title: Characterization of the Role of MANF in Regulating the Secretion of CRELD2.
abstract: We recently demonstrated that the secretion of two novel endoplasmic reticulum (ER) stress-inducible proteins, cysteine-rich with epidermal growth factor (EGF)-like domains 2 (CRELD2) and mesencephalic astrocyte-derived neurotrophic factor (MANF), are oppositely regulated by the overexpression of 78 kDa glucose-regulated protein (GRP78). In the present study, we found that the co-transfection of CRELD2 and MANF remarkably enhanced the secretion of CRELD2 without affecting the expression level of GRP78. To identify the structural features of CRELD2 and MANF involved in this process, we generated several CRELD2 and MANF expression constructs. The deletion of the four C-terminal amino acids, either REDL in CRELD2 or RTDL in MANF, abolished the increased secretion of CRELD2 induced by the co-expression of MANF. The deleted mutation of MANF partially abolished the increased secretion of wild type CRELD2 (wtCRELD2) as a positive action of wild type MANF (wtMANF), even when we added the amino acid sequence RTDL at the C-terminus of each mutated MANF construct. Enhanced green fluorescent protein (EGFP), which was tagged with the signal peptide sequence at the N-terminus and four C-terminal amino acids (KEDL, REDL or RTDL), were retained intracellularly, but they did not enhance the secretion of wtCRELD2. Taken together, our data demonstrate that MANF is a factor in regulating the secretion of CRELD2 through four C-terminal amino acids, RTDL and REDL, and the fluctuation of intracellular MANF seems to potentiate the secretion of CRELD2.
authors.affiliation: Department of Chemistry and Biomolecular Science, Faculty of Engineering, Gifu University; 1–1 Yanagido, 2. United Graduate School of Drug Discovery and Medical Information Sciences, Gifu University.
authors.forename: Kentaro
authors.initials: K
authors.lastname: Oh-hashi
authors.forename: Junpei
authors.initials: J
authors.lastname: Norisada
authors.forename: Yoko
authors.initials: Y
authors.lastname: Hirata
authors.forename: Kazutoshi
authors.initials: K
authors.lastname: Kiuchi
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism; D000818:Animals; D019556:COS Cells; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D002522:Chlorocebus aethiops; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D059865:Endoplasmic Reticulum Stress; D016326:Extracellular Matrix Proteins / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D057809:HEK293 Cells; D006360:Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D009414:Nerve Growth Factors / Q000378:metabolism*; D014162:Transfection
chemical_list: D000596:Amino Acids; C507070:CRELD2 protein, human; D015815:Cell Adhesion Molecules; D000091342:Endoplasmic Reticulum Chaperone BiP; D016326:Extracellular Matrix Proteins; C000717247:HSPA5 protein, human; D006360:Heat-Shock Proteins; C099695:MANF protein, human; D009414:Nerve Growth Factors; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins