Biology subjects

Shekar, P. C.

Publications and source records attributed to Shekar, P. C..

4 recordsLinked to original sources

bioRxiv2023

Derivation of trophoblast stem cells unveils unrestrained potential of mouse ESCs and epiblast

mESCs and epiblast are considered to follow strict lineage adherence and lack the potential to contribute to trophoectoderm. Here, we report the derivation of trophoblast stem cells (ESTS) from the mESCs. The single-cell transcriptome and molecular characterization of ESTS show similarity with TSCs. They efficiently integrate into the TE compartment of the blastocyst and contribute to the placenta during development. We discovered GSK3{beta} as a critical regulator of the TE fate of ESCs. It plays a vital stage-specific role during ESTS derivation. We further show {beta}-CATENIN and an intron-I regulatory element of Cdx2 are essential for the TE fate of ESCs. We further show that the mouse epiblast can readily differentiate into TE lineage. In contrast to the paradigm of the restricted potential of pluripotent ESCs and epiblast, our data shows that murine ESCs and epiblast have the unrestrained developmental potential for extraembryonic lineages.

developmental biology↗

bioRxiv2022

DIP2 is a unique regulator of diacylglycerol lipid homeostasis in eukaryotes

Chain-length specific subsets of diacylglycerol (DAG) lipids are proposed to regulate differential physiological responses ranging from signal transduction to modulation of the membrane properties. However, the mechanism or molecular players regulating the subsets of DAG species remains unknown. Here, we uncover the role of a conserved eukaryotic protein family, DISCO-interacting protein 2 (DIP2) as a homeostatic regulator of a chemically distinct subset of DAGs using yeast, fly and mouse models. Genetic and chemical screens along with lipidomics analysis in yeast reveal that DIP2 prevents the toxic accumulation of specific DAGs in the logarithmic growth phase, which otherwise leads to endoplasmic reticulum stress. We also show that the fatty acyl-AMP ligase-like domains of DIP2 are essential for the redirection of the flux of DAG subspecies to storage lipid, triacylglycerols. Such modulation of selective DAG abundance by DIP2 is found to be crucial for optimal vacuole-membrane fusion and consequently osmoadaptation in yeast. Thus, the study illuminates an unprecedented DAG metabolism route and provides new insights on how cell fine-tunes DAG subspecies for cellular homeostasis and environmental adaptation.

biochemistry↗

bioRxiv2022

TEX13B is important for germ cell development and male fertility

The recent epidemiological studies suggest that nearly one out of every 7 reproductive age couples face problem to conceive a child after trying for at least one year. Impaired fertility of the male partner is causative in approximately 50% of the infertile couples. However, the etiologies of large proportion of male infertility are still unclear. Our unpublished exome sequencing data identified several novel genes including TEX13B, which motivated us to further explore the role of TEX13B in male infertility in large infertile case control cohort. Hence in this study, we have examined the role of TEX13B in male infertility by whole gene sequencing 628 infertile and 427 control men and have demonstrated the functional role of Tex13b in spermatogonia GC1spg (GC1) cells. We identified 2 variants on TEX13B which are tightly associated with male infertility. TEX13B gene exclusively expressed in germ cells, but its molecular functions in germ cells are still unknown. Hence, we demonstrated the functional importance of Tex13b in GC1 cell line by genomic manipulation via CRISPR-Cas9 and mass spectrometry-based whole cell proteomics. The gene knock out in GC1 cell line clearly shows that Tex13b play an important role in germ cell growth and morphology. We demonstrate that Tex13b knockout or conditional overexpression in GC1 cells reprograms the metabolic status from an oxidative phosphorylation to glycolysis state and vice versa. In conclusion, our study clearly showed the importance of Tex13b in germ cells development and Its association with male infertility.

genetics↗

bioRxiv2021

FGF/ERK autocrine signaling is enhanced by NANOG in a subpopulation of pluripotent stem cells to execute autoregulation and induce heterogeneity

The self-renewal and differentiation potential of Embryonic stem cells (ESCs) is maintained by the regulated expression of core pluripotency factors. The expression level of core pluripotency factor Nanog is tightly regulated by a negative feedback autorepression loop. However, it remains unclear how the ESCs perceive the NANOG levels and execute autorepression. Here, we show that a dose-dependent induction of Fgfbp1 and Fgfr2 by NANOG activates an autocrine mediated ERK signaling in high-Nanog cells to trigger autorepression. pERK recruits NONO to Nanog locus to repress transcription by preventing POL2 loading. The Nanog autorepression process establishes a self-perpetuating NANOG-pERK reciprocal regulatory circuit. We further demonstrate that the reciprocal regulatory circuit induces the pERK heterogeneity and ERK signaling dynamics in pluripotent stem cells.

cell biology↗