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Distinct phylogeny and substrate specificities suggest that 12 Arabidopsis Ovarian Tumor domain-containing (OTU) deubiquitinases participate in conserved or plant-specific functions. The <i>otu5-1</i> 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 <i>FLC</i> and <i>MAF4-5</i> are responsible for the early flowering of <i>otu5-1</i>. qChIP analyses revealed a reduction and increase in activating and repressive histone marks, respectively, on <i>FLC</i> and <i>MAF4-5</i> in <i>otu5-1</i>. 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 <i>FLC</i> and <i>MAF4-5</i>. The OTU5-associated protein complex(es) appears to be distinct from SWR1c, as the molecular weights of OTU5 complex(es) were unaltered in <i>arp6-1</i> plants. Furthermore, the <i>otu5-1 arp6-1</i> double mutant exhibited synergistic phenotypes, and H2A.Z levels on <i>FLC/MAF4-5</i> were reduced in <i>arp6-1</i> but not <i>otu5-1</i>. Our results support the proposition that Arabidopsis OTU5, acting independently of SWR1c, suppresses flowering by activating <i>FLC</i> and <i>MAF4-5</i> through histone modification. Double-mutant analyses also indicate that <i>OTU5</i> acts independently of the HUB1-mediated pathway, but it is partially required for <i>FLC</i>-mediated flowering suppression in autonomous pathway mutants and <i>FRIGIDA</i>-Col.