title: PCP4: a regulator of aldosterone synthesis in human adrenocortical tissues.
abstract: Purkinje cell protein 4 (PCP4) is a calmodulin (CaM)-binding protein that accelerates calcium association and dissociation with CaM. It has been previously detected in aldosterone-producing adenomas (APA), but details on its expression and function in adrenocortical tissues have remained unknown. Therefore, we performed the immunohistochemical analysis of PCP4 in the following tissues: normal adrenal (NA; n=15), APA (n=15), cortisol-producing adenomas (n=15), and idiopathic hyperaldosteronism cases (IHA; n=5). APA samples (n=45) were also submitted to quantitative RT-PCR of PCP4, CYP11B1, and CYP11B2, as well as DNA sequencing for KCNJ5 mutations. Transient transfection analysis using PCP4 siRNA was also performed in H295R adrenocortical carcinoma cells, following ELISA analysis, and CYP11B2 luciferase assays were also performed after PCP4 vector transfection in order to study the regulation of PCP4 protein expression. In our findings, PCP4 immunoreactivity was predominantly detected in APA and in the zona glomerulosa of NA and IHA. In APA, the mRNA levels of PCP4 were significantly correlated with those of CYP11B2 (P<0.0001) and were significantly higher in cases with KCNJ5 mutation than WT (P=0.005). Following PCP4 vector transfection, CYP11B2 luciferase reporter activity was significantly higher than controls in the presence of angiotensin-II. Knockdown of PCP4 resulted in a significant decrease in CYP11B2 mRNA levels (P=0.012) and aldosterone production (P=0.011). Our results indicate that PCP4 is a regulator of aldosterone production in normal, hyperplastic, and neoplastic human adrenocortical cells.
authors.affiliation: Department of Pathology, Tohoku University Graduate School of Medicine, 2-1 Seiryo-machi, Aoba-ku, Sendai, Miyagi 980-8575, Japan Division of Nephrology, Endocrinology and Vascular Medicine, Tohoku University Hospital, Sendai, Japan Departments of Diagnostic Radiology Molecular Endocrinology, Tohoku University Graduate School of Medicine, 2-1 Seiryo-machi, Aoba-ku, Sendai, Miyagi 980-8575, Japan Department of Physiology and Medicine, University of Michigan, Ann Arbor, Michigan, USA.
authors.forename: Saulo J A
authors.initials: SJ
authors.lastname: Felizola
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Nakamura
authors.forename: Yoshikiyo
authors.initials: Y
authors.lastname: Ono
authors.forename: Kanako
authors.initials: K
authors.lastname: Kitamura
authors.forename: Kumi
authors.initials: K
authors.lastname: Kikuchi
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Onodera
authors.forename: Kazue
authors.initials: K
authors.lastname: Ise
authors.forename: Kei
authors.initials: K
authors.lastname: Takase
authors.forename: Akira
authors.initials: A
authors.lastname: Sugawara
authors.forename: Namita
authors.initials: N
authors.lastname: Hattangady
authors.forename: William E
authors.initials: WE
authors.lastname: Rainey
authors.forename: Fumitoshi
authors.initials: F
authors.lastname: Satoh
authors.forename: Hironobu
authors.initials: H
authors.lastname: Sasano
mesh_terms: D000236:Adenoma / Q000235:genetics / Q000473:pathology; D000302:Adrenal Cortex / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D000306:Adrenal Cortex Neoplasms / Q000235:genetics / Q000473:pathology; D000450:Aldosterone / Q000096:biosynthesis*; D000804:Angiotensin II / Q000494:pharmacology; D015153:Blotting, Western; D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology; D019405:Cytochrome P-450 CYP11B2 / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D057075:Enzyme Assays; D004797:Enzyme-Linked Immunosorbent Assay; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007150:Immunohistochemistry; D008156:Luciferases / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D013252:Steroid 11-beta-Hydroxylase / Q000235:genetics / Q000378:metabolism; D014162:Transfection
chemical_list: D009419:Nerve Tissue Proteins; D034741:RNA, Small Interfering; C050394:PCP4 protein, human; D000804:Angiotensin II; D005576:Colforsin; D000450:Aldosterone; D004247:DNA; D008156:Luciferases; D019405:Cytochrome P-450 CYP11B2; D013252:Steroid 11-beta-Hydroxylase
keywords: CYP11B2; Purkinje cell protein 4 (PCP4); adrenal cortex; aldosterone; calmodulin (CaM)