Shah, R. A. | bioRxiv Science

bioRxiv2022

Scavenger receptor B1 facilitates the endocytosis of Escherichia coli via TLR4 signaling in mammary gland infection

SCARB1 belongs to class B of Scavenger receptors (SRs) that are known to be involved in binding and endocytosis of various pathogens. SRs have emerging role in regulating innate immunity and host-pathogen interactions by acting in co-ordination with Toll-like receptors. Little is known about the function of SCARB1 in milk-derived mammary epithelial cells (MECs). This study reports the role of SCARB1 in infection and its potential association in TLR4 signaling on bacterial challenge in Goat mammary epithelial cells (GMECs). The novelty in the establishment of MEC culture lies in the method that aims to enhance the viability of the cells with intact characteristics upto a higher passage number. We represent MEC culture to be used as a potential infection model for deeper understanding of animal physiology especially around the mammary gland. On E.coli challenge the expression of SCARB1 was significant in induced GMECs at 6 h. Endoribonuclease-esiRNA based silencing of SCARB1 affects the expression of TLR4 and its pathways i.e. MyD88 and TRIF pathways on infection. Knockdown also affected the endocytosis of E.coli in GMECs demonstrating that E.coli uses SCARB1 function to gain entry in cells. Furthermore, we predict 3 unique protein structures of uncharacterized SCARB1 (Capra hircus) protein. Overall, we highlight SCARB1 as a main participant in host defence and its function in antibacterial advances to check mammary gland infections.

immunology↗

bioRxiv2022

Comparison of co-immunoprecipitation techniques for effective identification of SNTA1 interacting proteins in breast cancer cells

Diverse signal transduction pathways involve thousands of proteins acting in concert that contribute to breast cancer pathology. SNTA1 has emerged as a common link that functions to wire various complex signaling circuits within the cell. In our study we used endogenous SNTA1 protein as bait in pull-down assays to identify SNTA1 interactome from breast cancer cell extract. We compared the utility of two co-immunoprecipitation techniques conventional co-IP and cross-linking co-IP coupled to mass spectrometry (IP-MS) that intriguingly generated valuable inventory of SNTA1 associated proteins in breast cancer cells that may or may not interact physically but contribute to shared functions in breast cancer pathology. We observed that Pierce cross-link IP kit enhanced sensivity of the analysis as it decreased the number of background proteins with a remarkable increase in identified list of proteins. We have also observed that different approaches are able to consistently detect the same signaling outcome as observed by Gene Ontology. Identification of interacting partners of SNTA1 in breast cancer cell lines may depict the possible signaling mechanism of this protein in molecular pathogenesis of breast cancer and its possible use as a therapeutic target.

cancer biology↗