Biology subjects

Shah, K.

Publications and source records attributed to Shah, K..

At least 37 records · Page 2Linked to original sources

bioRxiv2022

Validation of saline, PBS and a locally produced VTM at varying storage conditions to detect the SARS-CoV-2 virus by qRT-PCR

Coronavirus Disease-2019 tests require a Nasopharyngeal (NP) and/or Oropharyngeal (OP) specimen from the upper airway, from which virus RNA is extracted and detected through quantitative reverse transcription-Polymerase Chain Reaction (qRT-PCR). The viability of the virus is maintained after collection by storing the NP/OP swabs in Viral Transport Media (VTM). We evaluated the performance of four transport media: locally manufactured ("REVITAL") Viral Transport Media (RVTM), Standard Universal Transport Media (SUTM), PBS and 0.9% (w/v) NaCl (normal saline). We used laboratory cultured virus to evaluate: i) viral recovery and maintaining integrity at different time periods and temperatures; ii) stability in yielding detectable RNA consistently for all time points and conditions; and iii) their overall accuracy. Four vials of SARS-CoV-2 cultured virus (2 high and 2 low concentration samples) and 1 negative control sample were prepared for each media type (SUTM, RVTM, PBS and normal saline) and stored at the following temperatures, -80{degrees}C, 4{degrees}C, room temperature (25{degrees}C) and 37{degrees}C for 7 days. Viral Ribonucleic acid (RNA) extractions and qRT-PCR were done on the following days after inoculation with the cultured virus, days 1, 2, 3, 4 and 7 to assess virus stability and viral recovery. CT values fell over time at room temperature, but normal saline, PBS, RVTM and SUTM all showed comparable performance in maintaining virus integrity and stability allowing for the detection of SARS-CoV-2 viral RNA. Overall, this study demonstrated that normal saline, PBS and the locally manufactured VTM can be used for COVID-19 sample collection and testing, thus expanding the range of SARS-CoV-2 viral collection media.

molecular biology↗

bioRxiv2022

S-nitrosoglutathione Inhibits the Growth of Androgen-Receptor Dependent and Castration Resistant Prostate Cancer by Modulating FOXM1 Signaling

Castration resistant prostate cancer (CRPC) is an advanced form of prostate cancer associated with loss of androgen receptor activity and resistance to AR-targeted therapies. CRPC is often associated with higher metastasis, invasion, and proliferation. Thus, theres a strong need for additional molecular markers in CRPC, and therapies that target them directly. Transcriptomic profiling of prostate cancer patients revealed an upregulated signature specific to high-grade tumors in cell cycle progression, E2F targets, G2M checkpoint, DNA repair, Aurora Kinase B, and FOXM1 pathway. Validation in a separate cohort of neuroendocrine prostate cancer patients, the most resistant and invasive form of CRPC which lacks AR expression, showed CCP signature as a dominant driver gene set for NEPC. Moreover, high expression of mRNA targets in each of these pathways is associated with poor overall survival in NEPC patients. Gene set variation analysis revealed a strong inverse correlation between AR signaling and CCP.31, set of 31 markers associated with high proliferation, and a positive correlation between CCP.31 and NEPC signature. FOXM1, a prioritized candidate in CCP is shown to be upregulated in CRPC cell lines and human clinical sections of Gleason 9 patients. GSNO, a nitric oxide donor, known for its inhibitory effect on cell proliferation and an effective inhibitor of cell cycle checkpoints, was able to reverse CCP signature both in-vitro and in-vivo. GSNO exhibits a dependence on AR signaling in-vitro by showing mild effect on CRPC clones, but successfully inhibits tumor growth of AR-null CRPC xenografts. FOXM1 is also targeted by GSNO in-vivo and in-vitro models of both CRPC and AD prostate cancer. This establishes an important link between androgen receptor status in prostate cancer and proposes nitric oxide donors as therapeutic intervention. GSNO is effective against AR-null castration resistant prostate cancer. SignificanceNitric oxide donor, S-glutathione-transferase inhibits tumor growth of castration resistant prostate cancer, regardless of androgen receptor expression status by targeting FOXM1, a critical regulator of cell cycle progression. O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

cancer biology↗

bioRxiv2022

Novel class of yellow emitting carbon dots stimulate collective cell migration and 3D uptake in vivo

We present a new class of nitrogen-doped yellow fluorescent carbon dots, synthesized using a one-step hydrothermal method. These bright fluorescent nanoparticles have excitation and emission spectra near the red region of the visible light spectrum that are quite useful for bioimaging applications. Using organic molecules like ortho- phenylenediamine (OPDA), L-ascorbic acid and urea, yellow fluorescent carbon dots (CDs) were synthesized. We obtained a scalable number of CDs having an average size of 3 nm. The CDs show significant emission spectra in the yellow fluorescence region ({lambda}em= 557 nm). The CDs show remarkable stability in their fluorescence in different pH conditions, ionic stability, photostability as well as thermal stability. These CDs are efficiently uptaken by mammalian cells through clathrin-mediated pathway. Apart from in vitro studies we have also used zebrafish larvae as a 3D in vivo model, and showed that CDs were uptaken efficiently by larvae showing maximum accumulation and fluorescence in the yolk sac region and the notochord region. The CDs also offer enhancement in cell proliferation, hence showing the application in wound healing. The fluorescence of CDs is quite robust and is not affected by most external stimuli, hence can be explored as a promising bioimaging tool for targeted bioimaging and biomedical applications.

bioengineering↗

bioRxiv2022

Green emitting carbon quantum dots (GCQDs) to probe endocytic pathways in cells; for tissue and in vivo bioimaging

Small sized, carbon-based organic nanoparticles have recently gained attention due their advantage of biocompatibility, photostability and biological non-toxicity as compared to their inorganic counterparts. Herein, a new class of small (5-8 nm), green emitting fluorescent carbon quantum dots (GCQDs) were synthesized using organic substrates like citric acid and ascorbic acid in aqueous solvent containing water and ethanol. The very small size and bright green photoluminescence prompted their use for both in vitro and in vivo bioimaging. GCQDs were uptaken via clathrin mediated pathways in mouse kidney and liver primary cells. Similarly, they showed active uptake and distribution in the zebrafish embryo model system. The optical tunability and surface modification properties of these GCQDs provide a platform to be explored for them to emerge as a new class of targeted bioimaging entities, as well as tools for biomedical applications.

bioengineering↗

bioRxiv2022

Phosphorylation of the novel mTOR substrate Unkempt regulates cellular morphogenesis

Mechanistic target of rapamycin (mTOR) is a protein kinase that integrates multiple inputs to regulate anabolic cellular processes. mTOR complex I (mTORC1) has key functions in growth control, autophagy and metabolism. Much less is known about the signalling components that act downstream of mTORC1 that regulate cellular morphology, a vital determinant of cellular function. Here we show that the RNA-binding protein Unkempt, a key regulator of cellular morphogenesis, is a novel substrate mTORC1. We find that Unkempt phosphorylation is regulated by nutrient levels and growth factors via mTORC1. Furthermore, Unkempt physically interacts with and is directly phosphorylated by mTORC1 through binding to the regulatory-associated protein of mTOR, Raptor. Phosphorylation of Unkempt, which we find is mTORC1-dependent in cultured mammalian cell lines as well as in primary tissues, occurs largely within the highly serine-rich intrinsically disordered region of Unkempt. Importantly, mutation analysis of this region indicates that phosphorylation inhibits the ability of Unkempt to induce a bipolar morphology. Our findings reveal a novel molecular link between mTORC1 signalling and cellular morphogenesis.

neuroscience↗

bioRxiv2021

Decrowding Expansion Pathology: Unmasking Previously Invisible Nanostructures and Cells in Intact Human Brain Pathology Specimens

Proteins are densely packed in cells and tissues, where they form complex nanostructures. Expansion microscopy (ExM) variants have been used to separate proteins from each other in preserved biospecimens, improving antibody access to epitopes. Here we present an ExM variant, decrowding expansion pathology (dExPath), which can expand proteins away from each other in human brain pathology specimens, including formalin-fixed paraffin-embedded (FFPE) clinical specimens. Immunostaining of dExPath-expanded specimens reveals, with nanoscale precision, previously unobserved cellular structures, as well as more continuous patterns of staining. This enhanced molecular staining results in observation of previously invisible disease marker-positive cell populations in human glioma specimens, with potential implications for tumor aggressiveness. dExPath results in improved fluorescence signals even as it eliminates lipofuscin-associated autofluorescence. Thus, this form of expansion-mediated protein decrowding may, through improved epitope access for antibodies, render immunohistochemistry more powerful in clinical science and diagnosis.

bioengineering↗

bioRxiv2021

Genetic analysis of Caenorhabditis elegans pry-1/Axin suppressors identifies genes involved in reproductive structure development, stress response, and aging

The Axin family of scaffolding proteins regulates a wide array of developmental and post-developmental processes in eukaryotes. Studies in the nematode, Caenorhabditis elegans, have shown that the Axin homolog, PRY-1, plays essential roles in multiple tissues. To understand the genetic network of pry-1, we focused on a set of genes that are differentially expressed in the pry-1-mutant transcriptome and are linked to reproductive structure development. Eight of the genes (ard-1, rpn-7, cpz-1, his-7, cdk-1, rnr-1, clsp-1, and spp-1), when knocked down by RNA interference, efficiently suppressed the plate-level multivulva phenotype of pry-1 mutants. In every case, other than clsp-1 and spp-1, the ectopic vulval precursor cell (VPC) induction was also inhibited. The suppressor genes are members of known gene families in eukaryotes and perform essential functions. Our genetic interaction experiments revealed that except for clsp-1, the genes participate in one or more pry-1-mediated biological events. While four of them (cpz-1, his-7, cdk-1, and rnr-1) function in VPC induction, stress response, and aging, the other three (spp-1, ard-1, and rpn-7) are specific to one or more of these processes. Further analysis of the genes involved in aging showed that his-7, cdk-1, and rnr-1 also interacted with daf-16/FOXO. The results of genetic epistasis experiments suggested that his-7 functions upstream of daf-16, whereas cdk-1 and rnr-1 act downstream of the pry-1-daf-16 pathway. Altogether, these findings demonstrate the important role of pry-1 suppressors in C. elegans. Given that all of the genes described in this study are conserved, future investigations of their interactions with Axin and their functional specificity promises to uncover the genetic network of Axin under normal and disease states.

genetics↗

bioRxiv2021

Small Intestinal Resident Eosinophils Maintain Gut Homeostasis Following Microbial Colonisation

Intestinal homeostasis following postnatal microbial colonization requires the coordination of multiple processes including the activation of immune cells, cell-cell communication, the controlled deposition of extracellular matrix, and epithelial cell turnover and differentiation. The intestine harbors the largest frequency of resident eosinophils of all homeostatic organs, yet the functional significance of eosinophil recruitment to this organ has long remained enigmatic. Eosinophils are equipped to both respond to, and modify, their local tissue environment and thus are able to regulate the adaption of tissues to environmental changes. We report a critical role for eosinophils in regulating villous structure, barrier integrity and motility in the small intestine. Notably, the microbiota was identified as a key driver of small intestinal eosinophil activation and function. Collectively our findings demonstrate a critical role for eosinophils in facilitating mutualistic interactions between host and microbiota and provide a rationale for the functional significance of their early life recruitment in the small intestine. HIGHLIGHTSO_LIThe microbiota is a critical regulator of eosinophil activation and turnover C_LIO_LIEosinophils uphold intestinal barrier integrity following microbial colonization C_LIO_LILoss of eosinophils at steady-state results in increased villous blunting and altered intestinal motility C_LI

immunology↗

bioRxiv2020

NOVEL NUCLEAR ROLE OF HDAC6 IN PROGNOSIS AND THERAPEUTIC TARGET FOR COLORECTAL CANCER

Histone deacetylase 6 (HDAC6) inhibition is a potential treatment of a wide range of cancer types via the acetylation of diverse proteins in the cytoplasm. However, the regulation of histone acetylation and the maintenance of higher-order chromatin structure remains unidentified. Here, we investigated the effect of selective inhibition of HDAC6 by histone acetylation, chromatin relaxation assays, co-immunoprecipitation, acetylome peptide array and in vivo RNA microarray. Our data shows that nuclear HDAC6 physically interacts with the Histone 4 lysine 12 residue, and that HDAC6 inhibition increases acetylation specifically at this residue in several cancer types. Inhibition induces major chromatin structure modulation, but has no equivalent effect on knockout HDAC6-/- MEF cells. We identified several novel HDAC6-deacetylated substrates and high expression of HDAC6 in colorectal cancer (CRC) tissue association with reduced levels of H4K12ac and independent of the key CRC driver mutations, but positively associated with EGFR expression. Furthermore, in vivo HDAC6 inhibition induces significant tumor regression in a CRC xenograft mice model with significant changes in the expression of functional nuclear genes. We also demonstrated that a DNA damaging agent in combination with selective HDAC6 inhibition is effective and acts synergistically, inducing chromatin relaxation and increased cell death in CRC cells. CRC tissues (Normal versus tumor; n=58 matched pairs) together with TCGA data analysis of 467 CRC patients showed that high HDAC6 expression is associated with metastasis, overall and disease-free survival, and is an independent risk factor of CRC stage progression. Our findings designate a new role for nuclear HDAC6 both in cancer prognosis and as a new therapeutic target for CRC and other types of cancer. HighlightHistone deacetylases 6 activity; Chromatin relaxation; Histone modifications; Gene array; DOX: doxorubicin; OXA: oxaliplatin; 5-FU: fluorouracil; Ac: acetylation; MNase: Micrococal nuclease.

cancer biology↗

bioRxiv2020

Mutations in fbiD(Rv2983) as a novel determinant of resistance to pretomanid and delamanid in Mycobacterium tuberculosis

The nitroimidazole pro-drugs delamanid and pretomanid comprise one of only two new antimicrobial classes approved to treat tuberculosis (TB) in 50 years. Prior in vitro studies suggest a relatively low barrier to nitroimidazole resistance in Mycobacterium tuberculosis, but clinical evidence is limited to date. We selected pretomanid-resistant M. tuberculosis mutants in two mouse models of TB using a range of pretomanid doses. The frequency of spontaneous resistance was approximately 10-5 CFU. Whole genome sequencing of 161 resistant isolates from 47 mice revealed 99 unique mutations, 91% of which occurred in 1 of 5 genes previously associated with nitroimidazole activation and resistance: fbiC (56%), fbiA (15%), ddn (12%), fgd (4%) and fbiB (4%). Nearly all mutations were unique to a single mouse and not previously identified. The remaining 9% of resistant mutants harbored mutations in Rv2983, a gene not previously associated with nitroimidazole resistance but recently shown to be a guanylyltransferase necessary for cofactor F420 synthesis. Most mutants exhibited high-level resistance to pretomanid and delamanid, although Rv2983 and fbiB mutants exhibited high-level pretomanid resistance, but relatively small changes in delamanid susceptibility. Complementing an Rv2983 mutant with wild-type Rv2983 restored susceptibility to pretomanid and delamanid. By quantifying intracellular F420 and its precursor Fo in overexpressing and loss-of-function mutants, we provide further evidence that Rv2983 is necessary for F420 biosynthesis. Finally, Rv2983 mutants and other F420H2-deficient mutants displayed hypersusceptibility to some antibiotics and to concentrations of malachite green found in solid media used to isolate and propagate mycobacteria from clinical samples.

microbiology↗

bioRxiv2020

BAP1 and YY1 regulate expression of death receptors in malignant pleural mesothelioma

Malignant pleural mesothelioma (MPM) is a rare, aggressive, and incurable cancer arising from the mesothelial lining of the lungs with few treatment options. We recently reported loss of function of the nuclear deubiquitinase BRCA associated protein-1 (BAP1), a frequent event in MPM, is associated with sensitivity to tumour necrosis factor-related apoptosis-inducing ligand (TRAIL). As a potential underlying mechanism, here we report that BAP1 negatively regulates the expression of TRAIL receptors: death receptors 4 (DR4) and 5 (DR5). Using tissue microarray (TMAs) of tumour samples from MPM patients, we found a strong inverse correlation between BAP1 and TRAIL receptors. BAP1 knockdown increased DR4 and DR5 expression, whereas overexpression of BAP1 had the opposite effect. Reporter assays confirmed wild-type BAP1, but not catalytically-inactive mutant BAP1, reduced promoter activities of DR4 and DR5, suggesting deubiquinase activity plays an important role in the regulation of gene expression. Co-IP studies demonstrated direct binding of BAP1 and the transcription factor Ying Yang 1 (YY1) and ChIP assays revealed BAP1 and YY1 to be enriched in the promoter regions of DR4 and DR5. Notably, shRNA knockdown of YY1 also increased DR4 and DR5 expression, and sensitivity to TRAIL. These results demonstrate that BAP1 and YY1 together negatively regulate transcriptional activity of TRAIL receptors. BAP1 and YY1 may both therefore be strong therapeutic targets to enhance the efficacy of TRAIL-induced apoptosis. Statement of significanceWe describe how the most-frequently mutated tumour suppressor gene in mesothelioma regulates the response to TNF-related apoptosis-inducing ligand (TRAIL). These findings will accelerate a biomarker-driven cancer therapy.

cancer biology↗

bioRxiv2020

Optimization and scaling of patient-derived brain organoids uncovers deep phenotypes of disease

Cerebral organoids provide unparalleled access to human brain development in vitro. However, variability induced by current culture methodologies precludes using organoids as robust disease models. To address this, we developed an automated Organoid Culture and Assay (ORCA) system to support longitudinal unbiased phenotyping of organoids at scale across multiple patient lines. We then characterized organoid variability using novel machine learning methods and found that the contribution of donor, clone, and batch is significant and remarkably consistent over gene expression, morphology, and cell-type composition. Next, we performed multi-factorial protocol optimization, producing a directed forebrain protocol compatible with 96-well culture that exhibits low variability while preserving tissue complexity. Finally, we used ORCA to study tuberous sclerosis, a disease with known genetics but poorly representative animal models. For the first time, we report highly reproducible early morphological and molecular signatures of disease in heterozygous TSC+/- forebrain organoids, demonstrating the benefit of a scaled organoid system for phenotype discovery in human disease models.

neuroscience↗

bioRxiv2020

Transient delivery of A-C/EBP protein perturbs differentiation of 3T3-L1 cells and induces preadipocyte marker genes

Transformation of committed 3T3-L1 preadipocytes to lipid-laden adipocytes involves timely appearance of numerous transcription factors (TFs), foremost among them C/EBP{beta}, is expressed during early phases of differentiation. Here we describe liposome-mediated protein transfection approach to rapidly downregulate C/EBP{beta} by A-C/EBP protein inhibitor. Signals from tagged A-C/EBP were observed in 3T3-L1 cells within 2hrs of protein inhibitor transfections whereas for gene transfection signals appeared in 48hrs. Following transient transfections, expression profiles of 24 marker genes belonging to pro- and anti-adipogenic, cell cycle, and preadipocytes pathways was analyzed. mRNA and protein expression profiles of adipocyte-marker genes showed lower expression in both A-C/EBP protein and gene transfected samples. Interestingly, for preadipocytes and cell fate determinant genes, striking differences were observed between protein and gene transfected samples. Preadipocyte differentiation factors Stat5a and Creb were downregulated in A-C/EBP protein samples. Five preadipocyte markers, namely, Pdgfr, Pdgfr{beta}, Ly6A, CD34 Itgb1 showed high expression in protein samples whereas only Ly6A and CD34 were expressed in gene transfected samples. Pdgfr and Pdgfr{beta}, two known cell fate markers were expressed in protein transfected samples 5-days post-differentiation suggesting a possible reversal of differentiation. Our study provides evidences for the robust and efficient knockdown of C/EBP{beta} protein to understand time-dependent gene regulation during adipogenesis.

molecular biology↗

bioRxiv2020

Super-human cell death detection with biomarker-optimized neural networks

Cell death is an essential process in biology that must be accounted for in live microscopy experiments. Nevertheless, cell death is difficult to detect without perturbing experiments with stains, dyes or biosensors that can bias experimental outcomes, lead to inconsistent results, and reduce the number of processes that can be simultaneously labelled. These additional steps also make live microscopy difficult to scale for high-throughput screening because of the cost, labor, and analysis they entail. We address this fundamental limitation of live microscopy with biomarker-optimized convolutional neural networks (BO-CNN): computer vision models trained with a ground truth biosensor that detect live cells with superhuman, 96% accuracy more than 100 times faster than previous methods. Our models learn to identify important morphological characteristics associated with cell vitality without human input or additional perturbations, and to generalize to other imaging modalities and cell types for which they have no specialized training. We demonstrate that we can interpret decisions from BO-CNN models to gain biological insight into the patterns they use to achieve superhuman accuracy. The BO-CNN approach is broadly useful for live microscopy, and affords a powerful new paradigm for advancing the state of high-throughput imaging in a variety of contexts.

cell biology↗

bioRxiv2020

Neurons from human mesenchymal stem cells display both spontaneous and stimuli responsive activity

Mesenchymal stem cells are one of the promising tissue specific stem cell source for neural tissue regeneration applications. Previous studies on human mesenchymal stem cell (hMSC) derived neurons have been limited and not statisfactory in terms of neuronal activity. In this study, we analysed the functionality of bone marrow hMSCs differentiated into neural protein expressing cells by a single step cytokine based induction protocol. Neurons from both primary hMSCs and hMSC cell line displayed spontaneous activity ([≥]75%) as demonstrated by Ca++ imaging. Furthermore, when electrically stimulated, hMSC induced neurons (hMd-Neuron) matched the response of a typical neuron in the process of maturation. Our results reveal that enriched neurothrophic factors enhance differentiation capacity of bone marrow hMSCs into high yielding functional neurons with spontaneous activity and mature into electrophysiologically active state. hMd-Neurons have the potential to be used as a tool for disease modelling of neuropathologies and neural differentiation studies.

neuroscience↗

bioRxiv2019

Global kinome silencing combined with 3D invasion screening of the tumor microenvironment identifies fibroblast-expressed PIK3Cδ involvement in triple-negative breast cancer progression

As there is growing evidence for the tumor microenvironments (TME) role in tumorigenesis, we sought to investigate the role of fibroblast-expressed kinases in triple negative breast cancer (TNBC). Using a high-throughput kinome screen combined with 3D invasion assays, we identified fibroblast-expressed PIK3C{delta} (f-PIK3C{delta}) as a key regulator of progression. Although PIK3C{delta} has been mainly described in leucocytes, we detected high expression in primary fibroblasts derived from TNBC patients, while PIK3C{delta} was undetectable in cancer epithelial cell lines. Genetic and pharmacologic gain- and loss-of functions experiments verified the contribution of f-PIK3C{delta} in TNBC cell invasion. By employing an integrated secretomics and transcriptomics analysis, we revealed a paracrine mechanism via which f-PIK3C{delta} confers its pro-tumorigenic effects. Inhibition of f-PIK3C{delta} promoted the secretion of factors, including PLGF and BDNF, which subsequently led to upregulation of NR4A1 in TNBC cells where it acts as a tumor suppressor. Inhibition of PIK3C{delta} in an orthotopic BC mouse model reduced tumor growth only after inoculation with fibroblasts, indicating a role of f-PIK3C{delta} in cancer progression. Similar results were observed in the MMTV-PyMT transgenic BC mouse model, in addition to a decrease on tumor metastasis emphasizing the potential immune-independent effects of PIK3C{delta} inhibition. Finally, analysis of BC patient cohorts and TCGA datasets identified f-PIK3C{delta} (protein and mRNA levels) as an independent prognostic factor for overall and disease free survival, highlighting it as a therapeutic target for TNBC.

cancer biology↗

bioRxiv2019

Resolvin D-series regulates Phospholipase D both during inflammation and resolution by modulating phagocyte functions

A successful acute inflammatory response results in the elimination of infectious agents by neutrophils and monocytes, followed by resolution and repair by tissue-resident and recruited macrophages. D-series resolvins are pro-resolving mediators involved in resolution and tissue repair, their intracellular signaling mechanism(s) are of interest. Here, we report that D-series resolvins activate phospholipase D (PLD), a ubiquitously expressed membrane lipase in modulating phagocyte functions. The mechanism for PLD-mediated actions of RvD5 in polarizing macrophages (M1-M2) was found to be two-pronged: (a) RvD5 inhibits post-transcriptional modifications, by miRs and 3exonulceases that process PLD2 mRNA, thus increasing PLD2 expression and activity; (b) RvD5 enhances PLD2-S6K signaling and Actin expression required for membrane expansion and efferocytosis. In addition, investigating RvD5s actions in second organ reflow injury, we found that RvD5 did not reduce lung neutrophil myeloperoxidase levels in PLD2-/- mice compared to WT and PLD1-/- mice, pointing to a unique role of PLD2 as the relevant isoform in RvD5-mediated resolution. These results demonstrate that RvD5-PLD2 are attractive targets for therapeutic interventions in vascular inflammation such as I/R injury and cardiovascular diseases.

immunology↗

bioRxiv2019

Androgen Receptor Signaling Regulates the Transcriptome of Prostate Cancer Cells by Modulating Global Alternative Splicing

Androgen receptor (AR), is a transcription factor and a member of a hormone receptor superfamily. AR plays a vital role in the progression of prostate cancer and is a crucial target for therapeutic interventions. While the majority of advanced-stage prostate cancer patients will initially respond to the androgen-deprivation, the disease often progresses to castrate-resistant prostate cancer (CRPC). Interestingly, CRPC tumors continue to depend on hyperactive AR signaling and will respond to potent second-line anti-androgen therapies, including bicalutamide (CASODEX(R)) and enzalutamide (XTANDI(R)). However, the progression-free survival rate for the CRPC patients on anti-androgen therapies is only 8 to 19 months. Hence, there is a need to understand the mechanisms underlying CRPC progression and eventual treatment resistance. Here, we have leveraged next-generation sequencing and newly developed analytical methodologies to evaluate the role of AR-signaling in regulating the transcriptome of prostate cancer cells. The genomic and pharmacologic stimulation- and inhibition-of AR activity demonstrates that AR regulates alternative splicing within cancer-relevant genes. Furthermore, by integrating transcriptomic data from in vitro experiments and in prostate cancer patients, we found that a significant number of AR-regulated splicing events are associated with tumor progression. For example, we found evidence for an inadvertent AR-antagonist mediated switch in IDH1 and PL2G2A isoform expression, which is associated with a decrease in overall survival of patients. Mechanistically, we discovered that the epithelial-specific splicing regulators (ESRP1 and ESRP2), flank many AR-regulated alternatively spliced exons. And, using 2D-invasion assays, we show that the inhibition of ESRPs can suppress AR-antagonist driven tumor invasion. In conclusion, until now, AR signaling has been primarily thought to modulate transcriptome of prostate epithelial cells by inducing or suppressing gene expression. Our work provides evidence for a new mechanism by which AR alters the transcriptome of prostate cancer cells by modulating alternative splicing. As such, our work has important implications for CRPC progression and development of resistance to treatment with bicalutamide and enzalutamide.

cancer biology↗