title: A Comparative Analysis of the Molecular Features of MANF and CDNF.
abstract: Cerebral dopamine neurotrophic factor (CDNF) is a paralogous protein of mesencephalic astrocyte-derived neurotrophic factor (MANF). Both proteins have been reported to show a common cytoprotective effect on dopaminergic neurons as a secretory protein containing the KDEL-like motif of the ER retrieval signal at the C-terminus, RTDL in MANF and [Q/K]TEL in CDNF among many species, although functions of paralogous proteins tend to differ from each other. In this study, we focused on post-translational regulations of their retention in the endoplasmic reticulum (ER) and secretion and performed comparative experiments on characterization of mouse MANF and mouse CDNF according to our previous report about biosynthesis and secretion of mouse MANF using a NanoLuc system. In this study, co-expression of glucose-regulated protein 78 kDa (GRP78), KDEL receptor 1 or mutant Sar1 into HEK293 cells similarly decreased MANF and CDNF secretion with some degree of variation. Next, we investigated whether CDNF affects the secretion of mouse cysteine-rich with EGF-like domains 2 (CRELD2) because mouse wild-type (wt) MANF but not its KDEL-like motif deleted mutant (ΔCMANF) was found to promote the CRELD2 release from the transfected cells. Co-expressing CRELD2 with wt or ΔC CDNF, we found that CDNF and ΔCMANF hardly elevated the CRELD2 secretion. We then investigated effects of the four or six C-terminal amino acids of MANF and CDNF on the CRELD2 secretion. As a result, co-transfection of mouse CDNF having the mouse MANF-type C-terminal amino acids (CDNFRTDL and CDNFSARTDL) increased the CRELD2 secretion to a small extent, but mouse CDNF having human CDNF-type ones (CDNFKTEL and CDNFHPKTEL) well increased the CRELD2 secretion. On the other hand, the replacement of C-terminal motifs of mouse MANF with those of mouse CDNF (MANFQTEL and MANFYPQTEL) enhanced the CRELD2 secretion, and the mouse MANF having human CDNF-type ones (MANFKTEL and MANFHPKTEL) dramatically potentiated the CRELD2 secretion. These results indicate that the secretion of mouse MANF and mouse CDNF is fundamentally regulated in the same manner and that the variation of four C-terminal amino acids in the MANF and CDNF among species might influence their intracellular functions. This finding could be a hint to identify physiological functions of MANF and CDNF.
authors.affiliation: United Graduate School of Drug Discovery and Medical Information Sciences, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan.
authors.forename: Junpei
authors.initials: J
authors.lastname: Norisada
authors.affiliation: United Graduate School of Drug Discovery and Medical Information Sciences, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan.|Department of Chemistry and Biomolecular Science, Faculty of Engineering, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan.
authors.forename: Yoko
authors.initials: Y
authors.lastname: Hirata
authors.affiliation: Department of Anesthesiology, Kyoto Prefectural University of Medicine, Kawaramachi-Hirokoji, Kamikyo-ku, Kyoto 602-0841, Japan.
authors.forename: Fumimasa
authors.initials: F
authors.lastname: Amaya
authors.affiliation: United Graduate School of Drug Discovery and Medical Information Sciences, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan.|Department of Chemistry and Biomolecular Science, Faculty of Engineering, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan.
authors.forename: Kazutoshi
authors.initials: K
authors.lastname: Kiuchi
authors.affiliation: United Graduate School of Drug Discovery and Medical Information Sciences, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan.|Department of Chemistry and Biomolecular Science, Faculty of Engineering, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan.
authors.forename: Kentaro
authors.initials: K
authors.lastname: Oh-hashi
mesh_terms: D000818:Animals; D019556:COS Cells; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002522:Chlorocebus aethiops; D000091342:Endoplasmic Reticulum Chaperone BiP; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D015870:Gene Expression; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D020559:Monomeric GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D020125:Mutation, Missense; D009414:Nerve Growth Factors / Q000235:genetics / Q000378:metabolism*; D021382:Protein Sorting Signals; D021381:Protein Transport
chemical_list: C561361:CRELD2 protein, mouse; D015815:Cell Adhesion Molecules; D000091342:Endoplasmic Reticulum Chaperone BiP; D016326:Extracellular Matrix Proteins; C000717247:HSPA5 protein, human; C000717248:Hspa5 protein, mouse; C534671:MANF protein, mouse; D009414:Nerve Growth Factors; D021382:Protein Sorting Signals; C522218:cdnf protein, mouse; C493083:SAR1A protein, human; D020559:Monomeric GTP-Binding Proteins
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