Biology subjects

Mehta, N. R.

Publications and source records attributed to Mehta, N. R..

3 recordsLinked to original sources

bioRxiv2025

4-Pi Stimulated Raman Scattering for Label-free Super-resolution Chemical Imaging

AO_SCPLOWBSTRACTC_SCPLOWSuper-resolution fluorescence microscopy has transformed the study of biological structures and functions beyond the diffraction limit. Unlike fluorescence methods, label-free chemical imaging, mostly based on Raman and infrared spectroscopy, provides intrinsic molecular contrast, enabling the study of biomolecules, nanostructures, drug molecules, and metabolites that cannot be easily tagged. However, while a wide range of fluorescence-based super-resolution techniques are well-established, extending super-resolution to label-free chemical imaging has remained challenging due to low signal levels and limited resolution improvement. Super-resolution stimulated Raman scattering microscopy (SRS) is most promising due to its high sensitivity and imaging speed. Similar to fluorescence, existing SRS super-resolution approaches are mostly based on either photoswitching/saturation of molecular labels or sample expansion, which suffers from poor sensitivity due to limitations in labeling density or signal dilution, respectively. Moreover, axial resolution is typically much worse than lateral resolution, yet most super-resolution SRS techniques focused on improving lateral resolution. In this work, we combine stimulated Raman scattering (SRS) with 4Pi-interferometry to significantly improve the axial resolution by nearly 7-fold. We report on the characterization of improvements in imaging sensitivity and axial resolution using 80 nm polystyrene beads. Harnessing the improved axial resolution, we demonstrate super-resolution 4Pi-SRS imaging in resolving small lipid droplet structures in mammalian cells and lipid membranes in E. coli cells. Because 4Pi-SRS uses interferometry to improve axial resolution, it is completely orthogonal to all previous super-resolution SRS techniques, including visible excitation, photoswitching, sample expansion, and computational approaches, thus it is straightforward to combine them to achieve much higher resolution chemical imaging than currently possible.

bioengineering↗

bioRxiv2023

The epigenetic response of mouse brown adipose tissue to cold stress: histone proteoform, DNA methylation, and RNA expression

Regulation of the thermogenic response by brown adipose tissue (BAT) is an important component of energy homeostasis with implications for the treatment of obesity and diabetes. Our preliminary analyses uncovered many nodes representing epigenetic modifiers that are altered in BAT in response to chronic thermogenic activation. Thus, we hypothesized that chronic thermogenic activation broadly alters epigenetic modifications of DNA and histones in BAT. Motivated to understand how BAT function is regulated epigenetically, we developed a novel method for the first-ever unbiased top- down proteomic quantitation of histone modifications in BAT and validated our results with a multi-omic approach. To test our hypothesis, wildtype male C57BL/6J mice were housed under chronic conditions of thermoneutral temperature (TN, 28.8{degrees}C), mild cold/room temperature (RT, 22{degrees}C), or severe cold (SC, 8{degrees}C) and BAT was analyzed for DNA methylation and histone modifications. Methylation of promoters and intragenic regions in genomic DNA decrease in response to chronic cold exposure. Integration of DNA methylation and RNA expression data suggest a role for epigenetic modification of DNA in gene regulation in response to cold. In response to cold housing, we observe increased bulk acetylation of histones H3.2 and H4, increased histone H3.2 proteoforms with di- and trimethylation of lysine 9 (K9me2 and K9me3), and increased histone H4 proteoforms with acetylation of lysine 16 (K16ac) in BAT. Taken together, our results reveal global epigenetically-regulated transcriptional "on" and "off" signals in murine BAT in response to varying degrees of chronic cold stimuli and establish a novel methodology to quantitatively study histones in BAT, allowing for direct comparisons to decipher mechanistic changes during the thermogenic response. Additionally, we make histone PTM and proteoform quantitation, RNA splicing, RRBS, and transcriptional footprint datasets available as a resource for future research.

systems biology↗

bioRxiv2020

COPB2 haploinsufficiency causes a coatopathy with osteoporosis and developmental delay

Coatomer complexes function in the sorting and trafficking of proteins between subcellular organelles. Pathogenic variants in coatomer subunits or associated factors have been reported in multi-systemic disorders, i.e., coatopathies, that can affect the skeletal and central nervous systems. We have identified loss-of-function variants in COPB2, a component of the coatomer complex I (COPI), in individuals presenting with osteoporosis, fractures and developmental delay of variable severity. Because the role of COPB2 in bone has not been characterized, we studied the effect of COPB2 deficiency on skeletal development in mice and zebrafish. Copb2+/- mice showed low bone mass and decreased bone strength. In zebrafish, larvae carrying a copb2 heterozygous frameshift variant showed delayed mineralization. copb2-null embryos showed endoplasmic reticulum (ER) and Golgi disorganization, and embryonic lethality. COPB2 siRNA-treated fibroblasts showed delayed collagen trafficking with retention of type I collagen in the ER and Golgi, and altered distribution of Golgi markers. Our data suggest that COPB2 haploinsufficiency leads to disruption of intracellular collagen trafficking and osteoporosis, which may improve with ascorbic acid supplementation. This work highlights the role of COPI complex as a critical regulator of bone mass and identifies a new form of coatopathy due to COPB2 deficiency.

genetics↗