GRP78 regulates milk biosynthesis and the proliferation of bovinemammaryepithelial cells through the mTOR signaling pathway

Abstract

BackgroundGlucose-regulated protein 78 (GRP78) is a member of the HSP70 protein family and a key endoplasmic reticulum chaperone. It has been revealed to play important roles both in the maturation, folding and transport of proteins and in cellproliferation. However, its involvement in milk biosynthesis or the proliferation of bovine primary mammary epithelial cells (BMECs) has yet to be established.MethodsThe expressions of GRP78 in BMECs stimulated with methionine, leucine, estrogen and prolactin were determined using western blotting and immunofluorescence assays. To explore the function of GRP78 in BMECs, the protein was overexpressed or knocked down, respectively using an overexpression vector or an siRNA mixture transfected into cells cultured in vitro. Flow cytometry was used to analyze cell proliferation and cell activity. The contents of lactose and triglyceride (TG) secreted from the treated BMECs were measured using lactose and TG assay kits, respectively. Western blotting analysis was used to measure the β-casein content and the protein levels of the signaling molecules known to be involved in milk biosynthesis and cell proliferation.ResultsGRP78overexpression significantly stimulated milk protein and milk fat synthesis, enhanced cell proliferation, positively regulated the phosphorylation of mammalian target of rapamycin (mTOR), and increased the amount of protein of cyclinD1andsterol regulatory element-binding protein 1c (SREBP-1c). GRP78 knockdown after siRNA transfection had the opposite effects. We further found that GRP78 was located in the cytoplasm of BMECs, and that stimulating methionine, leucine, estrogen and prolactin expression led to a significant increase in the protein expression of GRP78 in BMECs.ConclusionsThese data reveal that GRP78 is an important regulator of milk biosynthesis and the proliferation of BMECs through the mTOR signaling pathway.

Hydrogen sulfide inhibits endoplasmic reticulum stress through the GRP78/mTOR pathway in rat chondrocytes subjected to oxidative stress

“…In the present study, it was hypothesized that exogenous H 2 S could protect cartilage against TBHP-induced oxidative stress. The present study relied on the novel finding that GYY4137 displays physicochemical properties that may be related to the inhibition of ER stress-induced apoptosis ( 14 , 17 , 29 ) via the downregulation of the expression levels of CHOP, GRP78 and ATF6.…”

“…ER stress induces cH apoptosis, which is associated with the pathogenesis of OA (27). GRP78 has been reported to serve as a critical mediator of ER stress, as demonstrated by decreased mTOR induction (28)(29)(30)(31)(32)(33). GRP78 was selected as a novel biomarker, as its level associated with cell process, cell integrity failure, and cell proliferation.…”

Kisspeptin-10 Maintains the Activation of the mTOR Signaling Pathway by Inhibiting SIRT6 to Promote the Synthesis of Milk in Bovine Mammary Epithelial Cells

“…mTOR is one of the important protein kinases. It can regulate a variety of physiological and pathological metabolisms in different tissues and organs through two downstream effectors, 4EBP1 and S6K. , In addition, studies have also shown that mTOR can promote lipid de novo synthesis through SREBP1, which controls the expression of genes involved in fat biosynthesis and decomposition. , Currently, mTOR is also used as the axial pathway for milk synthesis in BMECs . It has been proven that amino acids, fatty acids, and hormones can affect milk synthesis by regulating mTOR. , Therefore, the intensity of mTOR signal transduction directly determines the ability of milk synthesis in BMECs.…”

Schisandrin ameliorates cognitive deficits, endoplasmic reticulum stress and neuroinflammation in streptozotocin (STZ)-induced Alzheimer’s disease rats

“…The ER stress is mainly caused by the accumulation of unfolded or misfolded protein inside the ER, and excess ER stress response also leads to cell damages [ 12 ]. GRP78 is a vital endoplasmic reticulum chaperone which can function in various processes except for ER stress including cell proliferation, protein maturation, folding and transport [ 17 ]. CHOP is a key molecule in the downstream of PERK, and ER stress can be mediated by the PERK-CHOP pathway [ 11 ].…”