Biology subjects

Patel, N.

Publications and source records attributed to Patel, N..

8 recordsLinked to original sources

bioRxiv2018

Slow delivery immunization enhances HIV neutralizing antibody and germinal center responses via modulation of immunodominance

The observation that humans can produce broadly neutralizing antibodies (bnAbs) against HIV-1 has generated enthusiasm about the potential for a bnAb vaccine against HIV-1. Conventional immunization strategies will likely be insufficient for the development of a bnAb HIV vaccine and vaccines to other difficult pathogens, due to the significant immunological hurdles posed, including B cell immunodominance and germinal center (GC) quantity and quality. Using longitudinal lymph node fine needle aspirates, we found that two independent methods of slow delivery immunization of rhesus macaques (RM) resulted in larger GCs, more robust and sustained GC Tfh cell responses, and GC B cells with improved Env-binding, which correlated with the development of ~20 to 30-fold higher titers of tier 2 HIV-1 nAbs. Using a new RM genomic immunoglobulin loci reference sequence, we identified differential IgV gene usage between slow delivery immunized and conventional bolus immunized animals. The most immunodominant IgV gene used by conventionally immunized animals was associated with many GC B cell lineages. Ab mapping of those GC B cell specificities demonstrated targeting of an immunodominant non-neutralizing trimer base epitope, while that response was muted in slow delivery immunized animals. Thus, alternative immunization strategies appear to enhance nAb development by altering GCs and modulating immunodominance of non-neutralizing epitopes.

immunology↗

bioRxiv2018

SREBP1 drives KRT80-dependent cytoskeletal changes and invasive behavior in endocrine resistant ERα breast cancer

Approximately 30% of women diagnosed with ER breast cancer relapse with metastatic disease following adjuvant treatment with endocrine therapies1,2. The connection between acquisition of drug resistance and invasive potential is poorly understood. In this study, we demonstrate that the type II keratin topological associating domain (TAD)3 undergoes epigenetic reprogramming in cells that develop resistance to aromatase inhibitors (AI), leading to keratin 80 (KRT80) upregulation. In agreement, an increased number of KRT80-positive cells are observed at relapse in vivo while KRT80 expression associates with poor outcome using several clinical endpoints. KRT80 expression is driven by de novo enhancer activation by sterol regulatory element-binding protein 14 (SREBP1). KRT80 upregulation directly promotes cytoskeletal rearrangements at the leading edge, increased focal adhesion maturation and cellular stiffening, which collectively promote cancer cell invasion. Shear-wave elasticity imaging of prospective patients shows that KRT80 levels correlate with stiffer tumors in vivo. Collectively, our data uncover an unpredicted and potentially targetable direct link between epigenetic and cytoskeletal reprogramming promoting cell invasion in response to chronic AI treatment.

cancer biology↗

bioRxiv2018

Identification of rare de novo epigenetic variations in congenital disorders

Certain human traits such as neurodevelopmental disorders (NDs) and congenital anomalies (CAs) are believed to be primarily genetic in origin. With recent dramatic advances in genomic technologies, genome-wide surveys of cohorts of patients with ND/CAs for point mutations and structural variations have greatly advanced our understanding of their genetic etiologies1,2. However, even after whole genome sequencing (WGS), a substantial fraction of such disorders remain unexplained3. In contrast, the possibility that constitutive epigenetic variations (epivariations) might underlie such traits has not been well explored. We hypothesized that some cases of ND/CA are caused by aberrations of DNA methylation that lead to a dysregulation of normal genome function. By comparing DNA methylation profiles from 489 individuals with ND/CAs against 1,534 population controls, we identified epivariations as a frequent occurrence in the human genome. De novo epivariations were significantly enriched in cases when compared to controls. RNAseq data from population studies showed that epivariations often have an impact on gene expression comparable to loss-of-function mutations. Additionally, we detected and replicated an enrichment of rare sequence mutations overlapping CTCF binding sites close to epivariations. Thus, some epivariations occur secondary to cis-linked mutations in regulatory regions, providing a rationale for interpreting non-coding genetic variation. We propose that epivariations likely represent the causative genomic defect in 5-10% of patients with unexplained ND/CAs. This constitutes a yield comparable to CNV microarrays, and as such has significant diagnostic relevance.

genomics↗

bioRxiv2017

Robust identification of deletions in exome and genome sequence data based on clustering of Mendelian errors

Multiple tools have been developed to identify copy number variants (CNVs) from whole exome (WES) and whole genome sequencing (WGS) data. Current tools such as XHMM for WES and CNVnator for WGS identify CNVs based on changes in read depth. For WGS, other methods to identify CNVs include utilizing discordant read pairs and split reads and genome-wide local assembly with tools such as Lumpy and SvABA, respectively. Here, we introduce a new method to identify deletion CNVs from WES and WGS trio data based on the clustering of Mendelian errors (MEs). Using our Mendelian Error Method (MEM), we identified 127 deletions (inherited and de novo) in 2,601 WES trios from the Pediatric Cardiac Genomics Consortium, with a validation rate of 88% by digital droplet PCR. MEM identified additional de novo deletions compared to XHMM, and also identified sample switches, DNA contamination, a significant enrichment of 15q11.2 deletions compared to controls and eight cases of uniparental disomy. We applied MEM to WGS data from the Genome In A Bottle Ashkenazi trio and identified deletions with 97% specificity. MEM provides a robust, computationally inexpensive method for identifying deletions, and an orthogonal approach for verifying deletions called by other tools.

genetics↗

bioRxiv2017

Hsf1 phosphorylation generates cell-to-cell variation in Hsp90 levels and promotes phenotypic plasticity

Clonal populations of cells exhibit cell-to-cell variation in the transcription of individual genes. In addition to this \"noise\" in gene expression, heterogeneity in the proteome and the proteostasis network expands the phenotypic diversity of a population. Heat shock transcription factor (Hsf1) regulates chaperone gene expression, thereby coupling transcriptional noise to proteostasis. Here we show that cell-to-cell variation in Hsf1 activity is an important determinant of phenotypic plasticity. Budding yeast cells with high Hsf1 activity were enriched for the ability to acquire resistance to an antifungal drug, and this enrichment depended on Hsp90 - a known \"phenotypic capacitor\" and canonical Hsf1 target. We show that Hsf1 phosphorylation promotes cell-to-cell variation, and this variation - rather than absolute Hsf1 activity - promotes antifungal resistance. We propose that Hsf1 phosphorylation enables differential tuning of the proteostasis network in individual cells, allowing populations to access a wide range of phenotypic states.

systems biology↗

bioRxiv2017

Hsf1 and Hsp70 constitute a two-component feedback loop that regulates the yeast heat shock response

Models for regulation of the eukaryotic heat shock response typically invoke a negative feedback loop consisting of the transcriptional activator Hsf1 and a molecular chaperone encoded by an Hsf1 target gene. Previously, we identified Hsp70 as the chaperone responsible for Hsf1 repression in Saccharomyces cerevisiae and constructed a mathematical model based on Hsp70-mediated negative feedback that recapitulated the dynamic activity of Hsf1 during heat shock. The model was based on two assumptions: dissociation of Hsp70 activates Hsf1, and transcriptional induction of Hsp70 deactivates Hsf1. Here we validated these assumptions. First, we severed the feedback loop by uncoupling Hsp70 expression from Hsf1 regulation. As predicted by the model, Hsf1 was unable to efficiently deactivate in the absence of Hsp70 transcriptional induction. Next we mapped a discrete Hsp70 binding site on Hsf1 to a motif in the C-terminal activation domain known as conserved element 2 (CE2). Removal of CE2 resulted in increased Hsf1 activity under non-heat shock conditions and delayed deactivation kinetics. In addition, we uncovered a role for the N-terminal domain of Hsf1 in negatively regulating DNA binding. These results reveal the quantitative control mechanisms underlying the feedback loop charged with maintaining cytosolic proteostasis.

biochemistry↗

bioRxiv2017

The effect of a lifestyle intervention in obese pregnant women on change in gestational metabolic profiles: findings form the UK Pregnancies Better Eating and Activity Trial (UPBEAT) RCT.

Background: Pregnancy metabolic disruption is believed to be enhanced in obese women and lead to adverse outcomes in them and their offspring. The UK Pregnancies Better Eating and Activity Trial (UPBEAT), a randomised controlled trial of a lifestyle intervention in obese pregnant women, has already been shown to improve diet and physical activity. We used UPBEAT to determine (a) the magnitude of change in metabolic profiles in obese pregnant women and (b) the impact of a lifestyle intervention that successfully improved diet and physical activity on these profiles. Methods: Detailed targeted metabolic profiling, with quantification of 158 metabolic features (129 lipid measures, 9 glycerides and phospholipids, and 20 low-molecular weight metabolites) was completed on three occasions (~17-, 28- and 35-weeks of gestation) in the UPBEAT participants using NMR. Random intercept and random slope models were used to quantify metabolite changes in obese women using the control (usual care) group only (N = 577). The effect of the intervention was determined by comparing rates of metabolite change between those randomised to intervention and usual care, using intention to treat analyses (N = 1158). Results: There were adverse changes across pregnancy in most lipoprotein subclasses, lipids, glycerides, phospholipids, several fatty acids and glucose. All extremely large, very large, large, medium, small and very small VLDL particles increased by 2 to 3 standard deviation units (SD), with IDL, and large, medium and small LDL particles increasing by 1 to 2SD, between 16- and 36-weeks. Triglycerides increased by 3 to 4SD and glucose increased by 2SD, with more modest changes in other metabolites. The intervention reduced the rate of increase in extremely large, very large, large and medium VLDL, in particular those containing triglycerides. Triglyceride to phosphoglyceride ratio was reduced and there were improvements in fatty acid profiles (increases in the proportion of all fatty acids that were linoleic, omega-6 and polyunsaturated and decreases in the proportion of saturated). Conclusion: Systemic metabolism is markedly disrupted in obese pregnant women, but a lifestyle intervention that improved their diet and physical activity has beneficial effects on some of these profiles; these effects might have long-term benefit. Clinical trial registration ID # ISRCTN89971375.

clinical trials↗

bioRxiv2017

Size uniformity of animal cells is actively maintained by a p38 MAPK-dependent regulation of G1-length

Animal cells within a tissue typically display a striking regularity in their size. To date, the molecular mechanisms that control this uniformity are still unknown. We have previously shown that size uniformity in animal cells is promoted, in part, by size-dependent regulation of G1 length. To identify the molecular mechanisms underlying this process, we performed a large-scale small molecule screen and found that the p38 MAPK pathway is involved in coordinating cell size and cell cycle progression. Small cells display higher p38 activity and spend more time in G1 than larger cells. Inhibition of p38 MAPK leads to loss of the compensatory G1 length extension in small cells, resulting in faster proliferation, smaller cell size and increased size heterogeneity. We propose a model wherein the p38 pathway responds to changes in cell size and regulates G1 exit accordingly, to increase cell size uniformity.\n\nOne-sentence summaryThe p38 MAP kinase pathway coordinates cell growth and cell cycle progression by lengthening G1 in small cells, allowing them more time to grow before their next division.

cell biology↗