Regulation of LRH-1 cellular trafficking: Role of Nuclear localization signals and sumoylation

• Main Authors: Feng-Ming Yang, 楊豐名
• Other Authors: 胡孟君
• Format: Others
• Language: en_US
• Published: 2010
• Online Access: http://ndltd.ncl.edu.tw/handle/62581164501326227113

博士 === 國立臺灣大學 === 生理學研究所 === 99 === Liver receptor homolog-1 (LRH-1; NR5A2), is an orphan nuclear receptor that is localized to the nucleus for the regulation of gene expression. In this study, I characterized two functional nuclear localization signals (NLSs) in LRH-1. The first motif, NLS1 (residues 117-168), overlaps the second zinc finger in the DNA binding domain. Mutagenesis analysis showed that the zinc finger structure and two basic clusters on either side of the zinc finger loop are critical for nuclear localization directed by NLS1. The motif NLS2 (residues 169-204) is located in the Ftz-F1 box that contains a bipartite signal. Either NLS1 or NLS2 alone was sufficient to target full-length LRH-1 to the nucleus. Both NLS1 and NLS2 interacted with importin α and importin β in vitro, suggesting that importins mediate the nuclear transport of LRH-1. Three crucial basic clusters in the NLSs are also involved in the DNA binding and transcriptional activities of LRH-1. I further showed that nuclear localization of LRH-1 was regulated by SUMO modification. SUMO conjugation targets LRH-1 to the transcriptionally inactive nuclear bodies. Lysine 289 is the major site for SUMO-1 conjugation in vitro. However, in COS-7 cells, mutation of either K173 or K289 prevented translocation of LRH-1 into nuclear bodies and reduced the conjugation by SUMO-1, suggesting that K289 and K173 are two important sites involved in SUMO-1 modification. In granulosa cells, three or more altered lysine residues were required for nucleoplasm retention. This result suggests that multiple lysine residues are targets for SUMO conjugation in vivo and that granulosa cells are more sensitive to SUMO-1-mediated LRH-1 localization to nuclear bodies. Nuclear body localization of LRH-1 was suppressed by forskolin and cholera toxin. Forskolin treatment could change the dynamic process of sumoylation and repress LRH-1 targeting to nuclear speckles in rat granulosa cells. Take together, the result of this study suggest that transcription function of LRH-1 is maintained via the nucleocytoplasmic transport system and regulated by sumoylation.