Biology subjects

Manjegowda, M. C.

Publications and source records attributed to Manjegowda, M. C..

4 recordsLinked to original sources

bioRxiv2023

Novel TRPM7 inhibitors with potent anti-inflammatory effects in vivo

TRPM7, a TRP channel with ion conductance and kinase activities, has emerged as an attractive drug target for immunomodulation. Reverse genetics and cell biological studies have already established a key role for TRPM7 in the inflammatory activation of macrophages. Advancing TRPM7 as a viable molecular target for immunomodulation requires selective TRPM7 inhibitors with in vivo tolerability and efficacy. Such inhibitors have the potential to interdict inflammatory cascades mediated by systemic and tissue-specialized macrophages. FTY720, an FDA-approved drug for multiple sclerosis inhibits TRPM7. However, FTY720 is a prodrug and its metabolite, FTY720-phosphate, is a potent agonist of sphingosine 1-phosphate (S1P) receptors. In this study, we tested non-phosphorylatable FTY720 analogs, which are inert against S1PRs and well tolerated in vivo, for activity against TRPM7 and tissue bioavailability. Using patch clamp electrophysiology, we show that VPC01091.4 and AAL-149 block TRPM7 current at low micromolar concentrations. In culture, they act directly on macrophages to blunt LPS-induced inflammatory cytokine expression, an effect that is predominantly but not solely mediated by TRPM7. We found that VPC01091.4 has significant and rapid accumulation in the brain and lungs, along with direct anti-inflammatory action on alveolar macrophages and microglia. Finally, using a mouse model of endotoxemia, we show VPC01091.4 to be an efficacious anti-inflammatory agent that arrests systemic inflammation in vivo. Together, these findings identify novel small molecule inhibitors that allow TRPM7 channel inhibition independent of S1P receptor targeting. These inhibitors exhibit potent anti-inflammatory properties that are mediated by TRPM7 and likely other molecular targets that remain to be identified.

immunology↗

bioRxiv2022

The G-protein-coupled estrogen receptor, a gene co-expressed with ERα in breast tumors, is regulated by estrogen-ERα signalling in ERα positive breast cancer cells.

PurposeThe purpose of this study was to assess the relationship between GPER, a seven transmembrane G-protein coupled estrogen receptor, and ER in breast tumors, and to make inroads into the mechanistic basis and clinical significance. MethodsTCGA-BRCA data was mined to examine the relationship between GPER and ER expression. GPER mRNA, and protein expression were analyzed in ER-positive or -negative breast tumors from two cohorts using immunohistochemistry, western blotting, or RT-qPCR. The Kaplan-Meier Plotter was employed for survival analysis. The influence of estrogen in vivo was studied by examining GPER expression levels in estrus or diestrus mouse mammary tissues, and the impact of 17{beta}-estradiol (E2) administration in juvenile or adult mice. The effect of E2, or propylpyrazoletriol (PPT, an ER agonist) stimulation on GPER expression was studied in MCF-7 and T47D cells, with or without tamoxifen or ER knockdown. ER-binding to the GPER locus was explored by analysing ChIP-seq data (ERP000380), in silico prediction of estrogen response elements, and chromatin immunoprecipitation assay. ResultsClinical data revealed significant positive association between GPER and ER expression in breast tumors. The median GPER expression in ER-positive tumors was significantly higher than ER-negative tumors. High GPER expression was significantly associated with longer overall survival of patients with ER-positive tumors. In vivo experiments showed a positive effect of E2 on GPER expression. E2 induced GPER expression in MCF-7 and T47D cells; an effect mimicked by PPT. Tamoxifen or ER-knockdown blocked the induction of GPER. Estrogen-mediated induction was associated with increased ER occupancy in the upstream region of GPER. ConclusionGPER expression is positively associated with ER in breast tumors, and a transcriptional target of the estrogen-ER signalling axis. More in-depth studies are required to establish the significance of GPER-ER co-expression, and their interplay in breast tumor development, progression, and treatment.

cancer biology↗

bioRxiv2021

DNA Methyltransferase 1 and 3a Expression in the Frontal Cortex Regulates Palatable Food Consumption

DNA methylation is an important regulatory mechanism in the control of neuronal function. Both during development and following exposure to salient stimuli, plasticity in the methylation of cytosine residues leads to a change in neuron excitability that subsequently sculpts animal behavior. However, although the response of DNA methyltransferase enzymes in adult neurons to stimuli such as drugs of abuse have been described, less is known about how these enzymes regulate methylation at specific loci to change the drive to ingest natural rewards. Specifically, we do not understand how changes in methylation within important brain areas known to regulate palatable food intake can affect ingestion, while a detailed investigation of the neurophysiological and genomic effects of perturbing methyltransferase function has not been pursued. By deleting DNA methyltransferase 1 and 3a in the mouse prefrontal cortex, we observed the requirement for these enzymes in the regulation of nutrient rich food consumption in the absence of any effect on the intake of low fat and low sugar chow. We also determined that the deletion profoundly affected neuron excitability within pyramidal cells resident in superficial layers II/III of the cortex but had little effect in deep layer V neurons. Finally, reduced representation bisulfite sequencing revealed both hypo and hypermethylation in response to methyltransferase deletion, an effect that was observed in binding sites for retinoic acid receptor beta (RAR{beta}) located within regulatory regions of genes known to affect neuronal function. Together, our data suggest that alterations in the actions of RAR{beta} could shift neuronal activity to reduce palatable food intake.

neuroscience↗

bioRxiv2019

Hdac3, Setdb1, and Kap1 mark H3K9me3/H3K14ac bivalent regions in young and aged liver.

Post-translational modifications of histone tails play a crucial role in gene regulation. Here, we performed chromatin profiling by quantitative targeted mass spectrometry to assess all possible modifications of the core histones. We discovered a novel bivalent combination, a dually-marked H3K9me3/H3K14ac modification in the liver, that is significantly decreased in old hepatocytes. Subsequent genome-wide location analysis (ChIP-Seq) identified 1032 and 668 bivalent regions in young and old livers, respectively, with 280 in common. Histone H3K9 deacetylase Hdac3, as well as H3K9 methyltransferase Setdb1, found in complex Kap1, occupied bivalent regions in both young and old livers, correlating to presence of H3K9me3. Expression of genes associated with bivalent regions in young liver, including those regulating cholesterol secretion and triglyceride synthesis, is upregulated in old liver once the bivalency is lost. Hence, H3K9me3/H3K14ac dually-marked regions define a poised inactive state that is resolved with loss of one or both of the chromatin marks, which subsequently leads to change in gene expression.

genomics↗