The Neural Progenitor Cell-Associated Transcription Factor FoxG1 Regulates Cardiac Epicardial Cell Proliferation.

The epicardium is a layer of mesothelial cells that covers the surface of the heart. During development, epicardial cells undergo epithelial-to-mesenchymal transition (EMT) to form multipotent precursors that migrate into the heart and contribute to the coronary vasculature by differentiating into adventitial fibroblasts, smooth muscle cells, and endothelial cells. Epicardial cells also provide paracrine signals to cardiac myocytes that are required for appropriate heart growth. In adult hearts, a similar process of epicardial cell EMT, migration, and differentiation occurs after myocardial infarction (MI, heart attack). Pathological cardiac hypertrophy is associated with fibrosis, negative remodeling, and reduced cardiac function. In contrast, aerobic exercises such as swimming and running promote physiological (i.e., beneficial) hypertrophy, which is associated with angiogenesis and improved cardiac function. As epicardial cell function(s) during physiological hypertrophy are poorly understood, we analyzed and compared the native epicardial cells isolated directly from the hearts of running-exercised mice and age-matched, nonrunning littermates. To obtain epicardial cells, we enzymatically digested the surfaces of whole hearts and performed magnetic-activated cell sorting (MACS) with antibodies against CD104 (integrin ß4). By cDNA microarray assays, we identified genes with increased transcription in epicardial cells after running exercise; these included FoxG1, a transcription factor that controls neural progenitor cell proliferation during brain development and Snord116, a small noncoding RNA that coordinates expression of genes with epigenetic, circadian, and metabolic functions. In cultured epicardial cells, shRNA-mediated FoxG1 knockdown significantly decreased cell proliferation, as well as Snord116 expression. Our results demonstrate that FoxG1 regulates epicardial proliferation, and suggest it may affect cardiac remodeling.

Determining the impact of vaccination on SARS-CoV-2 RT-PCR cycle threshold values and infectious viral titres.

Background: As the COVID-19 pandemic continues, efforts to better understand severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viral shedding and transmission in both unvaccinated and vaccinated populations remain critical to informing public health policies and vaccine development. The utility of using real time RT-PCR cycle threshold values (CT values) as a proxy for infectious viral litres from individuals infected with SARS-CoV-2 is yet to be fully understood. This retrospective observational cohort study compares quantitative infectious viral litres derived from a focus-forming viral titre assay with SARS-CoV-2 RT-PCR CT values in both unvaccinated and vaccinated individuals infected with the Delta strain. Methods: Nasopharyngeal swabs positive for SARS-CoV-2 by RT-PCR with a CT value <27 collected from 26 June to 17 October 2021 at the University of Vermont Medical Center Clinical Laboratory for which vaccination records were available were included. Partially vaccinated and individuals <18 years of age were excluded. Infectious viral litres were determined using a micro-focus forming assay under BSL-3 containment. Results: In total, 119 specimens from 22 unvaccinated and 97 vaccinated individuals met all inclusion criteria and had sufficient residual volume to undergo viral titring. A negative correlation between RT-PCR CT values and viral litres was observed in both unvaccinated and vaccinated groups. No difference in mean CT value or viral titre was detected between vaccinated and unvaccinated groups. Viral litres did not change as a function of time since vaccination. Conclusions: Our results add to the growing body of knowledge regarding the correlation of SARS-CoV-2 RNA levels and levels of infectious virus. At similar CT values, vaccination does not appear to impact an individual's potential infectivity when infected with the Delta variant.

Left Ventricular Gene Expression in Heart Failure With Preserved Ejection Fraction-Profibrotic and Proinflammatory Pathways and Genes.

BACKGROUND: Heart failure with preserved ejection fraction (HFpEF) is increasingly prevalent and has few treatments. The molecular mechanisms and resultant signaling pathways that underlie the development of HFpEF are poorly defined. It has been proposed that activation of proinflammatory pathways plays a role in the development of cardiac fibrosis. The signature of gene expression (transcriptome) of previously validated left ventricular biopsies obtained from patients with HFpEF and matched referent controls allows for an unbiased assessment of proinflammatory and profibrotic signaling pathways and genes. METHODS: Epicardial left ventricular biopsies from stringently selected HFpEF patients (HFpEF, n=16) and referent control patients (CTR, n=14) were obtained during aortocoronary bypass surgery. The subepicardial myocardium was flash-frozen to build a repository that was parallel-processed for RNA sequencing to allow for an unsupervised in-depth comparison of the left ventricular transcriptome. RESULTS: The average patient age was 67±10 years. When compared with controls, patients with HFpEF were hypertensive with a higher body mass index (kg/m2: 30±5 versus 37±6; P<0.01) and elevated NT-proBNP levels (pg/mL: 155 [89-328] versus 1554 [888-2178]; P<0.001). The transcriptome analysis revealed differential expression of 477 genes many of which were involved in profibrotic pathways including extracellular matrix production and posttranslational modification but no proinflammatory signature. CONCLUSIONS: The transcriptome analysis of left ventricular myocardial samples from patients with HFpEF confirms an overabundant extracellular matrix gene expression, the basis of myocardial fibrosis, without a signature of activated proinflammatory pathways or genes.

Targeting the fatty acid binding proteins disrupts multiple myeloma cell cycle progression and MYC signaling.

Multiple myeloma is an incurable plasma cell malignancy with only a 53% 5-year survival rate. There is a critical need to find new multiple myeloma vulnerabilities and therapeutic avenues. Herein, we identified and explored a novel multiple myeloma target: the fatty acid binding protein (FABP) family. In our work, myeloma cells were treated with FABP inhibitors (BMS3094013 and SBFI-26) and examined in vivo and in vitro for cell cycle state, proliferation, apoptosis, mitochondrial membrane potential, cellular metabolism (oxygen consumption rates and fatty acid oxidation), and DNA methylation properties. Myeloma cell responses to BMS309403, SBFI-26, or both, were also assessed with RNA sequencing (RNA-Seq) and proteomic analysis, and confirmed with western blotting and qRT-PCR. Myeloma cell dependency on FABPs was assessed using the Cancer Dependency Map (DepMap). Finally, MM patient datasets (CoMMpass and GEO) were mined for FABP expression correlations with clinical outcomes. We found that myeloma cells treated with FABPi or with FABP5 knockout (generated via CRISPR/Cas9 editing) exhibited diminished proliferation, increased apoptosis, and metabolic changes in vitro. FABPi had mixed results in vivo, in two pre-clinical MM mouse models, suggesting optimization of in vivo delivery, dosing, or type of FABP inhibitors will be needed before clinical applicability. FABPi negatively impacted mitochondrial respiration and reduced expression of MYC and other key signaling pathways in MM cells in vitro. Clinical data demonstrated worse overall and progression-free survival in patients with high FABP5 expression in tumor cells. Overall, this study establishes the FABP family as a potentially new target in multiple myeloma. In MM cells, FABPs have a multitude of actions and cellular roles that result in the support of myeloma progression. Further research into the FABP family in MM is warrented, especially into the effective translation of targeting these in vivo.

Multiple myeloma is a type of blood cancer for which only a few treatments are available. Currently, only about half the patients with multiple myeloma survive for five years after diagnosis. Because obesity is a risk factor for multiple myeloma, researchers have been studying how fat cells or fatty acids affect multiple myeloma tumor cells to identify new treatment targets. Fatty acid binding proteins (FABPs) are one promising target. The FABPs shuttle fatty acids and help cells communicate. Previous studies linked FABPs to some types of cancer, including another blood cancer called leukemia, and cancers of the prostate and breast. A recent study showed that patients with multiple myeloma, who have high levels of FABP5 in their tumors, have worse outcomes than patients with lower levels. But, so far, no one has studied the effects of inhibiting FABPs in multiple myeloma tumor cells or animals with multiple myeloma. Farrell et al. show that blocking or eliminating FABPs kills myeloma tumor cells and slows their growth in a dish (in vitro) and in some laboratory mice. In the experiments, the researchers treated myeloma cells with drugs that inhibit FABPs or genetically engineered myeloma cells to lack FABPs. They also show that blocking FABPs reduces the activity of a protein called MYC, which promotes tumor cell survival in many types of cancer. It also changed the metabolism of the tumor cell. Finally, the team examined data collected from several sets of patients with multiple myeloma and found that patients with high FABP levels have more aggressive cancer. The experiments lay the groundwork for more studies to determine if drugs or other therapies targeting FABPs could treat multiple myeloma. More research is needed to determine why inhibiting FABPs worked in some mice with multiple myeloma but not others, and whether FABP inhibitors might work better if combined with other cancer therapies. There were no signs that the drugs were toxic in mice, but more studies must prove they are safe and effective before testing the drugs in humans with multiple myeloma. Designing better or more potent FABP-blocking drugs may also lead to better animal study results.

Draft Genome Sequence of a Novel Calicivirus from a Brown Bullhead (Ameiurus nebulosus) from Lake Memphremagog, Vermont/Quebec.

We report a draft genome sequence of a previously undescribed calicivirus from a single brown bullhead inhabiting Lake Memphremagog, Vermont/Quebec. The genome is 7,413 nucleotides long and is most similar to the Atlantic salmon calicivirus (nucleotide identity; 64.7%).

Unique Transcriptomic Changes Underlie Hormonal Interactions During Mammary Histomorphogenesis in Female Pigs.

Successful lactation and the risk for developing breast cancer depend on growth and differentiation of the mammary gland (MG) epithelium that is regulated by ovarian steroids (17ß-estradiol [E] and progesterone [P]) and pituitary-derived prolactin (PRL). Given that the MG of pigs share histomorphogenic features present in the normal human breast, we sought to define the transcriptional responses within the MG of pigs following exposure to all combinations of these hormones. Hormone-ablated female pigs were administered combinations of E, medroxyprogesterone 17-acetate (source of P), and either haloperidol (to induce PRL) or 2-bromo-α-ergocryptine. We subsequently monitored phenotypic changes in the MG including mitosis, receptors for E and P (ESR1 and PGR), level of phosphorylated STAT5 (pSTAT5), and the frequency of terminal ductal lobular unit (TDLU) subtypes; these changes were then associated with all transcriptomic changes. Estrogen altered the expression of approximately 20% of all genes that were mostly associated with mitosis, whereas PRL stimulated elements of fatty acid metabolism and an inflammatory response. Several outcomes, including increased pSTAT5, highlighted the ability of E to enhance PRL action. Regression of transcriptomic changes against several MG phenotypes revealed 1669 genes correlated with proliferation, among which 29 were E inducible. Additional gene expression signatures were associated with TDLU formation and the frequency of ESR1 or PGR. These data provide a link between the hormone-regulated genome and phenome of the MG in a species having a complex histoarchitecture like that in the human breast, and highlight an underexplored synergy between the actions of E and PRL during MG development.

Caught in motion: human NTHL1 undergoes interdomain rearrangement necessary for catalysis.

Base excision repair (BER) is the main pathway protecting cells from the continuous damage to DNA inflicted by reactive oxygen species. BER is initiated by DNA glycosylases, each of which repairs a particular class of base damage. NTHL1, a bifunctional DNA glycosylase, possesses both glycolytic and ß-lytic activities with a preference for oxidized pyrimidine substrates. Defects in human NTHL1 drive a class of polyposis colorectal cancer. We report the first X-ray crystal structure of hNTHL1, revealing an open conformation not previously observed in the bacterial orthologs. In this conformation, the six-helical barrel domain comprising the helix-hairpin-helix (HhH) DNA binding motif is tipped away from the iron sulphur cluster-containing domain, requiring a conformational change to assemble a catalytic site upon DNA binding. We found that the flexibility of hNTHL1 and its ability to adopt an open configuration can be attributed to an interdomain linker. Swapping the human linker sequence for that of Escherichia coli yielded a protein chimera that crystallized in a closed conformation and had a reduced activity on lesion-containing DNA. This large scale interdomain rearrangement during catalysis is unprecedented for a HhH superfamily DNA glycosylase and provides important insight into the molecular mechanism of hNTHL1.

The Kidney-Associated Microbiome of Wild-Caught Artibeus spp. in Grenada, West Indies.

Bats are capable of asymptomatically carrying a diverse number of microorganisms, including human pathogens, due to their unique immune system. Because of the close contact between bats and humans, there is a possibility for interspecies transmission and consequential disease outbreaks. Herein, high-throughput sequencing was used to determine the kidney-associated microbiome of a bat species abundant in Grenada, West Indies, Artibeus spp. Results indicate that the kidney of these bats can carry potential human pathogens. An endogenous retrovirus, Desmodus rotundus endogenous retrovirus isolate 824, phylogenetically related to betaretroviruses from rodents and New World primates, was also identified.

Immunotranscriptomic profiling the acute and clearance phases of a human challenge dengue virus serotype 2 infection model.

About 20-25% of dengue virus (DENV) infections become symptomatic ranging from self-limiting fever to shock. Immune gene expression changes during progression to severe dengue have been documented in hospitalized patients; however, baseline or kinetic information is difficult to standardize in natural infection. Here we profile the host immunotranscriptome response in humans before, during, and after infection with a partially attenuated rDEN2Δ30 challenge virus (ClinicalTrials.gov NCT02021968). Inflammatory genes including type I interferon and viral restriction pathways are induced during DENV2 viremia and return to baseline after viral clearance, while others including myeloid, migratory, humoral, and growth factor immune regulation factors pathways are found at non-baseline levels post-viremia. Furthermore, pre-infection baseline gene expression is useful to predict rDEN2Δ30-induced immune responses and the development of rash. Our results suggest a distinct immunological profile for mild rDEN2Δ30 infection and offer new potential biomarkers for characterizing primary DENV infection.

The Intrinsic Activity of Thyroxine Is Critical for Survival and Growth and Regulates Gene Expression in Neonatal Liver.

Background: Thyroxine (T4) is generally considered to be a prohormone that requires conversion to triiodothyronine (T3) to exert biological activity. Although evidence suggests that T4 has intrinsic activity, it is questionable if this activity has any physiological relevance. Methods: To answer this question, triple knockout (KO) mice (Triples) that cannot express the types 1 (D1) and 2 (D2) deiodinase and the Pax8 genes were generated. Thus, they lack a thyroid and cannot convert T4 to T3. Triples were injected on alternate days with either vehicle or physiological doses of T4, T3, or T3+T4 from postnatal days 2-14. They were euthanized at P15, and RNA-seq was employed to profile gene expression in the liver. In another experiment, Pax8KO mice were injected with T3, T4, or T4+T3, and growth rate and survival to P84 were determined. Results: The growth retardation of Triples was not improved by either T3 or T4 alone but was significantly improved by T4+T3. In the liver, T4 significantly regulated the expression of genes that were also regulated by T3, but the proportion of genes that were negatively regulated was higher in mice treated with T4 than in mice treated with T3. Treatment with T4+T3 identified genes that were regulated synergistically by T3 and T4, and genes that were regulated only by T4+T3. Analysis of these genes revealed enrichment in mechanisms related to cell proliferation and cholesterol physiology, suggesting a unique contribution of T4 to these biological functions. Pax8KO mice all survived to P84 when injected with T4 or T4+T3. However, survival rate with T3 was only 50% and 10% at 3.5 and 12 weeks of life, respectively. Conclusions: T4 has intrinsic activity in vivo and is critical for survival and growth. At a physiological level, T4 per se can upregulate or downregulate many T3 target genes in the neonatal liver. While most of these genes are also regulated by T3, subsets respond exclusively to T4 or demonstrate enhanced or normalized expression only in the presence of both hormones. These studies demonstrate for the first time a complex dependency on both T4 and T3 for normal mammalian growth and development.

Unique Structural Features of Mammalian NEIL2 DNA Glycosylase Prime Its Activity for Diverse DNA Substrates and Environments.

Oxidative damage on DNA arising from both endogenous and exogenous sources can result in base modifications that promote errors in replication as well as generating sites of base loss (abasic sites) that present unique challenges to maintaining genomic integrity. These lesions are excised by DNA glycosylases in the first step of the base excision repair pathway. Here we present the first crystal structure of a NEIL2 glycosylase, an enzyme active on cytosine oxidation products and abasic sites. The structure reveals an unusual "open" conformation not seen in NEIL1 or NEIL3 orthologs. NEIL2 is predicted to adopt a "closed" conformation when bound to its substrate. Combined crystallographic and solution-scattering studies show the enzyme to be conformationally dynamic in a manner distinct among the NEIL glycosylases and provide insight into the unique substrate preference of this enzyme. In addition, we characterized three cancer variants of human NEIL2, namely S140N, G230W, and G303R.

Direct RT-qPCR detection of SARS-CoV-2 RNA from patient nasopharyngeal swabs without an RNA extraction step.

The ongoing COVID-19 pandemic has created an unprecedented need for rapid diagnostic testing. The World Health Organization (WHO) recommends a standard assay that includes an RNA extraction step from a nasopharyngeal (NP) swab followed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) to detect the purified SARS-CoV-2 RNA. The current global shortage of RNA extraction kits has caused a severe bottleneck to COVID-19 testing. The goal of this study was to determine whether SARS-CoV-2 RNA could be detected from NP samples via a direct RT-qPCR assay that omits the RNA extraction step altogether. The direct RT-qPCR approach correctly identified 92% of a reference set of blinded NP samples (n = 155) demonstrated to be positive for SARS-CoV-2 RNA by traditional clinical diagnostic RT-qPCR that included an RNA extraction. Importantly, the direct method had sufficient sensitivity to reliably detect those patients with viral loads that correlate with the presence of infectious virus. Thus, this strategy has the potential to ease supply choke points to substantially expand COVID-19 testing and screening capacity and should be applicable throughout the world.

Selective DNA Demethylation Accompanies T Cell Homeostatic Proliferation and Gene Regulation in Lupus-Prone lpr Mice.

Systemic lupus erythematosus (SLE) is characterized by increased DNA demethylation in T cells, although it is unclear whether this occurs primarily in a subset of SLE T cells. The process driving the DNA demethylation and the consequences on overall gene expression are also poorly understood and whether this represents a secondary consequence of SLE or a primary contributing factor. Lupus-prone lpr mice accumulate large numbers of T cells with age because of a mutation in Fas (CD95). The accumulating T cells include an unusual population of CD4-CD8-TCR-αß+ (DN) T cells that arise from CD8+ precursors and are also found in human SLE. We have previously observed that T cell accumulation in lpr mice is due to dysregulation of T cell homeostatic proliferation, which parallels an increased expression of numerous genes in the DN subset, including several proinflammatory molecules and checkpoint blockers. We thus determined the DNA methylome in lpr DN T cells compared with their CD8+ precursors. Our findings show that DN T cells manifest discrete sites of extensive demethylation throughout the genome, and these sites correspond to the location of a large proportion of the upregulated genes. Thus, dysregulated homeostatic proliferation in lpr mice and consequent epigenetic alterations may be a contributing factor to lupus pathogenesis.

DIRECT RT-qPCR DETECTION OF SARS-CoV-2 RNA FROM PATIENT NASOPHARYNGEAL SWABS WITHOUT AN RNA EXTRACTION STEP.

The ongoing COVID-19 pandemic has caused an unprecedented need for rapid diagnostic testing. The Centers for Disease Control and Prevention (CDC) and the World Health Organization (WHO) recommend a standard assay that includes an RNA extraction step from a nasopharyngeal (NP) swab followed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) to detect the purified SARS-CoV-2 RNA. The current global shortage of RNA extraction kits has caused a severe bottleneck to COVID-19 testing. We hypothesized that SARS-CoV-2 RNA could be detected from NP samples via a direct RT-qPCR assay that omits the RNA extraction step altogether, and tested this hypothesis on a series of blinded clinical samples. The direct RT-qPCR approach correctly identified 92% of NP samples (n = 155) demonstrated to be positive for SARS-CoV-2 RNA by traditional clinical diagnostic RT-qPCR that included an RNA extraction. Thus, direct RT-qPCR could be a front-line approach to identify the substantial majority of COVID-19 patients, reserving a repeat test with RNA extraction for those individuals with high suspicion of infection but an initial negative result. This strategy would drastically ease supply chokepoints of COVID-19 testing and should be applicable throughout the world.

Bioinformatics Core Survey Highlights the Challenges Facing Data Analysis Facilities.

Over the last decade, the cost of -omics data creation has decreased 10-fold, whereas the need for analytical support for those data has increased exponentially. Consequently, bioinformaticians face a second wave of challenges: novel applications of existing approaches (e.g., single-cell RNA sequencing), integration of -omics data sets of differing size and scale (e.g., spatial transcriptomics), as well as novel computational and statistical methods, all of which require more sophisticated pipelines and data management. Nonetheless, bioinformatics cores are often asked to operate under primarily a cost-recovery model, with limited institutional support. Seeing the need to assess bioinformatics core operations, the Association of Biomolecular Resource Facilities Genomics Bioinformatics Research Group conducted a survey to answer questions about staffing, services, financial models, and challenges to better understand the challenges bioinformatics core facilities are currently faced with and will need to address going forward. Of the respondent groups, we chose to focus on the survey data from smaller cores, which made up the majority. Although all cores indicated similar challenges in terms of changing technologies and analysis needs, small cores tended to have the added challenge of funding their operations largely through cost-recovery models with heavy administrative burdens.

Metagenomic analysis of Aedes aegypti and Culex quinquefasciatus mosquitoes from Grenada, West Indies.

The mosquitoes Aedes aegypti (Linnaeus, 1762) (Diptera: Culicidae) and Culex quinquefasciatus Say, 1823 (Diptera: Culicidae) are two major vectors of arthropod-borne pathogens in Grenada, West Indies. As conventional vector control methods present many challenges, alternatives are urgently needed. Manipulation of mosquito microbiota is emerging as a field for the development of vector control strategies. Critical to this vector control approach is knowledge of the microbiota of these mosquitoes and finding candidate microorganisms that are common to the vectors with properties that could be used in microbiota modification studies. Results showed that bacteria genera including Asaia, Escherichia, Pantoea, Pseudomonas, and Serratia are common to both major arboviral vectors in Grenada and have previously been shown to be good candidates for transgenetic studies. Also, for the first time, the presence of Grenada mosquito rhabdovirus 1 is reported in C. quinquefasciatus.

Critical Role for SLAM/SAP Signaling in the Thymic Developmental Programming of IL-17- and IFN-γ-Producing γδ T Cells.

During thymic development, mouse γδ T cells commit to either an IFN-γ- or an IL-17-producing phenotype through mechanisms that remain unclear. In this study, we investigated the extent to which the SLAM/SAP signaling pathway regulates the functional programming of γδ T cells. Characterization of SLAM family receptor expression revealed that thymic γδ T cell subsets were each marked by distinct coexpression profiles of SLAMF1, SLAMF4, and SLAMF6. In the thymus, Vγ1 and Vγ4 T cells that exhibited an SLAMF1+SLAMF6+ double positive phenotype were largely contained within immature CD24+CD73- and CD24+CD73+ subsets, whereas SLAMF1 single positive, SLAMF6 single positive, or SLAMF1SLAMF6 double negative cells were found within mature CD24-CD73+ and CD24-CD73- subsets. In the periphery, SLAMF1 and SLAMF6 expression distinguished IL-17- and IFN-γ-producing γδ T cells, respectively. Disruption of SLAM family receptor signaling through deletion of SAP resulted in impaired thymic Vγ1 and Vγ4 T cell maturation at the CD24+CD73-SLAMF1+SLAMF6+ double positive stage that was associated with a decreased frequency of CD44+RORγt+ γδ T cells. Impaired development was in turn associated with decreased γδ T cell IL-17 and IFN-γ production in the thymus as well as in peripheral tissues. The role for SAP was subset-specific, as Vγ1Vδ6.3, Vγ4, Vγ5, but not Vγ6 subsets were SAP-dependent. Together, these data suggest that the SLAM/SAP signaling pathway plays a larger role in γδ T cell development than previously appreciated and represents a critical checkpoint in the functional programming of both IL-17- and IFN-γ-producing γδ T cell subsets.

Life and its traces in Antarctica's McMurdo Dry Valley paleolakes: a survey of preservation.

The extremely cold and arid conditions of Antarctica make it uniquely positioned to investigate fundamental questions regarding the persistence of life in extreme environments. Within the McMurdo Dry Valleys and surrounding mountain ranges are multiple ancient relict lakes, paleolakes, with lacustrine deposits spanning from thousands to millions of years in age. Here we present data from light microscopy, scanning electron microscopy, electron dispersive x-ray spectroscopy, and radiocarbon dating to catalog the remarkable range of life preserved within these deposits. This includes intact microbes and nanobacteria-sized cocci, CaCO3 precipitations consistent with biogenic calcium, previously undescribed net-like structures, possible dormant spores, and long-extinct yet exquisitely preserved non-vascular plants. These images provide an important reference for further microbiome investigations of Antarctic paleolake samples. In addition, these findings may provide a visual reference for the use of subsurface groundwater microbial communities as an analog for paleolake subsurface water on planets such as Mars.

A Review of the Scientific Rigor, Reproducibility, and Transparency Studies Conducted by the ABRF Research Groups.

Shared research resource facilities, also known as core laboratories (Cores), are responsible for generating a significant and growing portion of the research data in academic biomedical research institutions. Cores represent a central repository for institutional knowledge management, with deep expertise in the strengths and limitations of technology and its applications. They inherently support transparency and scientific reproducibility by protecting against cognitive bias in research design and data analysis, and they have institutional responsibility for the conduct of research (research ethics, regulatory compliance, and financial accountability) performed in their Cores. The Association of Biomolecular Resource Facilities (ABRF) is a FASEB-member scientific society whose members are scientists and administrators that manage or support Cores. The ABRF Research Groups (RGs), representing expertise for an array of cutting-edge and established technology platforms, perform multicenter research studies to determine and communicate best practices and community-based standards. This review provides a summary of the contributions of the ABRF RGs to promote scientific rigor and reproducibility in Cores from the published literature, ABRF meetings, and ABRF RGs communications.

The NEIL1 G83D germline DNA glycosylase variant induces genomic instability and cellular transformation.

Base excision repair (BER) is a key genome maintenance pathway. The NEIL1 DNA glycosylase recognizes oxidized bases, and likely removes damage in advance of the replication fork. The rs5745906 SNP of the NEIL1 gene is a rare human germline variant that encodes the NEIL1 G83D protein, which is devoid of DNA glycosylase activity. Here we show that expression of G83D NEIL1 in MCF10A immortalized but non-transformed mammary epithelial cells leads to replication fork stress. Upon treatment with hydrogen peroxide, we observe increased levels of stalled replication forks in cells expressing G83D NEIL1 versus cells expressing the wild-type (WT) protein. Double-strand breaks (DSBs) arise in G83D-expressing cells during the S and G2/M phases of the cell cycle. Interestingly, these breaks result in genomic instability in the form of high levels of chromosomal aberrations and micronuclei. Cells expressing G83D also grow in an anchorage independent manner, suggesting that the genomic instability results in a carcinogenic phenotype. Our results are consistent with the idea that an inability to remove oxidative damage in an efficient manner at the replication fork leads to genomic instability and mutagenesis. We suggest that individuals who harbor the G83D NEIL1 variant face an increased risk for human cancer.