Fyodor Urnov, who studies genome editing at the University of California, Berkeley, but was not involved in any of the latest research.
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These studies,if borne out, offer new concerns (from Ledford’s June 25, 2020 article),
The changes are the result of DNA-repair processes harnessed by genome-editing tools. CRISPR–Cas9 uses a small strand of RNA to direct the Cas9 enzyme to a site in the genome with a similar sequence. The enzyme then cuts both strands of DNA at that site, and the cell’s repair systems heal the gap.
The edits occur during that repair: most often, the cell seals up the cut using an error-prone mechanism that can insert or delete a small number of DNA letters. If researchers provide a DNA template, the cell might sometimes use that sequence to mend the cut, resulting in a true rewrite. But broken DNA can also cause shuffling or loss of a large region of the chromosome.
Previous work using CRISPR in mouse embryos and other kinds of human cell had already demonstrated that editing chromosomes can cause large, unwanted effects4,5. But it was important to demonstrate the work in human embryos as well, says Urnov, because different cell types might respond to genome editing differently.
Such rearrangements could be missed in many experiments, which typically look for other unwanted edits, such as single DNA-letter changes or small insertions or deletions of only a few letters. The latest studies, however, looked specifically for large deletions and chromosomal rearrangements near the target site. [emphasis mine] “This is something that all of us in the scientific community will, starting immediately, take more seriously than we already have,” says Urnov. “This is not a one-time fluke.”
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Ledford’s article offers some description and analysis of each of the three papers.Note: All of the research was done with nonviable embryos. For anyone who wants to read the papers for themselves here are links and citations for each of the three,
Frequent loss-of-heterozygosity in CRISPR-Cas9-edited early human embryos by Gregorio Alanis-Lobato, Jasmin Zohren, Afshan McCarthy, Norah M.E. Fogarty, Nada Kubikova, Emily Hardman, Maria Greco, Dagan Wells, James M.A. Turner, Kathy K. Niakan. bioRxiv DOI: https://doi.org/10.1101/2020.06.05.135913 Posted: June 5, 2020
FREQUENT GENE CONVERSION IN HUMAN EMBRYOS INDUCED BY DOUBLE STRAND BREAKS by Dan Liang, Nuria Marti Gutierrez, Tailai Chen, Yeonmi Lee, Sang-Wook Park, Hong Ma, Amy Koski, Riffat Ahmed, Hayley Darby, Ying Li, Crystal Van Dyken, Aleksei Mikhalchenko, Thanasup Gonmanee, Tomonari Hayama, Han Zhao, Keliang Wu, Jingye Zhang, Zhenzhen Hou, Jumi Park, Chong-Jai Kim, Jianhui Gong, Yilin Yuan, Ying Gu, Yue Shen, Susan B. Olson, Hui Yang, David Battaglia, Thomas O’Leary, Sacha A. Krieg, David M. Lee, Diana H. Wu, P. Barton Duell, Sanjiv Kaul, Jin-Soo Kim, Stephen B. Heitner, Eunju Kang, Zi-Jiang Chen, Paula Amato, Shoukhrat Mitalipov. bioRxiv DOI: https://doi.org/10.1101/2020.06.19.162214 Posted June 20, 2020
These papers are open access.
A July 17, 2018 posting is probably the first time I featured work showing that CRISPR gene-editing can result in off-target effects; it was followed up by a September 20, 2019 posting on the topic.
