Drosophila Ulp1, a Nuclear Pore-associated SUMO Protease, Prevents Accumulation of Cytoplasmic SUMO Conjugates

Abstract

SUMO is a small ubiquitin-like protein that becomes covalently conjugated to a variety of target proteins, the large majority of which are found in the nucleus. Ulp1 is a member of a family of proteases that control SUMO function positively, by catalyzing the proteolytic processing of SUMO to its mature form, and negatively, by catalyzing SUMO deconjugation. In Drosophila S2 cells, depletion of Ulp1 by RNA interference results in a dramatic change in the overall spectrum of SUMO conjugates, indicating that SUMO deconjugation is substratespecific and plays a critical role in determining the steady state targets of SUMO conjugation. Ulp1 normally serves to prevent the accumulation of SUMO-conjugated forms of a number of proteins, including the aminoacyl-tRNA synthetase EPRS. In the presence of Ulp1, most SUMO conjugates reside in the nucleus. However, in its absence, SUMO-conjugated EPRS accumulates in the cytoplasm, contributing to an overall shift of SUMO from the nucleus to the cytoplasm. The ability of Ulp1 to restrict SUMO conjugates to the nucleus is independent of its role as a SUMO-processing enzyme because Ulp1-dependent nuclear localization of SUMO is even observed when SUMO is expressed in a preprocessed form. Studies of a Ulp1-GFP fusion protein suggest that Ulp1 localizes to the nucleoplasmic face of the nuclear pore complex. We hypothesize that, as a component of the nuclear pore complex, Ulp1 may prevent proteins from leaving the nucleus with SUMO still attached.The regulation of protein function by reversible post-translational modification is an effective means for regulating protein stability, activity, subcellular localization, and interaction specificity. One such modification involves the covalent attachment of the small ubiquitin-like modifier protein (SUMO) 1 (also called Smt3), to its target proteins via a series of enzymatic steps that closely resemble the reactions of the ubiquitylation pathway (1-3).Despite the sequence and structural homology between SUMO and ubiquitin, it is clear that sumoylation and ubiquitylation have distinct functional consequences. Ubiquitylation most frequently results in degradation of the substrate protein through targeting to the 26 S proteasome (4). The consequences of sumoylation are diverse and target-specific, with no evidence to suggest a role in promoting protein degradation. Conjugation of SUMO to RanGAP1 is required to target it to the nuclear pore complex (NPC), where it makes a stable interaction with the Ran-binding protein RanBP2 (5-7). Sumoylation of PML is essential for its recruitment to discrete intranuclear foci called PML oncogenic domains (PODs) (8 -11). For some sumoylation targets, including Smad4 and IB␣, this modification appears to increase protein stability by blocking ubiquitin-mediated degradation (12, 13). Finally, an increasing number of DNA binding transcription factors have been identified as substrates for sumoylation (14,15). Sumoylation of transcription factors can alter their activity in multiple ways. For example, in the ca...

Medea SUMOylation restricts the signaling range of the Dpp morphogen in the Drosophila embryo

“…Together, the data presented in Figure 5, B and C, strongly support the conclusion that Med is SUMO modified in the nucleus. This interpretation is also compatible with the existing data that SUMOylation is predominantly a nuclear process both in mammalian and Drosophila cells (Rodriguez et al 2001;Smith et al 2004). …”

“…The nature of the proteins shown to be SUMOylated suggests that SUMOylation is predominantly a nuclear process (Hay 2005). Consistent with this, the SUMOylation machinery is localized mainly to the nucleus in both mammalian and fly cells (Rodriguez et al 2001;Smith et al 2004). Moreover, addition of both a consensus SUMO site and a NLS are necessary to confer SUMOylation upon an artificial substrate (Rodriguez et al 2001).…”

“…The fate of SUMOylated Med is currently unknown. However, as Ulp1, one of the major SUMO deconjugating enzymes in Drosophila, is localized to the nuclear pore complex (Smith et al 2004), it is likely that Med is deSUMOylated upon export. We suggest that ultimately SUMOylated Med is either recycled following deSUMOylation or degraded.…”

“…Moreover, addition of both a consensus SUMO site and a NLS are necessary to confer SUMOylation upon an artificial substrate (Rodriguez et al 2001). SUMOylation is a reversible modification, and a family of SUMO-specific proteases has been identified that catalyze deSUMOylation (Smith et al 2004;Hay 2007).…”

SUMOylation Pathway in Trypanosoma cruzi: Functional Characterization and Proteomic Analysis of Target Proteins

“…Diverse aminoacyl-tRNA synthetases were detected in our list of potential SUMO targets. In Drosophila glutamyl-prolyl-tRNA synthetase and methionyl-tRNA synthetase were demonstrated to be conjugated with SUMO in vivo, and a potential role for this modification in increasing the traffic of tRNAs from the nucleus to the ribosomes was suggested (64).…”