Maintenance of the mammalian inactive X chromosome

Research Project

Project/Area Number	13672372
Research Category	Grant-in-Aid for Scientific Research (C)
Allocation Type	Single-year Grants
Section	一般
Research Field	Human genetics
Research Institution	HOKKAIDO UNIVERSITY
Principal Investigator	YOSHIDA Ikuya Hokkaido Univ., CAST., Instructor, 先端科学技術共同研究センター, 助手 (90240275)
Project Period (FY)	2001 – 2002
Project Status	Completed (Fiscal Year 2002)
Budget Amount *help	¥3,600,000 (Direct Cost: ¥3,600,000) Fiscal Year 2002: ¥700,000 (Direct Cost: ¥700,000) Fiscal Year 2001: ¥2,900,000 (Direct Cost: ¥2,900,000)
Keywords	inactive X chromosome / heterochromatinization / embryonal carcinoma cells / mouse / histone acetylation / Xist RNA / late DNA replication / macroH2A1.2 / X染色体 / RNA
Research Abstract	The mouse embryonal carcinoma cell line MC12 carries two X chromosomes, one of which replicates late in S phase and shares properties with the normal inactive X chromosome, and is therefore considered to be inactivated. Since the hypoxanthine phosphoribosyl transferase (HPRT) gene on the active X chromosome is mutated (hprt^-), MC12 cells lack HPRT activity. After subjecting MC12 cells to selection in HAT medium, however, a number of HAT-resistant clones (HAT^R) appeared. The high frequency of HAT resistance (3.18x10^<-4>) suggested reactivation of hprt^+ on the inactive X chromosome rather than reversion of hprt^-. Consistent with this view, cytological analyses showed that the reactivation occurred over the length of the inactive X chromosome in 11 out of 20 HAT^R clones isolated. The remaining 9 clones retained a normal heterochromatic inactive X chromosome. The spontaneous reactivation rate of the hprt^+ on the inactive X chromosome was relatively high (1.34x10^<-6>) and comparable to that observed for Xist-deleted somatic cells (Csankovszki et al., 2001), suggesting that the inactivated state is poorly maintained in MC12 cells.