Biology subjects

Venkatesan, M.

Publications and source records attributed to Venkatesan, M..

8 recordsLinked to original sources

bioRxiv2024

Neurofilament accumulation disrupts autophagy in giant axonal neuropathy

Neurofilament accumulation is a marker of several neurodegenerative diseases, but it is the primary pathology in Giant Axonal Neuropathy (GAN). This childhood onset autosomal recessive disease is caused by loss-of-function mutations in gigaxonin, the E3 adaptor protein that is essential for neurofilament degradation. Using a combination of genetic and RNA interference (RNAi) approaches, we found that dorsal root ganglia from mice lacking gigaxonin have impaired autophagy and lysosomal degradation through two mechanisms. First, neurofilament accumulations interfere with the distribution of autophagic organelles, impairing their maturation and fusion with lysosomes. Second, the accumulations sequester the chaperone 14-3-3, a protein responsible for the localization of the transcription factor EB (TFEB), a key regulator of autophagy. This dual disruption of autophagy likely contributes to the pathogenesis of other neurodegenerative diseases with neurofilament accumulations.

neuroscience↗

bioRxiv2024

Trends of Plasmodium falciparum molecular markers associated with resistance to artemisinins and reduced susceptibility to lumefantrine in Mainland Tanzania from 2016 to 2021

BackgroundTherapeutic efficacy studies (TESs) and detection of molecular markers of drug resistance are recommended by the World Health Organization (WHO) to monitor the efficacy of artemis inin combination therapy (ACT). This study assessed the trends of molecular markers of artemis inin resistance and/or reduced susceptibility to lumefantrine using samples collected in TES conducted in Mainland Tanzania from 2016 to 2021. MethodsA total of 2,015 samples were collected during TES of artemether-lumefantrine at eight sentinel sites (in Kigoma, Mbeya, Morogoro, Mtwara, Mwanza, Pwani, Tabora, and Tanga regions) between 2016 and 2021. Photo-induced electron transfer polymerase chain reaction (PET-PCR) was used to confirm presence of malaria parasites before capillary sequencing, which targeted two genes: Plasmodium falciparum kelch 13 propeller domain (k13) and P. falciparum multidrug resistance 1 (pfmdr1). ResultsSequencing success was [≥]87.8%, and 1,724/1,769 (97.5%) k13 wild-type samples were detected. Thirty-seven (2.1%) samples had synonymous mutations and only eight (0.4%) had non-synonymous mutations in the k13 gene; seven of these were not validated by WHO as molecular markers of resistance (I416V, E433D, R471S, P475S, A578S, and Q613E). One sample from Morogoro in 2020 had a k13 R622I mutation, which is a validated marker of artemisinin partial resistance. For pfmdr1, all except two samples carried N86 (wild-type), while mutations at Y184F increased from 33.9% in 2016 to about 60.5% in 2021, and only four samples (0.2%) had D1246Y mutations. pfmdr1 haplotypes were reported in 1,711 samples, with 985 (57.6%) NYD, 720 (42.1%) NFD, and six (0.4%) carrying minor haplotypes (three with NYY, 0.2%; YFD in two, 0.1%; and NFY in one sample, 0.1%). Between 2016 and 2021, NYD decreased from 66.1% to 45.2%, while NFD increased from 38.5% to 54.7%. ConclusionThis is the first report of the R622I (k13 validated mutation) in Tanzania. N86 and D1246 were nearly fixed, while increases in Y184F mutations and NFD haplotype were observed between 2016 and 2021. Despite the reports of ART-R in Rwanda and Uganda, this study did not report any other validated mutations in these study sites in Tanzania apart from R622I suggesting that intensified surveillance is urgently needed to monitor trends of drug resistance markers and their impact on the performance of ACTs.

molecular biology↗

bioRxiv2023

Targeted lipid A modification of Shigella vaccine strains reduced endotoxicity without compromising immunogenicity or invasiveness

Shigella infection contributes significantly to the global disease burden, especially affecting young children in developing countries. Currently, a vaccine against Shigella is unavailable and the prevalence of antibiotic resistance amongst Shigella species is continually rising. Live-attenuated Shigella vaccine candidates developed at Walter Reed Army Institute of Research have shown remarkable immunogenicity but exhibit adverse reactogenicity, most likely due to the highly toxic lipid A moiety present on the bacterial membrane. Previous attempts at reducing the endotoxicity have focused on deletion of intrinsic lipid A biosynthesis enzymes. In this study, we instead introduce exogenous lipid A modifying enzymes, generating targeted modifications in the lipid A structure, leading to a dampened TLR4 response within the host. In doing so, we generated vaccine candidates with detoxified lipid A and unaltered O-antigen structure thereby preserving the serotype-specific immunity while reducing endotoxicity.

microbiology↗

bioRxiv2023

Mitochondrial calcium signaling mediated transcriptional regulation of keratin filaments is a critical determinant of melanogenesis

Mitochondria are versatile organelles that regulate several physiological functions. Many mitochondria-controlled processes are driven by mitochondrial Ca2+ signaling. However, role of mitochondrial Ca2+ signaling in melanosome biology remains unknown. Here, we show that pigmentation requires mitochondrial Ca2+ uptake. In vitro gain and loss of function studies demonstrated that Mitochondrial Ca2+ Uniporter (MCU) is crucial for melanogenesis while the MCU rheostats, MCUb and MICU1 negatively control melanogenesis. Zebrafish and mouse models showed that MCU plays a vital role in pigmentation in vivo. Mechanistically, MCU controls activation of transcription factor NFAT2 to induce expression of three keratins (keratin 5, 7 and 8), which we report as positive regulators of melanogenesis. Interestingly, keratin 5 in turn modulates mitochondrial Ca2+ uptake thereby this signaling module acts as a negative feedback loop that fine-tunes both mitochondrial Ca2+ signaling and melanogenesis. Mitoxantrone, an FDA approved drug that inhibits MCU, decreases physiological melanogenesis. Collectively, our data demonstrates a critical role for mitochondrial Ca2+ signaling in vertebrate pigmentation and reveal the therapeutic potential of targeting MCU for clinical management of pigmentary disorders. Given the centrality of mitochondrial Ca2+ signaling and keratin filaments in cellular physiology, this feedback loop may be functional in a variety of other pathophysiological conditions. HighlightsO_LIMCU complex mediated mitochondrial Ca2+ uptake is a novel regulator of vertebrate pigmentation C_LIO_LIKeratin filaments bridge mitochondrial Ca2+ signaling to melanosome biogenesis and maturation C_LIO_LITranscription factor NFAT2 connects mitochondrial Ca2+ dynamics to keratins expression C_LIO_LIMCU-NFAT2-Keratin 5 signaling module generates a negative feedback loop to maintain mitochondrial Ca2+ homeostasis and to ensure optimal melanogenesis C_LIO_LIInhibiting MCU with mitoxantrone, an FDA approved drug, leads to reduction in physiological pigmentation C_LI

cell biology↗

bioRxiv2022

Molecular mechanism of plasmid-borne resistance to sulfonamides

The sulfonamides (sulfas) are the oldest class of synthetic antibacterial that target the essential, conserved dihydropteroate synthase (DHPS) enzyme, encoded by folP, through chemical mimicry of its substrate p-aminobenzoic acid (pABA). Resistance has complicated their clinical utility and is widespread in pathogenic species. Resistance is mediated by acquisition of sul genes on mobile genetic elements, which code for the so-called Sul enzymes that are divergent DHPS enzymes with intrinsic sulfa-insensitivity. Even decades after the discovery of this resistance mechanism, its molecular details have not been understood. In this study, we elucidate the molecular basis for intrinsic resistance of Sul enzymes using x-ray crystallography, enzymology, mutagenesis, intrinsic tryptophan fluorescence, antibiotic susceptibility of a contemporary {Delta}folP strain, and adaptive laboratory evolution of folP. We show that the active sites of Sul enzymes possess a modified pABA-interaction region based on insertion of a Phe-Gly sequence. This insertion is necessary for discrimination between pABA and sulfonamides, more than 1000-fold loss in binding affinity of sulfas to Sul enzymes, and robust pan-sulfonamide resistance. We detect no fitness cost due to this active site modification, as it does not compromise the rate of dihydropteroate biosynthesis and complements the thymidine-auxotrophy of an E. coli folP deletion strain. Lab-evolved sulfa-resistance folP recapitulated this mechanism through the same active site insertion. Finally, we show that this insertion and a nearby loop confer increased active site flexibility of Sul enzymes relative to DHPS. These results provide a molecular foundation for revisiting DHPS-targeted antibacterials to evade resistance.

microbiology↗

bioRxiv2022

Recombinant production of growth factors for application in cell culture

Culturing eukaryotic cells has widespread applications in research and industry, including the emerging field of cell-cultured meat production colloquially referred to as "cellular agriculture". These applications are often restricted by the high cost of growth medium necessary for cell growth. Mitogenic protein growth factors (GFs) are essential components of growth medium and account for upwards of 90% of the total costs. Here, we present a set of expression constructs and a simplified protocol for recombinant production of functionally active GFs, including FGF-2, IGF-1, PDGF-BB and TGF-{beta}1 in Escherichia coli. Using this expression system, we produced soluble GFs from species including bovine, chicken, and fish. Bioactivity analysis revealed orthologs with improved performance compared to commercially available alternatives. We estimated that the production cost of GFs using our methodology will significantly reduce the cost of cell culture medium, facilitating low-cost protocols tailored for cultured meat production and tissue engineering.

cell biology↗

bioRxiv2021

Oxytocin promotes epicardial cell activation and heart regeneration after cardiac injury

Cardiovascular disease (CVD) is one of the leading causes of mortality worldwide, and frequently leads to massive heart injury and the loss of billions of cardiac muscle cells and associated vasculature. Critical work in the last two decades demonstrated that these lost cells can be partially regenerated by the epicardium, the outermost mesothelial layer of the heart, in a process that highly recapitulates its role in heart development. Upon cardiac injury, mature epicardial cells activate and undergo an epithelial-mesenchymal transition (EMT) to form epicardial-derived progenitor cells (EpiPCs), multipotent progenitors that can differentiate into several important cardiac lineages, including cardiomyocytes and vascular cells. In mammals, this process alone is insufficient for significant regeneration, but it might be possible to prime it by administering specific reprogramming factors, leading to enhanced EpiPC function. Here, we show that oxytocin (OXT), a hypothalamic neuroendocrine peptide, induces epicardial cell proliferation, EMT, and migration in a model of human induced pluripotent stem cell (hiPSC)-derived epicardial cells. In addition, we demonstrate that OXT is produced after cardiac cryoinjury in zebrafish, and that it elicits significant epicardial activation promoting heart regeneration. Oxytocin signaling is also critical for proper epicardium development in zebrafish embryos. The above processes are significantly impaired when OXT signaling is inhibited chemically or genetically through RNA interference. RNA sequencing data suggests that the transforming growth factor beta (TGF-{beta}) pathway is the primary mediator of OXT-induced epicardial activation. Our research reveals for the first time an evolutionary conserved brain-controlled mechanism inducing cellular reprogramming and regeneration of the injured mammalian and zebrafish heart, a finding that could contribute to translational advances for the treatment of cardiac injuries.

developmental biology↗

bioRxiv2019

Zoonotic tuberculosis in India: looking beyond Mycobacterium bovis

BackgroundZoonotic tuberculosis (zTB) is the transmission of Mycobacterium tuberculosis complex (MTBC) subspecies from animals to humans. zTB is generally quantified by determining the proportion of human isolates that are Mycobacterium bovis. Although India has the worlds largest number of human TB cases and the largest cattle population, where bovine TB is endemic, the burden of zTB is unknown. MethodsTo obtain estimates of zTB in India, a PCR-based approach was applied to sub-speciate positive MGIT(R) cultures from 940 patients (548 pulmonary, 392 extrapulmonary disease) at a large referral hospital in India. Twenty-five isolates of interest were subject to whole genome sequencing (WGS) and compared with 715 publicly available MTBC sequences from South Asia. FindingsA conclusive identification was obtained for 939 samples; wildtype M. bovis was not identified (95% CI: 0 - 0.4%). There were 912 M. tuberculosis sensu stricto (97.0%, 95% CI: 95.7 - 98.0), 7 M. orygis (95% CI: 0.3 - 1.5%); 5 M. bovis BCG, and 15 non-tuberculous mycobacteria. WGS analysis of 715 MTBC sequences again identified no M. bovis (95% CI: 0 - 0.4%). Human and cattle MTBC isolates were interspersed within the M. orgyis and M. tuberculosis sensu stricto lineages. InterpretationM. bovis prevalence in humans is an inadequate proxy of zTB in India. The recovery of M. orygis from humans, together with the finding of M. tuberculosis in cattle, underscores the need for One Health investigations to assess the burden of zTB in countries with endemic bovine TB. FundingBill & Melinda Gates Foundation, Canadian Institutes for Health Research

microbiology↗