title: Nuclear translocation of CBP/p300-interacting protein CITED1 induced by parathyroid hormone requires serine phosphorylation at position 79 in its 63-84 domain.
abstract: The transcriptional cofactor CITED1 inhibits osteoblastic differentiation and blunts the stimulation of osteoblastic differentiation by parathyroid hormone (PTH). In the MC3T3-E1 osteoblastic cell line, we found that CITED1 was located predominantly in the cytoplasm and that hPTH(1-34) increased translocation of CITED1 from the cytoplasm to the nucleus. This response to hPTH(1-34) was not observed when all 9 serine residues within the 63-84 domain of CITED1 were mutated to alanines (CITED1 9S>A) or when a single serine to alanine mutation was made at position 79 (CITED1 S(79)>A). CITED1 containing mutations of these 9 serines to glutamic acid (9S>E) retained the same nuclear translocation response to hPTH(1-34) as the wild type CITED1. ALP activity and formation of mineralized nodules were inhibited in cells transfected with pcDNA3-CFP-CITED1 or with pcDNA3-CFP-CITED1 9S>E with or without hPTH(1-34) treatment (all P<0.05); these changes were not observed using CITED1 9S>A. Cells exposed to intermittent treatment with hPTH(1-34) expressed more ALP2, Runx2 and osteocalcin than vehicle-treated cells. These effects of hPTH(1-34) were inhibited in cells transfected with pcDNA3-CFP-CITED1 or pcDNA3-CFP-CITED1 9S>E, but were slightly enhanced by the alanine mutants. PKC activator (TPA) increased nuclear translocation of CITED1, whereas a PKC inhibitor (Go6983) blunted the effect of hPTH(1-34) on the nuclear translocation of wildtype CITED1 but not of CITED1 S(79)>E. The data indicated that serine phosphorylation at position 79 in the 63-84 domain is associated with PKC activation, and is required for both CITED1 nuclear translocation induced by PTH and the negative effects of CITED1 on osteoblastic differentiation and mineralization.
authors.affiliation: Department of Spinal Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong 510515, China.
authors.forename: Zhen
authors.initials: Z
authors.lastname: Lin
authors.affiliation: Department of Spinal Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong 510515, China.
authors.forename: Ruiqiang
authors.initials: R
authors.lastname: Feng
authors.affiliation: Department of Spinal Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong 510515, China.
authors.forename: Junqing
authors.initials: J
authors.lastname: Li
authors.affiliation: Department of Spinal Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong 510515, China.
authors.forename: Yue
authors.initials: Y
authors.lastname: Meng
authors.affiliation: Department of Spinal Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong 510515, China.
authors.forename: Liang
authors.initials: L
authors.lastname: Yuan
authors.affiliation: Department of Spinal Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong 510515, China.
authors.forename: Zhaozong
authors.initials: Z
authors.lastname: Fu
authors.affiliation: Endocrine Unit, Their 11, Massachusetts General Hospital, Boston, MA 02114, USA.
authors.forename: Jun
authors.initials: J
authors.lastname: Guo
authors.affiliation: Endocrine Unit, Their 11, Massachusetts General Hospital, Boston, MA 02114, USA.
authors.forename: F Richard
authors.initials: FR
authors.lastname: Bringhurst
authors.affiliation: Department of Spinal Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong 510515, China. Electronic address: drmyang@yahoo.com.
authors.forename: Dehong
authors.initials: D
authors.lastname: Yang
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000502:physiology; D019943:Amino Acid Substitution; D000818:Animals; D051017:Apoptosis Regulatory Proteins; D002113:Calcification, Physiologic / Q000187:drug effects / Q000502:physiology*; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology*; D002460:Cell Line; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism*; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D008301:Maleimides / Q000494:pharmacology; D051379:Mice; D020125:Mutation, Missense; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism*; D010281:Parathyroid Hormone / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects / Q000502:physiology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D047428:Protein Kinase Inhibitors; D012694:Serine / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*
chemical_list: C465664:2-(1-(3-dimethylaminopropyl)-5-methoxyindol-3-yl)-3-(1H-indol-3-yl)maleimide; D051017:Apoptosis Regulatory Proteins; C102208:Cited1 protein, mouse; D050659:Core Binding Factor Alpha 1 Subunit; D007211:Indoles; D008301:Maleimides; D009687:Nuclear Proteins; D010281:Parathyroid Hormone; D047428:Protein Kinase Inhibitors; C493716:Runx2 protein, mouse; D015534:Trans-Activators; D012694:Serine; D011493:Protein Kinase C
keywords: CITED1; Nuclear translocation; Osteoblastic differentiation; Osteoblastic mineralization; Parathyroid hormone; Phosphorylation