Selected Grantee Publications
Deploying MMEJ using MENdel in Precision Gene Editing Applications for Gene Therapy and Functional Genomics
Martínez-Gálvez et al., Nucleic Acids Research. 2021.
https://academic.oup.com/nar/article/49/1/67/6030233
Gene-editing experiments commonly elicit the error-prone non-homologous end joining for DNA double-strand break (DSB) repair. Martinez-Galvez et al. compared three DSB repair prediction algorithms - MENTHU, inDelphi, and Lindel. MENTHU correctly identified 46% of all PreMAs available, a ∼2- and ∼60-fold sensitivity increase compared to inDelphi and Lindel, respectively. The investigators report the new algorithm MENdel, a combination of MENTHU and Lindel, that achieves the most predictive coverage of homogeneous out-of-frame mutations. They suggest that the use of MENdel helps researchers use MMEJ at scale for reverse genetics screenings to be viable for nearly all loss-of-function based gene editing therapeutic applications. Supported by ORIP (R24OD020166) and NIGMS.
Antibody-Mediated Depletion of Viral Reservoirs is Limited in SIV-Infected Macaques Treated Early With Antiretroviral Therapy
Swanstrom et al., Journal of Clinical Investigation. 2021.
https://doi.org/10.1172/JCI142421
Virus-specific strategies to target the latent HIV reservoir in individuals on combination antiretroviral therapy (cART) have been limited by inefficient induction of viral protein expression. Researchers used rhesus macaques to investigate an antibody-mediated reservoir targeting strategy, targeting the CD4 molecule rather than a viral protein, to deplete potential viral target cells irrespective of infection status. Despite profound CD4+ T cell depletion in blood and lymph nodes, time to viral rebound following cART cessation was not delayed in anti-CD4 treated animals compared with controls, likely due to the limited antibody-mediated cell depletion that occurred in rectal tissue and lymphoid follicles. Supported by ORIP (R24OD010976), NCI, and NIAID.
Lung Expression of Human Angiotensin-Converting Enzyme 2 Sensitizes the Mouse to SARS-CoV-2 Infection
Han et al., American Journal of Respiratory Cell and Molecular Biology. 2021.
https://doi.org/10.1165/rcmb.2020-0354OC
A rapidly deployable mouse model that recapitulates a disease caused by a novel pathogen would be a valuable research tool during a pandemic. Researchers were able to produce C57BL/6J mice with lung expression of human angiotensin-converting enzyme 2 (hACE2), the receptor for SARS-CoV-2. They did so by oropharyngeal delivery of a recombinant human adenovirus type 5 expressing hACE2. The transduced mice were then infected with SARS-CoV-2. Thereafter, the mice developed interstitial pneumonia with perivascular inflammation, exhibited higher viral load in lungs compared to controls, and displayed a gene expression phenotype resembling the clinical response in lungs of humans with COVID-19. Supported by ORIP (P51OD011104, R21OD024931), NHLBI, and NIGMS.
Sequence Diversity Analyses of an Improved Rhesus Macaque Genome Enhance its Biomedical Utility
Warren et al., Science. 2020.
https://science.sciencemag.org/content/370/6523/eabc6617
Investigators sequenced and assembled an Indian-origin female rhesus macaque (RM) genome using a multiplatform genomics approach that included long-read sequencing, extensive manual curation, and experimental validation to generate a new comprehensive annotated reference genome. As a result, 99.7% of the gaps in the earlier draft genome are now closed, and more than 99% of the genes are represented. Whole-genome sequencing of 853 RMs of both sexes identified 85.7 million single-nucleotide variants and 10.5 million indel variants, including potentially damaging variants in genes associated with human autism and developmental delay. The improved assembly of segmental duplications, new lineage-specific genes and expanded gene families provide a framework for developing noninvasive NHP models for human disease, as well as studies of genetic variation and phenotypic consequences. Supported by ORIP (P51OD011106, P51OD011107, P51OD011132, P51OD011104, U42OD024282, U42OD010568, R24OD011173, R24OD021324, R24OD010962), NHGRI, NIMH, NHLBI, and NIGMS.
Imbalance of Regulatory and Cytotoxic SARS-CoV-2-Reactive CD4+ T Cells in COVID-19
Meckiff et al., Cell. 2020.
https://pubmed.ncbi.nlm.nih.gov/33096020/
It is not clear why COVID-19 is deadly in some people and mild in others. To understand the underlying mechanism, investigators studied the contribution of CD4+ T cells in immune responses to SARS-CoV-2 infection. They analyzed single-cell transcriptomic data of >100,000 viral antigen-reactive CD4+ T cells from 40 COVID-19 patients. In hospitalized patients compared to non-hospitalized patients, they found increased proportions of cytotoxic follicular helper cells (TFH) and cytotoxic T helper (TH) cells responding to SARS-CoV-2 and reduced proportion of SARS-CoV-2-reactive regulatory T cells (TREG). Importantly, in hospitalized COVID-19 patients, a strong cytotoxic TFH response was observed early in the illness, which correlated negatively with antibody levels to SARS-CoV-2 spike protein. Polyfunctional TH1 and TH17 cell subsets were underrepresented in the repertoire of SARS-CoV-2-reactive CD4+ T cells compared to influenza-reactive CD4+ T cells. Together, these analyses provided insights into the gene expression patterns of SARS-CoV-2-reactive CD4+ T cells in distinct disease severities. Supported by ORIP (S10RR027366, S10OD025052), NIAID, NHLBI, and NIGMS.
Fructose Stimulated De Novo Lipogenesis Is Promoted by Inflammation
Jelena et al., Nature Metabolism. 2020.
https://pubmed.ncbi.nlm.nih.gov/32839596
Non-alcoholic fatty liver disease (NAFD) affects 30% of adult Americans. While NAFD starts as simple steatosis with little liver damage, its severe manifestation as non-alcoholic steatohepatitis (NASH) is a leading cause of liver failure, cirrhosis, and cancer. Fructose consumption is proposed to increase the risk of hepatosteatosis and NASH. Excessive intake of fructose causes barrier deterioration and low-grade endotoxemia. Using a mouse model, the study examined the mechanism of how fructose triggers these alterations and their roles in hepatosteatosis and NASH pathogenesis. The results demonstrated that microbiota-derived Toll-like receptor (TLR) agonists promote hepatosteatosis without affecting fructose-1-phosphate (F1P) and cytosolic acetyl-CoA. Activation of mucosal-regenerative gp130 signaling, administration of the YAP-induced matricellular protein CCN1 or expression of the antimicrobial peptide Reg3b (beta) counteract fructose-induced barrier deterioration, which depends on endoplasmic-reticulum stress and subsequent endotoxemia. Endotoxin engages TLR4 to trigger TNF production by liver macrophages, thereby inducing lipogenic enzymes that convert F1P and acetyl-CoA to fatty acid in both mouse and human hepatocytes. The finding may be of relevance to several common liver diseases and metabolic disorders. Supported by ORIP (S10OD020025), NCI, NIEHS, NIDDK, NIAID, and NIAAA.
Intra-Strain Genetic Variation of Platyfish (Xiphophorus maculatus) Strains Determines Tumorigenic Trajectory
Lu et al., Frontiers in Genetics . 2020.
https://www.frontiersin.org/articles/10.3389/fgene.2020.562594/full
Xiphophorus interspecies hybrids represent a valuable model system to study heritable tumorigenesis. Although the ancestors of the two X. maculatus parental lines, Jp163 A and Jp163 B, were siblings produced by the same mother, backcross interspecies hybrid progeny between X. hellerii and X. maculatus Jp163 A develop spontaneous melanoma initiating at the dorsal fin due to a regulator encoded by the X. maculatus genome; the backcross hybrid progeny with X. hellerii or X. couchianus and Jp163 B exhibit melanoma on their flanks. Comparative genomic analyses revealed genetic differences are associated with pathways highlighting fundamental cellular functions. Disruption of these baselines may give rise to spontaneous or inducible tumorigenesis. Supported by ORIP (R24OD011120), NCI, and NIGMS.
Induction and Characterization of Pancreatic Cancer in a Transgenic Pig Model
Boas et al., PLOS One. 2020.
https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0239391
Preclinical testing of new therapies for pancreatic cancer has been challenging due to lack of a suitable large animal model. Pigs, however, have similar physiology and immune response to humans. Boas et al report the development of a porcine model for pancreatic cancer. H&E and immunohistochemical stains revealed undifferentiated carcinomas, like those of human pancreatobiliary systems. In several pigs, angiographies revealed that the artery supplying the pancreatic tumor could be catheterized using a 2.4 F microcatheter. In summary, pancreatic cancer can be induced in a transgenic pig, and intra-arterial procedures using catheters designed for human interventions were feasible in this model. Supported by ORIP (U42OD011140) and NCI.