Biology subjects

Jacob, R. S.

Publications and source records attributed to Jacob, R. S..

5 recordsLinked to original sources

bioRxiv2024

α-Synuclein acts as a cholesteryl-ester sensor on lipid droplets regulating organelle size and abundance.

While aggregated alpha-Synuclein (Syn) is commonly associated with Parkinsons disease, its physiological function as a membrane-binding protein is poorly understood. Here, we show that endogenous Syn binds lipid droplets (LDs) in multiple human cell lines and in stem cell-derived dopaminergic neurons. LD-binding encompasses Syn residues 1-100, which masks their detection by immunofluorescence microscopy, probably explaining the scarcity of similar observations in earlier studies. Syn-LD interactions are highly temperature-sensitive and selective for cholesteryl-ester-rich LDs. They promote the formation of Syn multimers that dissociate from LDs at non-permissive temperatures. Syn remains LD-bound throughout starvation-induced lipolysis, whereas siRNA-knockdown diminishes LD abundance and compromises cell viability upon nutrient depletion, without affecting LD biosynthesis. Reciprocally, excess Syn stimulates LD accumulation in dependence of lipid availability, restricts organelle size and ensures intracellular LD organization, which strictly depends on functional membrane-binding. Supporting a general role of Syn in cellular lipid and cholesterol metabolism, our results point to additional loss-of-function similarities between Parkinsons, Alzheimers and Gauchers disease.

cell biology↗

bioRxiv2021

Novel microRNAs targeting NMDA receptor subunits in animal models of schizophrenia

N-methyl-D-aspartate receptors (NMDAR) are downregulated in schizophrenia possibly through microRNAs (miRNAs) that are differentially expressed in this condition. We screened the miRNAs that are altered in schizophrenia against the targets, Grin2A and Grin2B subunits of NMDAR using bioinformatic tools. Among the predicted miRNAs some interacted with the 3-UTR sequences of Grin2A (miR-296, miR-148b, miR-129-2, miR-137) and Grin2B (miR-296, miR-148b, miR-129-2, miR-223) in dual luciferase assays. This was supported by downregulation of the GluN2B protein in primary hippocampal neurons upon overexpressing Grin2B targeting miRNAs. In two models of schizophrenia-pharmacological MK-801 model and neurodevelopmental methylazoxymethanol acetate (MAM) model which showed cognitive deficits - protein levels of GluN2A and GluN2B were downregulated but their transcript levels were upregulated. MiR-296-3p, miR-148b-5p and miR-137 levels showed upregulation in both models which could have interacted with Grin2A/Grin2B transcripts resulting in translational arrest. In MAM model, reciprocal changes in the expression of the 3p and 5p forms of miR-148b and miR-137 were observed. Expression of neuregulin 1 (NRG1), BDNF and CaMKII, genes implicated in schizophrenia, were also altered in these models. This is the first report of downregulation of GluN2A and GluN2B by miR-296, miR-148b and miR-129-2. Mining miRNAs regulating NMDA receptors might give insights into the pathophysiology of this disorder, providing avenues in therapeutics.

neuroscience↗

bioRxiv2021

Co-aggregation and secondary nucleation in the life cycle of human prolactin/galanin functional amyloids

Synergistic-aggregation and cross-seeding by two different amyloid proteins/peptides are well evident in various neurological disorders. However, this phenomenon is not well studied in functional amyloid aggregation. Here, we show Prolactin (PRL) is associated with lactation in mammals and neuropeptide galanin (GAL), which are co-stored in the lactotrophs facilitates the synergic aggregation in the absence of secretory granules helper molecules glycosaminoglycans (GAGS). Interestingly, although each partner possesses homotypic seeding ability, a unidirectional cross-seeding of GAL aggregation can be mediated by PRL seeds. The specificity of co-aggregation by PRL and GAL along with unidirectional cross-seeding suggests tight regulation of functional amyloid formation during co-storage of these hormones in secretory granule biogenesis of female rat lactotrophs. Further mixed fibrils release the constituent functional hormone much faster than the corresponding individual amyloid formed in presence of GAGs, suggesting that co-aggregation of functionally distant hormones might have evolved for efficient storage, synergistic and rapid release of both hormones upon stimulation. The co-aggregation and cross seeding by two different hormones of completely different structures and sequences (PRL and GAL) suggest a novel mechanism of heterologous amyloid formation both in disease and functional amyloids.

biophysics↗

bioRxiv2020

α-Synuclein plasma membrane localization correlates with cellular phosphatidylinositol polyphosphate levels.

The Parkinsons disease protein -synuclein (Syn) promotes membrane fusion and fission by interacting with various negatively charged phospholipids. Despite postulated roles in endocytosis and exocytosis, plasma membrane (PM) interactions of Syn are poorly understood. Here, we show that phosphatidylinositol 4,5-bisphosphate (PIP2) and phosphatidylinositol 3,4,5-trisphosphate (PIP3), two highly acidic components of inner PM leaflets, mediate plasma membrane localization of endogenous pools of Syn in A2780, HeLa, SH-SY5Y and SK-MEL-2 cells. We demonstrate that Syn binds reconstituted PIP2-membranes in a helical conformation in vitro and that PIP2 synthesizing kinases and hydrolyzing phosphatases reversibly redistribute Syn in cells. We further delineate that Syn-PM targeting follows phosphoinositide-3 kinase (PI3K)-dependent changes of cellular PIP2 and PIP3 levels, which collectively suggests that phosphatidylinositol polyphosphates contribute to Syns cellular function(s) at the plasma membrane.

cell biology↗

bioRxiv2020

An NMR-based biosensor to measure stereo-specific methionine sulfoxide reductase (MSR) activities in vitro and in vivo.

Oxidation of protein methionines to methionine-sulfoxides (MetOx) is associated with several age-related diseases. In healthy cells, MetOx is reduced to methionine by two families of conserved methionine sulfoxide reductase enzymes, MSRA and MSRB that specifically target the S- or R-diastereoisomers of methionine-sulfoxides, respectively. To directly interrogate MSRA and MSRB functions in cellular settings, we developed an NMR-based biosensor that we call CarMetOx to simultaneously measure both enzyme activities in single reaction setups. We demonstrate the suitability of our strategy to delineate MSR functions in complex biological environments that range from native cell lysates to zebrafish embryos. Thereby, we establish differences in substrate specificities between prokaryotic and eukaryotic MSRs and introduce CarMetOx as a highly sensitive tool for studying therapeutic targets of oxidative stress-related human diseases and redox regulated signaling pathways. Our approach further extends high-resolution in-cell NMR measurements of exogenously delivered biomolecules to an entire multicellular organism.

biophysics↗