Tag Archives: Chromosome

A novel approach to quantify personal information contained within gut metagenome data

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In a recent study published in Nature Microbiology, researchers used shotgun sequencing to extract human reads from deoxyribonucleic acid (DNA) in fecal samples of 343 Japanese individuals comprising the main dataset of this study.

They used this gut metagenome data to reconstruct personal information. Some study participants also provided whole genome sequencing (WGS) data for ultra-deep metagenome shotgun sequencing analysis.

Study: Reconstruction of the personal information from human genome reads in gut metagenome sequencing data. Image Credit: KaterynaKon/Shutterstock.comStudy: Reconstruction of the personal information from human genome reads in gut metagenome sequencing data. Image Credit: KaterynaKon/Shutterstock.com

Background

The knowledge regarding the human microbiome, microorganisms inhabiting the human body, has expanded considerably in the last ten years, thanks to rapid advancements in technologies like metagenome shotgun sequencing.

This technology allows the sequencing of the non-bacterial component of the microbiome samples, including host DNA. For instance, in fecal samples, the amount of host DNA is less than 10% but is removed to protect the privacy of donors.

Human germline genotype in metagenome data is substantial to enable the re-identification of individuals. However, researchers and donors should recognize that it is highly confidential, so sharing it with the community requires careful consideration.

Apart from ethical concerns related to sharing this data, it is necessary to understand that if human reads in metagenome data are not removed before deposition, what kind of personal information (e.g., sex and ancestry) could this data help recover?

In addition, human reads in gut metagenome data could be a good resource for stool-based forensics, robust variant calling, and polygenic risk scores based estimates of disease risks (e.g., type 2 diabetes).

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Since this data could help quantitatively and precisely reconstruct genotype information, it could complement human WGS data.

About the study

In the present study, researchers applied a few humans reads in the gut metagenome data of the main study dataset to reconstruct personal information, including genetic sex and ancestry. For predicting genetic sex and the ancestries of these 343 individuals, they used sequencing depth of the sex chromosomes and modified likelihood score-based method, respectively.

In addition, the researchers developed methods to re-identify a person from a genotype dataset. Furthermore, they combined two harmonized genotype-calling approaches, the direct calling of rare variants and the two-step imputation of common variants, to reconstruct genotypes.

The main dataset of the study included 343 Japanese participants, whereas the validation dataset for the genetic sex prediction analysis comprised 113 Japanese individuals.

The multi-ancestry dataset, which helped the researchers validate ancestry prediction analysis, comprised 73 individuals of various nationalities, including samples from individuals in New Delhi, India.

The female and male participants in each dataset were 196 & 147, 65 & 48, and 25 & 48, respectively. Likewise, the age range for these three datasets was 20 to 88, 20 to 81, and 20 to 61 years, respectively.

Results and conclusion

Given that human reads in the gut metagenome data were derived consistently from all chromosomes, the read depth of the X chromosome was nearly double in females and that of the Y chromosome in males.

So, in a logistic regression analysis, when the researchers applied a 0.43 Y:X chromosome read-depth ratio to the validation dataset, which correctly predicted the genetic sex of 97.3% of the study samples.

In human microbiome and genetic research, the feasibility of sex prediction using human gut metagenome data could help remove mislabelled samples.

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The study analysis also helped researchers remarkably predict ancestry in 98.3% of individuals using 1,000 Genomes Project (1KG) data as a reference.

However, the likelihood score-based method often misclassified South Asian (SAS) samples as American (AMR) and European (EUR), especially when the number of human reads was small. It is understandable because the genetic diversity of the SAS population is complex.

The likelihood score-based method also efficiently utilized the data from genomic areas with low coverage demonstrating the quantitative power of gut metagenome data to re-identify individuals and successfully re-identified 93.3% of individuals.

Despite ethical concerns, the re-identification method used in this study could help in the quality control of multi-omics datasets comprising gut metagenome and human germline genotype data.

In addition, the authors successfully reconstructed genome-wide common variants using genomic approaches. Historically researchers used stool samples as a source of germline genomes for wild and domestic animals but not humans.

Thus, further development of suitable methodologies could help efficiently utilize the human genome in gut metagenome data and benefit animal research.

Nonetheless, the study remarkably demonstrated that optimized methods could help reconstruct personal information from the human reads in gut metagenome data.

Moreover, the findings of this study could serve as a guiding resource to devise best practices for using the already accumulated gut metagenome data of humans.

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Journal reference:

Scientists discover antibiotic resistance genes in clouds

The atmosphere is a large-scale dissemination route for bacteria carrying antibiotic-resistance genes. A research team from Université Laval and Université Clermont Auvergne has shown that these genes can be transported by clouds.

“This is the first study to show that clouds harbor antibiotic resistance genes of bacterial origin in concentrations comparable to other natural environments,” says Florent Rossi, first author of the study and postdoctoral fellow in the team of Caroline Duchaine, a professor at Université Laval’s Faculty of Science and Engineering and a researcher at the Quebec Heart and Lung Institute-Université Laval.

To observe this phenomenon, the team sampled clouds at the Puy de Dôme summit, a dormant volcano in France’s Massif Central. At an atmospheric research station perched 1,465 meters above ground, the scientists conducted 12 cloud sampling sessions over two years using high-flow rate “vacuums.”

Analysis of these samples revealed that they contained about 8,000 bacteria per milliliter of cloud water, on average. “These bacteria usually live on the surface of vegetation or soil. They are aerosolized by the wind or by human activities, and some of them rise into the atmosphere and participate in the formation of clouds,” explains Florent Rossi. The concentrations are variable: they range from 330 to more than 30,000 bacteria per milliliter of cloud water. Between 5% and 50% of these bacteria could be alive and potentially active.

Various sources

With all their data, the scientists measured the concentration of 29 subtypes of antibiotic-resistance genes carried in atmospheric air masses. The clouds contained, on average, 20,800 copies of antibiotic-resistance genes per milliliter of cloud water.

“Oceanic clouds and continental clouds each have their signature of antibiotic resistance genes. For example, continental clouds contain more antibiotic resistance genes used in animal production,” explains Florent Rossi.

Although airborne transport of antibiotic resistance genes is a natural phenomenon, the widespread use of antibiotics in agriculture and medicine has contributed to the proliferation of these resistant strains and their dissemination in the environment.

“Our study shows that clouds are an important pathway for antibiotic-resistance genes spreading over short and long ranges. Ideally, we would like to locate emission sources resulting from human activities to limit the dispersal of these genes.”

The health effect of the spread of these antibiotic-resistant genes will be something to investigate in future research.

  • Florent Rossi, Raphaëlle Péguilhan, Nathalie Turgeon, Marc Veillette, Jean-Luc Baray, Laurent Deguillaume, Pierre Amato, Caroline Duchaine. Quantification of antibiotic resistance genes (ARGs) in clouds at a mountain site (puy de Dôme, central France). Science of The Total Environment, 2023; 865: 161264 DOI: 10.1016/j.scitotenv.2022.161264
  • Université Laval

    Elucidating the function of BRCA2 gene offers insight into cancer development

    A new study shows exactly how the gene BRCA2, linked to susceptibility to breast and ovarian cancer, functions to repair damaged DNA. By studying BRCA2 at the level of single molecules, researchers at the University of California, Davis, have generated new insights into the mechanisms of DNA repair and the origins of cancer. The work was published the week of March 27 in the Proceedings of the National Academy of Sciences.

    Elucidating the function of BRCA2 is essential for understanding the molecular etiology of cancer development in breast and ovarian cells, as well as many other cell types including prostate.”

    Stephen Kowalczykowski, distinguished professor of microbiology and molecular genetics, UC Davis College of Biological Sciences

    By visualizing BRCA2 function at a single molecule level, Kowalczykowski’s team discovered that it acts as a molecular chaperone, delivering another protein, RAD51, to single-stranded DNA. It ensures formation of a functional filament of RAD51 and the repair of broken DNA.

    “When BRCA2 is defective, broken DNA is not faithfully repaired, the genome loses integrity, and cancer ultimately ensues,” Kowalczykowski said.

    Mutations in the BRCA2 gene are linked to an increased risk of cancer, especially breast and ovarian cancer. In 2010, teams led by Kowalczykowski and by Professor Wolf-Dietrich Heyer in the same department at UC Davis succeeded in purifying the BRCA2 protein and showed that it plays a key role in DNA repair.

    The new work, using techniques developed in Kowalczykowski’s lab to image single proteins and DNA molecules in real time, gives new insight into the mechanics of this repair process.

    Our DNA is under constant assault by both processes inside cells and by outside factors, such as sunlight or chemical exposures. Accumulating damage to DNA can cause cells to become cancerous. Fortunately, our cells have several mechanisms to repair DNA. One of these is homologous recombination to repair double-stranded breaks.

    Repairing double-stranded breaks

    When a break crosses both strands of the DNA double helix, one strand is trimmed back a little to leave a single exposed strand. This strand then goes hunting for its counterpart in the same gene in the matching paired chromosome. It inserts into the healthy DNA and uses it as a template for repair.

    For this insertion to work, the single strand of DNA has to be coated with RAD51. Earlier work from Kowalczykowski’s lab measured how quickly RAD51 could be added onto DNA, like beads on a string.

    The function of BRCA2 is to load up with RAD51 (each BRCA2 can carry up to six RAD51s), push another protein called RPA out of the way and put the proteins onto the DNA.

    Postdoctoral researcher Jason Bell carried out the experiments observing RAD51 and BRCA2 working their way along the DNA. Bell manipulated pieces of DNA with a single-stranded gap and exposed them to RAD51 with and without BRCA2 under different conditions.

    The resulting movies show how BRCA2 chaperones RAD51 onto single-stranded DNA, displacing RPA.

    Understanding the role of BRCA2 in DNA repair has two important implications. First, it helps us understand why mutations of BRCA2 lead to an increased risk of cancer. Second, some drugs to treat cancer work by damaging DNA. By understanding how DNA repair works, we can develop new drugs to target it specifically in cancer cells.

    Additional co-authors on the paper are Christopher Dombrowski and Jody Plank, both at UC Davis, and Ryan Jensen, formerly at UC Davis and now at the Yale University School of Medicine. The work was supported by grants from the National Institutes of Health.

    Source:
    Journal reference:

    Bell, J. C., et al. (2023). BRCA2 chaperones RAD51 to single molecules of RPA-coated ssDNA. Proceedings of the National Academy of Sciences. doi.org/10.1073/pnas.2221971120.

    Epigenome reprogramming after SARS-CoV-2 infection

    In a recent article in published in the journal Nature Microbiology, researchers in Texas, United States (US) performed a three-dimensional (3D) evaluation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infected human cells to show a direct cell-autonomous effect elicited by SARS-CoV-2 on the host chromatin.

    The study aimed at improving the understanding of coronavirus disease 2019 (COVID-19)-related perturbations in the genome and epigenome of a host cell.

    Study: SARS-CoV-2 restructures host chromatin architecture. Image Credit:FUNFUNPHOTO/Shutterstock.com

    Study: SARS-CoV-2 restructures host chromatin architecture. Image Credit:FUNFUNPHOTO/Shutterstock.com

    Background

    The 3D folding of chromatin in mammals, including humans, influences deoxyribonucleic acid (DNA) replication, recombination, DNA damage repair, and transcription. It is a key determinant of how human cells act and function. Viruses, including SARS-CoV-2, antagonize host defense by rewiring their chromatin architecture, which typically has several layers, e.g., A/B compartments, chromatin loops, and topological associating domains (TADs).

    The A and B compartments superimpose transcriptionally active euchromatin and relatively inactive heterochromatin, respectively. However, studies have barely investigated these effects.

    In addition, epigenetic alterations impact gene expression and resulting phenotypes in the long term. Thus, a sneak peek into the interactions between the virus, host chromatin, and epigenome could help find novel methods to fight SARS-CoV-2 in the acute phase. In addition, it could unravel the molecular basis of post-acute SARS-CoV-2 sequelae or long COVID and subsequently mitigate it.

    About the study

    At 24 hours post-infection (24 hpi), human A549 cells expressing angiotensin-converting enzyme 2 (ACE2), infected with SARS-CoV-2 at a multiplicity of infection (MOI) of 0.1, had high levels of infection. This was shown by ribonucleic acid-sequencing (RNA-seq). Immunofluorescence of the SARS-CoV-2 spike (S) glycoprotein also substantiated an elevated infection ratio.

    So, in the present study, researchers used an improved version of in situ Hi-C high-throughput chromosome conformation capture (Hi-C) 3.0 to study host chromatin changes in these cells at 24 hpi and mock-infected cells (Mock).

    In addition, the team evaluated the epigenetic features of the altered chromatin regions to understand the vulnerability to compartmental changes due to infection. To this end, they used chromatin immunoprecipitation (ChIP-seq) methods to generate data on representative histone markers and polymerase II (Pol2) in A549-ACE2 cells. This analysis covered four histone markers, viz., H3K27ac, H3K4me3, H3K9me3, and H3K27me3.

    It helped them examine the epigenetic features of these six categories of bins. They ranked E1-score changes for each genomic bin to sort bins. They dubbed bins showing E1-score increase and decrease as ‘A-ing’ and ‘B-ing’ bins, respectively.

    Results

    The Hi-C analysis showed extensive alterations in the hosts’ 3D genome after SARS-CoV-2 infection. The researchers also plotted a Pearson correlation map of their Hi-C analysis that reaffirmed these changes alongside indicating modified chromatin compartmentalization.

    A focused view of the ~0.7 Mb region showed a weakening of the rectangle-shaped chromatin domains and deregulation of chromatin loops. While SARS-CoV-2 prompted a global decline in near-diagonal short-range chromatin contacts (<560 kilobases), as seen in a P(s) curve, chromatin contacts far-separated from the diagonal (>28 megabases) were often deregulated.

    Further, a P(s) curve showed that SARS-CoV-2 elicited modest and enhanced interactions in middle-to-long-distance contacts (~560 kb to 8.9 Mb) and far-positioned regions, respectively.

    Fold changes in inter-chromosomal interactions or trans-vs-cis contact ratios also depicted the effect of SARS-CoV-2 infection on inter-chromosomal contacts. The enhancement of inter- and intra-chromosomal interactions indicated changes in chromatin compartmentalization. Consequently, principal component analysis (PCA) of a 100-kb bin on Hi-C background showed noticeable defects of chromatin compartmentalization in virus-infected cells.

    The total PCA E1 scores quantifying E1 changes in ~30% of genomic regions showed a widespread diminishing of the A compartment, A-to-B switching, or strengthening of the B compartment post-SARS-CoV-2 infection.

    Among all, A to weaker A changes were the most common and occurred in ~18% of the genome, which indicated that SARS-CoV-2 extensively weakened the host euchromatin.

    Further analysis showed that the ‘B-ing’ and ‘A-ing’ genomic regions were historically enriched in active chromatin markers (e.g., H3K27ac) and repressive histone markers, especially H3K27me3. Unexpectedly, SARS-CoV-2 infection selectively modified the H3K4me3 marker of phytochrome interacting factors (PIF) gene promoters, suggesting unappreciated mechanisms at these promoters that confer deviating inflammation in COVID-19.

    A flawed chromatin compartmentalization likely caused the historically well-partitioned A or B compartments to lose their identity. A saddle plot illustrating inter-compartment chromatin interactions across the genome showed these global changes.

    The authors also noted weakened compartmentalization between chromosomes. For instance, in chromosomes 17 & 18, while A–B interactions were amplified, A–A/B–B homotypic interactions appeared to have become compromised.

    Moreover, SARS-CoV-2 infection mechanistically depleted the cohesin complex in a pervasive but selective manner from intra-TAD regions. These changes provided a molecular explanation for the weakening of intra-TAD interactions.

    It supported the notion that defective cohesin loop extrusion inside TADs releases this chromatin to engage in long-distance associations. Intriguingly, chromatin in SARS-CoV-2-infected cells exhibited a higher frequency of long-distance intra-chromosomal and inter-chromosomal interactions.

    Conclusions

    SARS-CoV-2 infection markedly restructured 3D host chromatin, featuring widespread compartment A weakening and A–B mixing and global reduction in intra-TAD chromatin contacts.

    However, it is still unknown exactly how SARS-COV-2 infection restructures host chromatin. Likely, open reading frame 8 (ORF8) disrupts the host epigenome, suggesting that some viral factors are involved in host chromatin rewiring.

    It also altered the host epigenome, including a global reduction in active chromatin mark H3K27ac and a specific increase in H3K4me3 at pro-inflammatory gene promoters. Intriguingly, all these host chromatin alterations were unique to SARS-CoV-2 infection, and other common-cold coronaviruses or immune stimuli did not elicit these changes.

    Journal reference:

    Avanced genome editing technology could be used as a one-time treatment for CD3 delta SCID

    A new UCLA-led study suggests that advanced genome editing technology could be used as a one-time treatment for the rare and deadly genetic disease CD3 delta severe combined immunodeficiency.

    The condition, also known as CD3 delta SCID, is caused by a mutation in the CD3D gene, which prevents the production of the CD3 delta protein that is needed for the normal development of T cells from blood stem cells.

    Without T cells, babies born with CD3 delta SCID are unable to fight off infections and, if untreated, often die within the first two years of life. Currently, bone marrow transplant is the only available treatment, but the procedure carries significant risks.

    In a study published in Cell, the researchers showed that a new genome editing technique called base editing can correct the mutation that causes CD3 delta SCID in blood stem cells and restore their ability to produce T cells.

    The potential therapy is the result of a collaboration between the laboratories of Dr. Donald Kohn and Dr. Gay Crooks, both members of the Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA and senior authors of the study.

    Kohn’s lab has previously developed successful gene therapies for several immune system deficiencies, including other forms of SCID. He and his colleagues turned their attention to CD3 delta SCID at the request of Dr. Nicola Wright, a pediatric hematologist and immunologist at the Alberta Children’s Hospital Research Institute in Canada, who reached out in search of a better treatment option for her patients.

    CD3 delta SCID is prevalent in the Mennonite community that migrates between Canada and Mexico.

    Because newborns are not screened for SCID in Mexico, I often see babies who have been diagnosed late and are returning to Canada quite sick.”

    Dr. Nicola Wright, pediatric hematologist and immunologist at the Alberta Children’s Hospital Research Institute

    When Kohn presented Wright’s request to his lab, Grace McAuley, then a research associate who joined the lab at the end of her senior year at UCLA, stepped up with a daring idea.

    “Grace proposed we try base editing, a very new technology my lab had never attempted before,” said Kohn, a distinguished professor of microbiology, immunology and molecular genetics, and of pediatrics.

    Base editing is an ultraprecise form of genome editing that enables scientists to correct single-letter mutations in DNA. DNA is made up of four chemical bases that are referred to as A, T, C and G; those bases pair together to form the “rungs” in DNA’s double-helix ladder structure.

    While other gene editing platforms, like CRISPR-Cas9, cut both strands of the chromosome to make changes to DNA, base editing chemically changes one DNA base letter into another -; an A to a G, for example -; leaving the chromosome intact.

    “I had a very steep learning curve in the beginning, when base editing just wasn’t working,” said McAuley, who is now pursuing an M.D.-Ph.D. at UC San Diego and is the study’s co-first author. “But I kept pushing forward. My goal was help get this therapy to the clinic as fast as was safely possible.”

    McAuley reached out to the Broad Institute’s David Liu, the inventor of base editing, for advice on how to evaluate the technique’s safety for this particular use. Eventually, McAuley identified a base editor that was highly efficient at correcting the disease-causing genetic mutation.

    Because the disease is extremely rare, obtaining patient stem cells for the UCLA study was a significant challenge. The project got a boost when Wright provided the researchers with blood stem cells donated by a CD3 delta SCID patient who was undergoing a bone marrow transplant.

    The base editor corrected an average of almost 71% of the patient’s stem cells across three laboratory experiments.

    Next, McAuley worked with Dr. Gloria Yiu, a UCLA clinical instructor in rheumatology, to test whether the corrected cells could give rise to T cells. Yiu used artificial thymic organoids, which are stem cell-derived tissue models developed by Crooks’ lab that mimic the environment of the human thymus -; the organ where blood stem cells become T cells.

    When the corrected blood stem cells were introduced into the artificial thymic organoids, they produced fully functional and mature T cells.

    “Because the artificial thymic organoid supports the development of mature T cells so efficiently, it was the ideal system to show that base editing of patients’ stem cells could fix the defect seen in this disease,” said Yiu, who is also a co-first author of the study.

    As a final step, McAuley studied the longevity of the corrected stem cells by transplanting them into a mouse. The corrected cells remained four months after transplant, indicating that base editing had corrected the mutation in true, self-renewing blood stem cells. The findings suggest that corrected blood stem cells could persist long-term and produce the T cells patients would need to live healthy lives.

    “This project was a beautiful picture of team science, with clinical need and scientific expertise aligned,” said Crooks, a professor of pathology and laboratory medicine. “Every team member played a vital role in making this work successful.”

    The research team is now working with Wright on how to bring the new approach to a clinical trial for infants with CD3 delta SCID from Canada, Mexico and the U.S.

    This research was funded by the Jeffrey Modell Foundation, the National Institutes of Health, the Bill and Melinda Gates Foundation, the Howard Hughes Medical Institute, the V Foundation and the A.P. Giannini Foundation.

    The therapeutic approach described in this article has been used in preclinical tests only and has not been tested in humans or approved by the Food and Drug Administration as safe and effective for use in humans. The technique is covered by a patent application filed by the UCLA Technology Development Group on behalf of the Regents of the University of California, with Kohn and McAuley listed as co-inventors.

    Source:
    Journal reference:

    McAuley, G.E., et al. (2023) Human T cell generation is restored in CD3δ severe combined immunodeficiency through adenine base editing. Cell. doi.org/10.1016/j.cell.2023.02.027.

    Decreased viral infection severity in females may be due to extra copy of X chromosome-linked gene

    It has long been known that viral infections can be more severe in males than females, but the question as to why has remained a mystery – until possibly now. The key may lie in an epigenetic regulator that boosts the activity of specialized anti-viral immune cells known as natural killer (NK) cells.

    In a study published March 16 in the peer-reviewed journal Nature Immunology, a collaborative team of UCLA researchers have found that female mouse and human NK cells have an extra copy of an X chromosome-linked gene called UTX. UTX acts as an epigenetic regulator to boost NK cell anti-viral function, while repressing NK cell numbers.

    While it is well-known that males have more NK cells compared to females, we did not understand why the increased number of NK cells was not more protective during viral infections. It turns out that females have more UTX in their NK cells than do males, which allows them to fight viral infections more efficiently.”

    Dr. Maureen Su, co-senior author, professor of microbiology immunology and molecular genetics, and of pediatrics, at the David Geffen School of Medicine at UCLA

    The researchers noted that this held true whether or not the mice had gonads (ovaries in females; testes in males), indicating that the observed trait was not linked to hormones. Furthermore, female mice with lower UTX expression had more NK cells which were not as capable of controlling viral infection.

    “This implicates UTX as a critical molecular determinant of sex differences in NK cells,” said the study’s lead author Mandy Cheng, graduate student in molecular biology at UCLA.

    The findings suggest that therapies involving immune responses need to move beyond a “one-size-fits-all” approach and toward a precision medicine model, also known as personalized medicine, that tailors treatments that take into account people’s individual differences, such as genetics, environment and other factors that influence health and disease risk, the researchers write.

    “Given the recent excitement with using NK cells in the clinic, we will need to incorporate sex as a biological factor in treatment decisions and immunotherapy design,” said co-senior author Tim O’Sullivan, assistant professor of microbiology, immunology and molecular genetics at the Geffen School.

    Source:
    Journal reference:

    Cheng, M.I., et al. (2023) The X-linked epigenetic regulator UTX controls NK cell-intrinsic sex differences. Nature Immunology. doi.org/10.1038/s41590-023-01463-8.

    New discoveries made regarding autism onset in mouse models

    Although autism is a common neurodevelopmental disorder, the multiple factors behind its onset are still not fully understood. Animal models of idiopathic autism, especially mice, are often used to help researchers understand the complicated mechanisms behind the disorder, with BTBR/J being the most commonly used mouse model in the world.

    Now, an international research collaboration including Kobe University’s Professor TAKUMI Toru and Researcher Chia-wen Lin et al. have made new discoveries regarding autism onset in mouse models.

    In their detailed series of experiments and analyses of BTBR/J mice and the other subspecies BTBR/R, they revealed that endogenous retrovirus activation increases a fetus’s susceptibility to autism. They also discovered that BTBR/R exhibits autistic-like behaviors without reduced learning ability, making it a more accurate model of autism than the widely-used BTBR/J model.

    It is hoped that further research will contribute towards better classification of autism types, as well as the creation of new treatment strategies for neurodevelopmental disorders.

    These research results were published in Molecular Psychiatry on March 7, 2023

    Main points

    • The researchers analyzed BTBR/J, a widely used mouse model of autism, and its subspecies BTBR/Rusing MRI. This revealed that the corpus callosum, which connects the left and right hemispheres of the brain, was impaired in BTBR/J mice but not in BTBR/R mice.
    • Genome and transcription analysis showed that BTBR mice have increased levels of endogenous retrovirus genes.
    • Furthermore, single-cell RNA analysis of BTBR/R mice revealed changes in the expression of various genes (including stress response genes) that are indicative of endogenous retrovirus activation.
    • Even though BTBR/J and BTBR/R mice have the same ancestry, the results of various behavioral analysis experiments revealed differences in spatial learning ability and other behaviors between the two types of model mice.

    Research background

    Autism (autism spectrum disorder) is a neurodevelopmental disorder that remains largely unexplored despite the rapidly increasing number of patients. Reasons for this continuing increase in people diagnosed with autism include changes to diagnostic criteria and older fathers becoming more common. Autism is strongly related to genetic factors and can be caused by abnormalities in DNA structure, such as copy number variations. Animal models, especially mice, are often used in research to illuminate the pathology of autism. Among these models, BTBR/J is a mouse model of the natural onset of autism that is commonly used. Studies have reported various abnormalities in BTBR/J mice including impairment of the corpus callosum (which connects the left and right hemispheres of the brain) and excessive immune system signaling. However, it is not fully understood why this particular lineage displays autistic-like behavioral abnormalities.

    The aim of the current study was to shed light on the onset mechanism of these autistic-like behavioral abnormalities by conducting comparative analysis on BTBR/J and its subspecies BTBR/R.

    Research findings

    First of all, the researchers conducted MRI scans on BTBR/J and BTBR/R mice to investigate structural differences in each region of the brain. The results revealed that there were differences between BTBR/J and BTBR/R mice in 33 regions including the amygdala. A particularly prominent difference discovered was that even though BTBR/J’s corpus callosum is impaired, BTBR/R’s is normal.

    Next, the research group used the array CGH method to compare BTBR/R’s copy number variations with that of a normal mouse model (B6). They revealed that BTBR/R mice had significantly increased levels of endogenous retroviruses (ERV) in comparison to B6 mice. Furthermore, qRT-PCR tests revealed that these retroviruses were activated in BTBR/R mice. On the other hand, in B6 mice there was no change in the expression of LINE ERV (which is classified in the same repetitive sequence), indicating that this retroviral activation is specific to BTBR.

    Subsequently, the researchers carried out single-cell RNA analysis on the tissue of embryonic BTBR mice (on the AGM and yolk sac). The results provide evidence of ERV activation in BTBR mice, as expression changes were observed in a group of genes downstream of ERV.

    Lastly, the researchers comprehensively investigated the differences between BTBR/J and BTBR/R on a behavioral level. BTBR/R mice were less anxious than BTBR/J and showed qualitative changes in ultrasound vocalizations, which are measured as a way to assess communicative ability in mice. BTBR/R mice also exhibited more self-grooming behaviors and buried more marbles in the marble burying test. These two tests were designed to detect repetitive behavioral abnormalities in autistic individuals. From the results, it was clear that BTBR/R exhibits more repetitive behaviors (i.e. it is more symptomatic) than BTBR/J. The 3-chamber social interaction test, which measures how closely a mouse will approach another mouse, also revealed more pronounced social deficits in BTBR/R than BTBR/J mice (Figure 4i). In addition, a Barnes maze was used to conduct a spatial learning test, in which BTBR/J mice exhibited reduced learning ability compared to B6 (normal mice). BTBR/R mice, on the other hand, exhibited similar ability to B6.

    Overall, the study revealed that retrovirus activation causes the copy number variants in BTBR mice to increase, which leads to the differences in behavior and brain structure seen in BTBR/J and BTBR/R mice (Figure 5).

    Further developments

    BTBR/J mice are widely used by researchers as a mouse model of autism. However, the results of this study highlight the usefulness of the other lineage of BTBR/R mice because they exhibit autistic-like behavior without compromised spatial learning ability. The results also suggest that it may be possible to develop new treatments for autism that suppress ERV activation. Furthermore, it is necessary to classify autism subtypes according to their onset mechanism, which is a vital first step towards opening up new avenues of treatment for autism.

    Source:
    Journal reference:

    Lin, C-W., et al. (2023) An old model with new insights: endogenous retroviruses drive the evolvement toward ASD susceptibility and hijack transcription machinery during development. Molecular Psychiatry. doi.org/10.1038/s41380-023-01999-z.

    Simple blood tests for telomeric protein could provide a valuable screen for certain cancers

    Once thought incapable of encoding proteins due to their simple monotonous repetitions of DNA, tiny telomeres at the tips of our chromosomes seem to hold a potent biological function that’s potentially relevant to our understanding of cancer and aging.

    Reporting in the Proceedings of the National Academy of Science, UNC School of Medicine researchers Taghreed Al-Turki, PhD, and Jack Griffith, PhD, made the stunning discovery that telomeres contain genetic information to produce two small proteins, one of which they found is elevated in some human cancer cells, as well as cells from patients suffering from telomere-related defects.

    Based on our research, we think simple blood tests for these proteins could provide a valuable screen for certain cancers and other human diseases. These tests also could provide a measure of ‘telomere health,’ because we know telomeres shorten with age.”

    Jack Griffith, PhD, the Kenan Distinguished Professor of Microbiology and Immunology and Member of the UNC Lineberger Comprehensive Cancer Center

    Telomeres contain a unique DNA sequence consisting of endless repeats of TTAGGG bases that somehow inhibit chromosomes from sticking to each other. Two decades ago, the Griffith laboratory showed that the end of a telomere’s DNA loops back on itself to form a tiny circle, thus hiding the end and blocking chromosome-to-chromosome fusions. When cells divide, telomeres shorten, eventually becoming so short that the cell can no longer divide properly, leading to cell death.

    Scientist first identified telomeres about 80 years ago, and because of their monotonous sequence, the established dogma in the field held that telomeres could not encode for any proteins, let alone ones with potent biological function.

    In 2011 a group in Florida working on an inherited form of ALS reported that the culprit was an RNA molecule containing a six-base repeat which by a novel mechanism could generate a series of toxic proteins consisting of two amino acids repeating one after the other. Al-Turki and Griffith note in their paper a striking similarity of this RNA to the RNA generated from human telomeres, and they hypothesized that the same novel mechanism might be in play.

    They conducted experiments – as described in the PNAS paper – to show how telomeric DNA can instruct the cell to produce signaling proteins they termed VR (valine-arginine) and GL (glycine-leucine). Signaling proteins are essentially chemicals that trigger a chain reaction of other proteins inside cells that then lead to a biological function important for health or disease.

    Al-Turki and Griffith then chemically synthesized VR and GL to examine their properties using powerful electron and confocal microscopes along with state-of-the-art biological methods, revealing that the VR protein is present in elevated amounts in some human cancer cells, as well as cells from patients suffering from diseases resulting from defective telomeres.

    “We think it’s possible that as we age, the amount of VR and GL in our blood will steadily rise, potentially providing a new biomarker for biological age as contrasted to chronological age,” said Al-Turki, a postdoctoral researcher in the Griffith lab. “We think inflammation may also trigger the production of these proteins.”

    Griffith noted, “When you go against current thinking, you are usually wrong because you are bucking many people who’ve worked so diligently in their fields. But occasionally scientists have failed to put observations from two very distant fields together and that’s what we did. Discovering that telomeres encode two novel signaling proteins will change our understanding of cancer, aging, and how cells communicate with other cells.

    “Many questions remain to be answered, but our biggest priority now is developing a simple blood test for these proteins. This could inform us of our biological age and also provide warnings of issues, such as cancer or inflammation.”

    Source:
    Journal reference:

    Al-Turki, T., et al. (2023) Mammalian Telomeric RNA (TERRA) can be translated to produce valine-arginine and glycine-leucine dipeptide repeat proteins. PNAS. doi.org/10.1073/pnas.2221529120.

    Tracking the global spread of antimicrobial resistance

    An international research team has provided valuable new information about what drives the global spread of genes responsible for antimicrobial resistance (AMR) in bacteria.

    The collaborative study, led by researchers at the Quadram Institute and the University of East Anglia, brought together experts from France, Canada, Germany and the UK and will provide new information to combat the global challenge of AMR.

    By examining the whole genome sequences of around two thousand resistant bacteria, predominantly Escherichia coli collected between 2008 and 2016, the team found that different types of AMR genes varied in their temporal dynamics. For example, some were initially found in North America and spread to Europe, while for others the spread was from Europe to North America.

    Not only did the study look at bacteria from different geographic regions but also from diverse hosts including humans, animals, food (meat) and the environment (wastewater), to define how these separate but interconnected factors influenced the development and spread of AMR. Understanding this interconnectivity embodies the One Health approach and is vital for understanding transmission dynamics and the mechanisms by which resistance genes are transmitted.

    The study, published in the journal Nature Communications, was supported by the Joint Programming Initiative on Antimicrobial Resistance (JPIAMR), a global collaboration spanning 29 countries and the European Commission that is tasked with turning the tide on AMR. Without concerted efforts on a global scale, AMR will undoubtedly make millions more people vulnerable to infections from bacteria and other microorganisms that can currently be tackled with antimicrobials.

    The team focussed on resistance to one particularly important group of antimicrobials, the Extended-Spectrum Cephalosporins (ESCs). These antimicrobials have been classed as critically important by the World Health Organization because they are a ‘last resort’ treatment for multidrug resistant bacteria; despite this, since their introduction, efficacy has declined as bacteria have developed resistance.

    Bacteria that are resistant to ESCs achieve this through the production of specific enzymes, called beta-lactamases, that are able to inactivate ESCs.

    The instructions for making these enzymes are encoded in genes, particularly two key types of genes: extended-spectrum beta-lactamases (ESBLs), and AmpC beta-lactamases (AmpCs).

    These genes may be found on the chromosomes of bacteria where they are passed to progeny during clonal multiplication, or in plasmids, which are small DNA molecules separate to the bacterium’s main chromosome. Plasmids are mobile and can move directly between individual bacteria representing an alternative way of exchanging genetic material.

    This study identified how some resistance genes proliferated through clonal expansion of particularly successful bacterial subtypes while others were transferred directly on epidemic plasmids across different hosts and countries.

    Understanding the flow of genetic information within and between bacterial populations is key to understanding AMR transmission and the global spread of resistance. This knowledge will contribute to the design of vitally needed interventions that can halt AMR in the real world where bacteria from diverse hosts and environmental niches interact, and where international travel and trade mean that these interactions are not limited by geography.

    Professor Alison Mather, group leader at the Quadram Institute and the University of East Anglia, said: “By assembling such a large and diverse collection of genomes, we were able to identify the key genes conferring resistance to these critically important drugs. We were also able to show that the majority of resistance to extended spectrum cephalosporins is spread by only a limited number of predominant plasmids and bacterial lineages; understanding the mechanisms of transmission is key to the design of interventions to reduce the spread of AMR.”

    Lead author Dr Roxana Zamudio said “Antimicrobial resistance is a global problem, and it is only by working collaboratively with partners in multiple countries that we can get a holistic understanding of where and how AMR is spreading.”

    n; the gut and the microbiome; food innovation and population health.

    Story Source:

    Materials provided by University of East Anglia. Note: Content may be edited for style and length.

    Journal Reference:

  • Roxana Zamudio, Patrick Boerlin, Racha Beyrouthy, Jean-Yves Madec, Stefan Schwarz, Michael R. Mulvey, George G. Zhanel, Ashley Cormier, Gabhan Chalmers, Richard Bonnet, Marisa Haenni, Inga Eichhorn, Heike Kaspar, Raquel Garcia-Fierro, James L. N. Wood, Alison E. Mather. Dynamics of extended-spectrum cephalosporin resistance genes in Escherichia coli from Europe and North America. Nature Communications, 2022; 13 (1) DOI: 10.1038/s41467-022-34970-7
  • University of East Anglia