title: Sumoylation activates the transcriptional activity of Pax-6, an important transcription factor for eye and brain development.
abstract: Pax-6 is an evolutionarily conserved transcription factor regulating brain and eye development. Four Pax-6 isoforms have been reported previously. Although the longer Pax-6 isoforms (p46 and p48) bear two DNA-binding domains, the paired domain (PD) and the homeodomain (HD), the shorter Pax-6 isoform p32 contains only the HD for DNA binding. Although a third domain, the proline-, serine- and threonine-enriched activation (PST) domain, in the C termini of all Pax-6 isoforms mediates their transcriptional modulation via phosphorylation, how p32 Pax-6 could regulate target genes remains to be elucidated. In the present study, we show that sumoylation at K91 is required for p32 Pax-6 to bind to a HD-specific site and regulate expression of target genes. First, in vitro-synthesized p32 Pax-6 alone cannot bind the P3 sequence, which contains the HD recognition site, unless it is preincubated with nuclear extracts precleared by anti-Pax-6 but not by anti-small ubiquitin-related modifier 1 (anti-SUMO1) antibody. Second, in vitro-synthesized p32 Pax-6 can be sumoylated by SUMO1, and the sumoylated p32 Pax-6 then can bind to the P3 sequence. Third, Pax-6 and SUMO1 are colocalized in the embryonic optic and lens vesicles and can be coimmunoprecipitated. Finally, SUMO1-conjugated p32 Pax-6 exists in both the nucleus and cytoplasm, and sumoylation significantly enhances the DNA-binding ability of p32 Pax-6 and positively regulates gene expression. Together, our results demonstrate that sumoylation activates p32 Pax-6 in both DNA-binding and transcriptional activities. In addition, our studies demonstrate that p32 and p46 Pax-6 possess differential DNA-binding and regulatory activities.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, NE 68198, USA.
authors.forename: Qin
authors.initials: Q
authors.lastname: Yan
authors.forename: Lili
authors.initials: L
authors.lastname: Gong
authors.forename: Mi
authors.initials: M
authors.lastname: Deng
authors.forename: Lan
authors.initials: L
authors.lastname: Zhang
authors.forename: Shuming
authors.initials: S
authors.lastname: Sun
authors.forename: Jiao
authors.initials: J
authors.lastname: Liu
authors.forename: Haili
authors.initials: H
authors.lastname: Ma
authors.forename: Dan
authors.initials: D
authors.lastname: Yuan
authors.forename: Pei-Chao
authors.initials: PC
authors.lastname: Chen
authors.forename: Xiaohui
authors.initials: X
authors.lastname: Hu
authors.forename: Jinping
authors.initials: J
authors.lastname: Liu
authors.forename: Jichao
authors.initials: J
authors.lastname: Qin
authors.forename: Ling
authors.initials: L
authors.lastname: Xiao
authors.forename: Xiao-Qin
authors.initials: XQ
authors.lastname: Huang
authors.forename: Jian
authors.initials: J
authors.lastname: Zhang
authors.forename: David Wan-Cheng
authors.initials: DW
authors.lastname: Li
mesh_terms: D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000254:growth & development*; D004247:DNA / Q000378:metabolism; D005123:Eye / Q000254:growth & development*; D005136:Eye Proteins / Q000235:genetics*; D005786:Gene Expression Regulation; D018398:Homeodomain Proteins / Q000235:genetics*; D051379:Mice; D000071841:PAX6 Transcription Factor; D051761:Paired Box Transcription Factors / Q000235:genetics*; D011485:Protein Binding; D020033:Protein Isoforms; D012097:Repressor Proteins / Q000235:genetics*; D025842:SUMO-1 Protein / Q000378:metabolism; D058207:Sumoylation / Q000502:physiology*; D014157:Transcription Factors; D015533:Transcriptional Activation
chemical_list: D005136:Eye Proteins; D018398:Homeodomain Proteins; D000071841:PAX6 Transcription Factor; D051761:Paired Box Transcription Factors; C000605732:Pax6 protein, mouse; D020033:Protein Isoforms; D012097:Repressor Proteins; D025842:SUMO-1 Protein; D014157:Transcription Factors; D004247:DNA