Berberine induced autophagic cell death by elevating GRP78 level in cancer cells
作者单位：Institute of BiotechnologyKey Laboratory of Chemical Biology and Molecular Engineering of Nationa1 Ministry of EducationShanxi University Institutes of Biomedical SciencesShanxi University School of Life ScienceShanxi University
会议名称：《2017第七届泛环渤海生物化学与分子生物学会学术交流会》
会议日期：2017年
学科分类：1006[医学-中西医结合] 10[医学] 100602[医学-中西医结合临床]
关 键 词：berberine autophagy cancer cell death GRP78 VPS34
摘 要：Autophagy is the cellular process that delivers damaged organelles and invasive bacteria to lysosomes for degradation so as to maintain the intracellular homeostasis during various cell stresses.These substrates are engulfed along with bulk cytoplasm by double-membrane,with the extension and fusion of membrane edge,which results in the formation of the autophagosome.After the fusion of autophagosome and lysosome,the degradation products are recycled back into cytosol and reused to enhance cell survival during nutrient deprivation.One potential regulator of autophagy is the activation of the unfolded protein response(UPR),an endoplasmic reticulum stress pathway.GRP78,a key upstream activator of the UPR,participates in promoting protein folding.assembly and degradation,endoplasmic reticulum stress sensing,and cellular calcium homeostasis.High level of GRP78 contributes to the stress-induced autophagy and functional blockade of the proteasome induces GRP78 to promote autophagosome formation.As a major active compound of Coptidis Rhizoma(Huanglian in Chinese),berberine is an isoquinoline alkaloid,which has long been used as a nonprescription oral drug in China for the treatment of gut infections and diarrhea.Importantly,the antineoplastic activities of berberine were intensive studied and reported since 1990s.A great portion of studies made effort on revealing of cytotoxicity,metastasis inhibition and antiangiogenic of berberine.Other studies focus on cell-cycle checkpoint control,DNA repair and regulation of initiating oxidative stress in cancer cells,which bring some new mechanisms underlying the antitumor of berberine to light.Recent studies have revealed the berberine can trigger autophagic cell death of tumor,and its beneficial effects are receded when the autophagy process is genetically or pharmacologically inactivated,which suggests that autophagy is indispensable for the protective effects of berberine.Here,we reported that berberine could induce autophagic cancer cell death by the elevation of GRP78.The results further demonstrated that berberine enhanced GRP78 by the suppression of ubiquitination/proteasomal degradation of GRP78 and activation of ATF6.Hence,ATF6 may be the new target of berberine,which is responsible for berberine-induced GRP78 elevation.Moreover,fluorescence spectrumassay revealed thatberberine had strong binding with GRP78 and only one binding site exist in GRP78 with berberine at 37℃.At the same time,thermodynamics parameters and nature of the binding forces revealed that hydrogen bonding and vander Waals forces played a dominantrole in the binding of berberine to GRP78.Previous studies indicated that AMPK is a major intermediatein facilitating the beneficial effects of berberine.Indeed,we observedthe increased p-AMPKin berberine-treated HCT-116 and DLD1 cells,and siGRP78 largely decreased the berberine-induced p-AMPK level,implying AMPK may functionas a downstream regulator of GRP78.Thus,our study provides a link between AMPK and GRP78 in berberine-induced cancercell autophagic death.Finally,Co-IP analysis showed that the ability of GRP78 bind to VPS34 was increased with berberine treatment,revealing that GRP78 is the important regulator of autophagic paradox.Taken together,our results suggested that berberine induced autophagic cancer cell death via enhanced GRP78 level and the ability of GRP78 bind to VPS34,which may be a promising candidate for cancer therapy.