不同的系统发育和底物特异性表明12种拟南芥卵巢肿瘤结构域(OTU)去泛素酶参与保守或植物特异性功能。otu5-1无效突变体表现出多效表型,包括早期开花,模仿在亚基中具有缺陷的突变体(例如,参与组蛋白H2A的SWR1复合物(SWR1c)的ARP6)。Z沉积。转录组和RT-qPCR分析表明,下调的FLC和MAF4-5负责otu5-1的早期开花。qChIP分析显示,激活和抑制组蛋白标记减少和增加,分别,在otu5-1中的FLC和MAF4-5上。亚细胞分馏,GFP融合表达,和MNase处理染色质表明OTU5是富含细胞核和染色质相关的。此外,发现OTU5与FLC和MAF4-5相关。OTU5相关蛋白复合物似乎与SWR1c不同,因为在arp6-1植物中OTU5复合物的分子量没有改变。此外,otu5-1arp6-1双突变体表现出协同表型,H2A。FLC/MAF4-5上的Z水平在arp6-1中降低,但在otu5-1中没有降低。我们的结果支持拟南芥OTU5独立于SWR1c起作用的命题,通过组蛋白修饰激活FLC和MAF4-5来抑制开花。双突变分析还表明OTU5独立于HUB1介导的途径起作用,但在自主途径突变体和FRIGIDA-Col中,FLC介导的开花抑制是部分必需的。
Distinct phylogeny and substrate specificities suggest that 12 Arabidopsis Ovarian Tumor domain-containing (OTU) deubiquitinases participate in conserved or plant-specific functions. The otu5-1 null mutant displayed a pleiotropic phenotype, including early flowering, mimicking that of mutants harboring defects in subunits (e.g., ARP6) of the SWR1 complex (
SWR1c) involved in histone H2A.Z deposition. Transcriptome and RT-qPCR analyses suggest that downregulated FLC and MAF4-5 are responsible for the early flowering of otu5-1. qChIP analyses revealed a reduction and increase in activating and repressive histone marks, respectively, on FLC and MAF4-5 in otu5-1. Subcellular fractionation, GFP-fusion expression, and MNase treatment of chromatin showed that OTU5 is nucleus-enriched and chromatin-associated. Moreover, OTU5 was found to be associated with FLC and MAF4-5. The OTU5-associated protein complex(es) appears to be distinct from
SWR1c, as the molecular weights of OTU5 complex(es) were unaltered in arp6-1 plants. Furthermore, the otu5-1 arp6-1 double mutant exhibited synergistic phenotypes, and H2A.Z levels on FLC/MAF4-5 were reduced in arp6-1 but not otu5-1. Our results support the proposition that Arabidopsis OTU5, acting independently of
SWR1c, suppresses flowering by activating FLC and MAF4-5 through histone modification. Double-mutant analyses also indicate that OTU5 acts independently of the HUB1-mediated pathway, but it is partially required for FLC-mediated flowering suppression in autonomous pathway mutants and FRIGIDA-Col.