Biology subjects

Ravichandran, K.

Publications and source records attributed to Ravichandran, K..

8 recordsLinked to original sources

bioRxiv2025

Long non-coding RNA Cerox1 targets components of the mitochondrial electron transport chain to regulate the memory impairment caused by sleep deprivation

AbstractSleep deprivation (SD) impairs long-term memory, but the molecular mechanisms underlying the impact of sleep loss on memory are poorly understood. Molecular changes driven by SD have thus far focused on transcription and translation. Long non-coding RNAs (lncRNAs), a class of regulatory RNAs, have recently been recognized as an important player in memory research. However, it remains unclear how sleep deprivation modulates the expression of lncRNAs or their targets to lead to memory impairment. In this study, we explored the role of lncRNAs in the disruption of spatial memory caused by SD. We examined a set of synapse-associated lncRNAs that were identified through a transcriptome analysis after SD. Among them, we discovered that the lncRNA Cerox1 is downregulated in dorsal hippocampus following SD, and its levels recover after 2.5 hours of rebound sleep. Sleep is critical for the regulation of metabolism and sleep loss impairs mitochondrial function. Both sleep deprivation and Cerox1 knockdown were found to reduce complex I activity of the mitochondrial electron transport chain. This reduction of complex I activity is linked to the decrease in expression of a subset of complex I subunits including Ndufs1, Ndufs3, Ndufa3 and Ndufs6. Overexpression of Cerox1 has the opposite effect, leading to increased complex I activity. Sleep deprivation reduced ATP levels in the dorsal hippocampus, while Cerox1 overexpression restored it. SD disrupted memory consolidation, and this impairment was rescued when Cerox1 was overexpressed. Cerox1 transcript contains multiple miRNA binding sites that regulate the activity of the lncRNA. Notably, overexpression of Cerox1 transcript lacking miRNA binding sites did not rescue the memory deficit caused by SD. Our findings demonstrated that the impairment of memory consolidation after SD is linked to lncRNA-mediated control of mitochondrial electron transport chain activity essential for sustaining energy requirements. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=141 SRC="FIGDIR/small/676326v1_ufig1.gif" ALT="Figure 1000"> View larger version (48K): org.highwire.dtl.DTLVardef@6d72d5org.highwire.dtl.DTLVardef@18e3590org.highwire.dtl.DTLVardef@15e83c9org.highwire.dtl.DTLVardef@1e1f92_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2025

Innate detection of Salmonella replication triggers caspase-8-dependent apoptosis via TLR-driven TNF signaling and NLRC4-mediated sensing of the SPI-2 Type III secretion system

Salmonella enterica comprises over 2500 serovars that are responsible for over 90 million annual infections and 100,000 deaths worldwide. Despite this diversity, our understanding of innate immune responses to Salmonella is based on extensive study of a few serovars, primarily Typhimurium, including strains that cannot replicate within primary murine macrophages. Non-replicating Salmonella trigger caspase-1 and -11-dependent pyroptosis. Whether the innate immune system distinguishes between replicating and non-replicating intracellular Salmonella is poorly defined. Here we demonstrate that replicating Salmonella enterica induce a distinct pathway of TNF- and caspase-8-driven apoptosis via host TLR4 and Salmonella Pathogenicity Island-2 activity. This pathway is independent of gasdermin D and involves the apoptotic pore protein Pannexin-1. Combined loss of Pannexin-1 and gasdermin D resulted in defective control of systemic Salmonella, indicating that these pathways function together to promote anti-Salmonella host defense. Altogether, our findings uncover a previously unappreciated pathway by which macrophages sense intracellular replicating bacteria.

microbiology↗

bioRxiv2024

Effects of intergenerational transmission of small intestinal bacteria cultured from stunted Bangladeshi children with enteropathy

Environmental enteric dysfunction (EED), a small intestinal disorder found at a high prevalence in stunted children, is associated with gut mucosal barrier disruption and decreased absorptive capacity1-4. To test the hypothesis that intergenerational transmission of a perturbed small intestinal microbiota contributes to undernutrition by inducing EED5, we characterized two consortia of bacterial strains cultured from duodenal aspirates from stunted Bangladeshi children with EED - one of which induced local and systemic inflammation in female gnotobiotic mice. Offspring of dams colonized with the inflammatory consortium exhibited impaired prenatal and postnatal growth, as well as immunologic changes phenocopying features of EED in children. Dam-to-pup transmission of the inflammatory consortium produced, in recently weaned offspring, alterations in inter-cellular signaling pathways related to intestinal epithelial cell renewal, barrier integrity and immune function. Cohousing of mice harboring the inflammatory or non-inflammatory consortia and subsequent screening of candidate disease-promoting bacterial isolates identified Campylobacter concisus, an organism typically found in the oral microbiota, as a contributor to enteropathy. The C. concisus strain induced, in a host nitric oxide synthase (NOS)-dependent manner, pro-inflammatory cytokine signaling. Moreover, host-derived nutrients generated by NOS augmented C. concisus growth. This preclinical model should facilitate identification of small intestinal microbiota-targeted therapeutics for (intergenerational) undernutrition.

systems biology↗

bioRxiv2023

G1 length dictates heterochromatin landscape

Stem cells have lower facultative heterochromatin as defined by trimethylation of histone H3 lysine 27 (H3K27me3) compared to differentiated cells. However, the mechanisms underlying these differential H3K27me3 levels remain elusive. Because H3K27me3 levels are diluted two-fold in every round of replication and then restored through the rest of the cell cycle, we reasoned that the cell cycle length could be a key regulator of total H3K27me3 levels. Here, we propose that a fast cell cycle restricts H3K27me3 levels in stem cells. To test this model, we determined changes to H3K27me3 levels in mESCs globally and at specific loci upon G1 phase lengthening - accomplished by thymidine block or growth in the absence of serum (with the "2i medium"). H3K27me3 levels in mESC increase with G1 arrest when grown in serum and in 2i medium. Additionally, we observed via CUT&RUN and ChIP-seq that regions that gain H3K27me3 in G1 arrest and 2i media overlap, supporting our model of cell cycle length as a critical regulator of the stem cell epigenome and cellular identity. Furthermore, we demonstrate the inverse effect - that G1 shortening in differentiated cells results in a loss of H3K27me3 levels. Finally, in tumor cells with extreme H3K27me3 loss, lengthening of the G1 phase leads to H3K27me3 recovery despite the presence of the dominant negative, sub-stoichiometric H3.1K27M mutation. Our results indicate that G1 length is an essential determinant of H3K27me3 landscapes across diverse cell types.

molecular biology↗

bioRxiv2023

GDF15 mediates inflammation-associated bone loss through a brain-bone axis

Metabolic mediators play an important role in regulating chronic inflammation in the body. Here we report an unexpected role for GDF15 (Growth Differentiation Factor 15), a central mediator of food intake, in inflammation-associated bone loss. GDF15 serum levels were found to be elevated in arthritis patients and inversely correlated with bone density. Despite being associated with inflammation, we found that GDF15 itself does not cause, nor contribute to, clinical or histopathological arthritis. Rather, under inflammatory conditions, GDF15 mediates trabecular bone loss through its receptor GFRAL, which is exclusively expressed in the hindbrain. GDF15-GFRAL binding results in {beta}-adrenergic activation of MALPs (Marrow Adipocytic Lineage Precursors) in the bone marrow, which stimulate osteoclasts and trigger bone loss. These data suggest a metabolic mediator-controlled brain-bone axis in inflammation, through which bone loss is induced in a contextual rather than general manner. These findings may lead to more specific therapeutic interventions to protect bone.

immunology↗

bioRxiv2023

CRISPR based programmable RNA editing in primary neurons

Investigating the RNA regulation landscape primarily relies on our understanding of how RNA-protein interactions are governed in various cell types, including neurons. Analysis of RNA-protein interactions in physiological environments warrants the development of new tools that rely on RNA manipulation. Recently, A CRISPR-based RNA-editing tool (dCas13b-ADAR2DD) was developed to mitigate disease associated point mutations in cell lines. Here, we have explored the targeted sequence editing potential of the tool (dCas13b-ADAR2DD system) by adapting it to manipulate RNA function with an aim to visualize RNA editing in primary hippocampal neurons. This is a two-component system that includes a programmable guide RNA (gRNA) complementary to the target RNA, and a catalytically dead version of the Cas13b enzyme fused to ADAR. The RNA editing protocol outlined in this manuscript relies on using the gRNA-dependent targeting of dCas13b-ADAR fusion protein to the mutant form of mDendra transcript. We first abrogated the fluorescence of Dendra2 by introducing a nonsense mutation that precludes the formation of the functional protein. To visualize the efficacy of the RNA editing in neurons, we used the dCas13b-ADAR2DD system to edit specific nucleotides within the Dendra2 mRNA to restore the amino acid codes critical for Dendra2 fluorescence. This method therefore lays the foundation to future studies on the dynamicity of activity-induced RNA-protein interactions in neurons and can be extended to manipulate the endogenous RNome in diverse neuronal subtypes. Furthermore, this methodology will enable investigators to visualize the spatial and temporal resolution of RNA-protein interactions without altering the genomes via conventional methods. HighlightsO_LICRISPR-Cas13 based application that enables site specific A-to-I in target RNAs directed by gRNA; optimized in neurons. C_LIO_LIEnables the temporal mapping of developmentally relevant RNAs and their cis-interacting RNA binding proteins. C_LI

neuroscience↗

bioRxiv2022

Amount of Pannexin 1 in smooth muscle cells regulates sympathetic nerve induced vasoconstriction

Pannexin 1 (Panx1) forms high conductance channels that secrete ATP upon stimulation. The role of Panx1 in mediating constriction in response to direct sympathetic nerve stimulation is not known. Additionally, it is unknown how the expression level of Panx1 in SMCs influences a-adrenergic responses. We hypothesized that the amount of Panx1 in SMCs dictates the levels of sympathetic constriction and blood pressure. To test this hypothesis, we used genetically modified mouse models enabling expression of Panx1 in vascular cells to be varied. Genetic deletion of SMC Panx1 prevented constriction by electric field stimulation of sympathetic nerves. Conversely, over-expression of Panx1 in SMCs using a ROSA26 transgenic model increased sympathetic nerve-mediated constriction. Cx43 hemichannel inhibitors did not alter constriction. Next, we evaluated the effects of altered SMC Panx1 expression on blood pressure. To do this, we created mice combining a global Panx1 deletion, with ROSA26-Panx1 under the control of an inducible SMC specific Cre (Myh11). This resulted in mice that could express only human Panx1, only in SMCs. After tamoxifen, these mice had increased blood pressure that was acutely decreased by the Panx1 inhibitor spironolactone. Control mice genetically devoid of Panx1 did not respond to spironolactone. These data suggest Panx1 in SMCs could regulate the extent of sympathetic nerve constriction and blood pressure. The results also show the feasibility humanized Panx1-mouse models to test pharmacological candidates.

physiology↗

bioRxiv2020

Ribonucleotide reductase, a novel target for gonorrhea

Antibiotic resistant Neisseria gonorrhoeae (Ng) are an emerging public health threat due to increasing numbers of multidrug resistant (MDR) organisms. We identified two novel orally active inhibitors, PTC-847 and PTC-672, that exhibit a narrow spectrum of activity against Ng including MDR isolates. By selecting organisms resistant to the novel inhibitors and sequencing their genomes, we identified a new therapeutic target, the class Ia ribonucleotide reductase (RNR). Activity studies and negative stain electron microscopy of the Ng Ia RNR suggest that these inhibitors potentiate conversion of its active 2{beta}2 state to an inactive 4{beta}4 similar to states first identified with the Escherichia coli (Ec) Ia RNR. Resistance mutations in Ng map to the N-terminal, ATP cone domain of its subunit and disrupt the interaction with the {beta} subunit required to form the specific quaternary inhibited state. Oral administration of PTC-672 reduces Ng infection in a mouse model and may have therapeutic potential for treatment of Ng that is resistant to current drugs.

microbiology↗