Biology subjects

Sharma, B.

Publications and source records attributed to Sharma, B..

42 records · Page 3Linked to original sources

bioRxiv2022

Acute sleep deprivation induces synaptic remodeling at the soleus muscle neuromuscular junction in rats

Sleep is important for cognitive and physical performance. Sleep deprivation not only affects neural functions but also results in muscular fatigue. A good nights sleep reverses these functional derangements caused by sleep deprivation. The role of sleep in brain function has been extensively studied. However, its role in neuromuscular junction or skeletal muscle morphology is sparsely addressed although skeletal muscle atonia and suspended thermoregulation during rapid eye movement sleep possibly provides a conducive environment for the muscle to rest and repair; somewhat similar to slow-wave sleep for synaptic downscaling. In the present study, we have investigated the effect of 24 h sleep deprivation on the neuromuscular junction morphology and neurochemistry using electron microscopy and immunohistochemistry in the rat soleus muscle. Acute sleep deprivation altered synaptic ultra-structure viz. mitochondria, synaptic vesicle, synaptic proteins, basal lamina, and junctional folds needed for neuromuscular transmission. Further acute sleep deprivation showed the depletion of the neurotransmitter acetylcholine and the overactivity of its degrading enzyme acetylcholine esterase at the neuromuscular junction. The impact of sleep deprivation on synaptic homeostasis in the brain has been extensively reported recently. The present evidence from our studies shows new information on the role of sleep on neuromuscular junction homeostasis and its functioning. Statement of significanceSleep causes synaptic downscaling in the brain, and allows the brain to carry out various housekeeping functions. Here we have reported that the function of the sleep-wake cycle on the synaptic homeostasis extends beyond the brain. Acute sleep deprivation caused significant alteration at ultra and macrostructure of antigravity muscle and the neuromuscular junction along with adaptation to new fiber type in rats. These morpho-functional changes were well correlated with the biochemical assessment of the acetylcholine at the neuromuscular junction. These changes were partially recovered when the rats were allowed to recover from sleep deprivation. The findings suggest a new avenue for a sleep study; employing the neuromuscular junction for exploring the effect of sleep at energy and synaptic homeostasis levels.

neuroscience↗

bioRxiv2022

Complete sequences of the velvet worm slime proteins reveal that slime formation is enabled by disulfide bonds and intrinsically disordered regions

The slime of velvet worms (Onychophora) is a strong and fully biodegradable protein material, which upon ejection undergoes a fast liquid-to-solid transition to ensnare prey. However, the molecular mechanisms of slime self-assembly are still not well understood, notably because the primary structures of slime proteins are yet unknown. Combining transcriptomic and proteomic studies, we have obtained the complete primary sequences of slime proteins and identified key features for slime self-assembly. The high molecular weight slime proteins contain Cys residues at the N- and C-termini that mediate the formation of multi-protein complexes via disulfide bonding. Low complexity domains in the N-termini were also identified and their propensity for liquid-liquid phase separation established, which may play a central role for slime biofabrication. Using solid-state nuclear magnetic resonance, rigid and flexible domains of the slime proteins were mapped to specific peptide domains. The complete sequencing of major slime proteins is an important step towards sustainable fabrication of polymers inspired by the velvet worm slime.

bioengineering↗

bioRxiv2021

Endocardium-to-coronary artery differentiation during heart development and regeneration involves sequential roles of Bmp2 and Cxcl12/Cxcr4.

Regenerating coronary blood vessels has the potential to ameliorate ischemic heart disease, yet there is currently no method of stimulating clinically effective cardiac angiogenesisis. Endocardial cells-- a particularly plastic cell type during development--line the heart lumen and are natural coronary vessel progenitors. Their intrinsic angiogenic potential is lost in adults, but studying the endocardial- to-coronary developmental pathway could identify methods of re-instating this process in diseased hearts. Here, we use a combination of mouse genetics and scRNAseq of lineage-traced endothelial cells to identify novel regulators of endocardial angiogenesis and precisely assess the role of Cxcl12/Cxcr4 signaling. Time-specific lineage tracing demonstrated that endocardial cells differentiated earlier than previously thought, largely at mid-gestation. A new mouse line reporting the activity of Cxcr4 revealed that, despite widespread Cxcl12 and Cxcr4 expression, only a small subset of these coronary endothelial cells activated the receptor, which were mostly in arteries. In accordance with these two findings, Cxcr4 deletion in the endocardial lineage only affected artery formation and only when deleted before mid-gestation. Integrating scRNAseq data of coronary endothelial cells from the endocardial lineage at both mid- and late-gestation identified a transitioning population that was specific to the earlier timepoint that specifically expressed Bmp2. Recombinant Bmp2 stimulated endocardial angiogenesis in an in vitro explant assay and in neonatal mouse hearts upon myocardial infarction. Our data shed light on how understanding the molecular mechanisms underlying endocardial-to-coronary transitions can identify new potential therapeutic targets that could promote revascularization of the injured heart.

developmental biology↗

bioRxiv2021

Cell lineage as a predictor of immune response in neuroblastoma

Immunotherapy for patients with neuroblastoma has met with limited success, partly due to an incomplete understanding of the mechanisms underlying immune responsiveness in this clinically and genetically heterogenic tumor. Here, we undertook an unbiased analysis using dimension reduction and UMAP visualization of transcriptional signatures derived from 498 primary neuroblastoma tumors. Four distinct clusters based on differentially expressed genes emerged, of which one, representing about 30% and comprising mainly of MYCN-nonamplified tumors, was notable for the high expression of genes associated with both immune response activation and suppression. This capacity to elicit a productive immune response resided exclusively in tumors with dominant populations of undifferentiated, neural crest-like or mesenchymal cells; by contrast, tumors comprising primarily of committed, adrenergic neuron-like cells were less immunogenic. Mesenchymal neuroblastoma cells were enriched for innate and adaptive immune gene signatures, demonstrated engagement with cytotoxic T and natural killer cells, and induced immune cell infiltration in an immunocompetent mouse model. Transcriptional or targeted therapy-induced reprogramming of adrenergic cells to the mesenchymal state led to reactivation of tumor cell-intrinsic immune genes. Key immune response genes in adrenergic tumor cells were found to be epigenetically silenced by the PRC2 complex, and such repression could be relieved by either mesenchymal cell state reprogramming or EZH2 inhibition, leading to increased activation of natural killer cells by the tumor cells. These data identify cell lineage as a major determinant of the immunogenic potential in neuroblastoma that could be used to stratify patients who are most likely to benefit from immunotherapy.

cancer biology↗

bioRxiv2020

Conjugates of neuroprotective chaperone L-PGDS provide MRI contrast for detection of amyloid β-rich regions in live Alzheimer's Disease mouse model brain

Endogenous brain proteins can recognize the toxic oligomers of amyloid-{beta} (A{beta}) peptides implicated in Alzheimers disease (AD) and interact with them to prevent their aggregation. Lipocalin-type Prostaglandin D Synthase (L-PGDS) is a major A{beta}-chaperone protein in the human cerebrospinal fluid. Here we demonstrate that L-PGDS detects amyloids in diseased mouse brain. Conjugation of L-PGDS with magnetic nanoparticles enhanced the contrast for magnetic resonance imaging. We conjugated the L-PGDS protein with ferritin nanocages to detect amyloids in the AD mouse model brain. We show here that the conjugates administered through intraventricular injections co-localize with amyloids in the mouse brain. These conjugates can target the brain regions through non-invasive intranasal administration, as shown in healthy mice. These conjugates can inhibit the aggregation of amyloids in vitro and show potential neuroprotective function by breaking down the mature amyloid fibrils.

biochemistry↗

bioRxiv2018

CDK12 loss in cancer cells affects DNA damage response genes through premature cleavage and polyadenylation

The cyclin-dependent kinase 12 (CDK12) modulates transcription elongation by phosphorylating the carboxy-terminal domain of RNA polymerase II and appears to selectively affect the expression of DNA damage response (DDR) and mRNA processing genes. Yet, the mechanism(s) by which it achieves this selectivity remains unclear. Using a highly selective CDK12/13 inhibitor, THZ531, and nascent RNA sequencing, we show that CDK12 inhibition results in gene length-dependent elongation defects, leading to premature cleavage and polyadenylation (PCPA) as well as loss of expression of long (>45 kb) genes, a substantial proportion of which participate in the DDR. This early termination phenotype correlated with an increased proportion of intronic polyadenylation sites, a feature that was especially prominent among DDR genes. Finally, phosphoproteomic analysis indicated that pre-mRNA processing factors, including those involved in PCPA, are direct phosphotargets of CDKs 12 and 13. These results support a model in which DDR genes are uniquely susceptible to CDK12 inhibition due primarily to their relatively longer lengths and lower ratios of U1 snRNP binding to intronic polyadenylation sites.

cancer biology↗