Role of Doa1 in the <i>Saccharomyces cerevisiae</i> DNA Damage Response

Lis, Ewa; Romesberg, Floyd E.

doi:10.1128/mcb.01640-05

Cited by 35 publications

(36 citation statements)

References 52 publications

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“…While more detailed structural insights into the interaction between Cdc48 and Ufd3 have to await a cocrystal structure analysis of both full-length proteins, the phenotypic analysis by Qiu et al is difficult to reconcile, as the well-characterized canavanine hypersensitivity of ufd3 mutants (Fig. 6a) (39,44) was not observed in their experimental system (50). The authors also failed to demonstrate mutual loss of binding for the ufd3 and cdc48 mutants used, leaving the possibility that residual interactions between Cdc48 and Ufd3 caused the lack of some phenotypes.…”

Section: Discussionmentioning

confidence: 99%

“…The basis underlying ubiquitin depletion in ufd3 mutants is not precisely known but is likely to reflect the inability to recycle ubiquitin in the context of proteasomal protein degradation (47,56). ⌬ufd3 strains exhibit a number of stress phenotypes and negative genetic interactions, many of which are an indirect consequence of ubiquitin depletion (32,39,44,47,56). However, defects of ⌬ufd3 in the monoubiquitylation of histone H2B (39), in the sorting of ubiquitylated membrane proteins into multivesicular bodies for vacuolar degradation (54), and in ribophagy (48) were shown to persist upon ubiquitin overexpression, suggesting that Ufd3 is directly involved in these processes.…”

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confidence: 99%

“…The absence of any general growth defects in the cdc48 shuffle strains enabled us to specifically analyze the effects of mutating the C-terminal tail of Cdc48 on known activities of Ufd2 and Ufd3 in vivo. ufd3 mutants are sensitive to a number of stress conditions, mostly as an indirect consequence of reduced ubiquitin pools in these strains (32,39,44,47,56). We compared the sensitivities of our shuffle strains to the arginine analogue canavanine with that of a ⌬ufd3 strain (Fig.…”

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confidence: 99%

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Cellular Functions of Ufd2 and Ufd3 in Proteasomal Protein Degradation Depend on Cdc48 Binding

Böhm

Lamberti

Fernández‐Sáiz

et al. 2011

Molecular and Cellular Biology

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The chaperone-related AAA ATPase Cdc48 (p97/VCP in higher eukaryotes) segregates ubiquitylated proteins for subsequent degradation by the 26S proteasome or for nonproteolytic fates. The specific outcome of Cdc48 activity is controlled by the evolutionary conserved cofactors Ufd2 and Ufd3, which antagonistically regulate the substrates' ubiquitylation states. In contrast to the interaction of Ufd3 and Cdc48, the interaction between the ubiquitin chain elongating enzyme Ufd2 and Cdc48 has not been precisely mapped. Consequently, it is still unknown whether physiological functions of Ufd2 in fact require Cdc48 binding. Here, we show that Ufd2 binds to the C-terminal tail of Cdc48, unlike the human Ufd2 homologue E4B, which interacts with the N domain of p97. The binding sites for Ufd2 and Ufd3 on Cdc48 overlap and depend critically on the conserved residue Y834 but are not identical. Saccharomyces cerevisiae cdc48 mutants altered in residue Y834 or lacking the C-terminal tail are viable and exhibit normal growth. Importantly, however, loss of Ufd2 and Ufd3 binding in these mutants phenocopies defects of ⌬ufd2 and ⌬ufd3 mutants in the ubiquitin fusion degradation (UFD) and Ole1 fatty acid desaturase activation (OLE) pathways. These results indicate that key cellular functions of Ufd2 and Ufd3 in proteasomal protein degradation require their interaction with Cdc48.

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Section: Discussionmentioning

confidence: 99%

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Cellular Functions of Ufd2 and Ufd3 in Proteasomal Protein Degradation Depend on Cdc48 Binding

Böhm

Lamberti

Fernández‐Sáiz

et al. 2011

Molecular and Cellular Biology

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“…The direct interaction between DOA1 and Ub was suggested by recent studies (7,9). Moreover, DOA1 also plays roles in the monoubiquitination of histone H2B and proliferating cell nuclear antigen (10) and in sorting ubiquitinated membrane proteins into multivesicular bodies (11).…”

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confidence: 86%

Structural Basis for Ubiquitin Recognition by a Novel Domain from Human Phospholipase A2-activating Protein

Zhou

Gao

et al. 2009

Journal of Biological Chemistry

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Ubiquitin (Ub) is an essential modifier conserved in all eukaryotes from yeast to human. Phospholipase A 2 -activating protein (PLAA), a mammalian homolog of yeast DOA1/UFD3, has been proposed to be able to bind with Ub, which plays important roles in endoplasmic reticulum-associated degradation, vesicle formation, and DNA damage response. We have identified a core domain from the PLAA family ubiquitin-binding region of human PLAA (residues 386 -465, namely PFUC) that can bind Ub and elucidated its solution structure and Ubbinding mode by NMR approaches. The PFUC domain possesses equal population of two conformers in solution by cis/ trans-isomerization, whereas the two isomers exhibit almost equivalent Ub binding abilities. This domain structure takes a novel fold consisting of four ␤-strands and two ␣-helices, and the Ub-binding site on PFUC locates in the surface of ␣2-helix, which is to some extent analogous to those of UBA, CUE, and UIM domains. This study provides structural basis and biochemical information for Ub recognition of the novel PFU domain from a PLAA family protein that may connect ubiquitination and degradation in endoplasmic reticulum-associated degradation.The eukaryotic secreted proteins are translocated into the endoplasmic reticulum after synthesis in cytosol. Misfolded or abnormally assembled proteins should be targeted for degradation through the endoplasmic reticulum-associated degradation (ERAD) 3 pathway (1, 2). This pathway involves many molecular steps: unfolded protein response in the endoplasmic reticulum lumen, retrotranslocation back into the cytosol, ubiquitin (Ub) conjugation, delivery of ubiquitinated proteins to proteasome, and degradation of the substrates by proteases (3, 4).A yeast protein DOA1/UFD3 has been shown to bind to CDC48 by both indirect and direct ways (5-7), suggesting that DOA1 may be involved in ERAD. Evidence indicates that DOA1 directly competes with UFD2 at the same docking site on CDC48, which determines whether a substrate is multiubiquitinated and routed to the proteasome for degradation or deubiquitinated and released for other purposes (8). The direct interaction between DOA1 and Ub was suggested by recent studies (7, 9). Moreover, DOA1 also plays roles in the monoubiquitination of histone H2B and proliferating cell nuclear antigen (10) and in sorting ubiquitinated membrane proteins into multivesicular bodies (11).The mammalian homolog of DOA1 is called phospholipase A 2 -activating protein (PLAA), which can bind to P97/VCP (a CDC48 homolog) with its C-terminal domain PUL (7). Having high sequence similarity (31% identity) with DOA1, PLAA is proposed to possess similar function of DOA1. Like DOA1, PLAA has an N-terminal WD40 domain with yet unknown function. The central region of PLAA contains a putative PLAA family ubiquitin-binding (PFU) domain, which is supposed to bind with Ub as observed in yeast DOA1 (7). Although the mechanism underlying the function of PLAA remains unclear, Ub binding of PLAA might be the central role that connects ubiquitin...

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“…Since overexpression of UBI4 (encoding Ub) suppresses the phenotypes of doa1Δ (Finley et al 1987;Lis and Romesberg 2006), we reasoned that it might suppress the lethality of doa1Δ strains overexpressing CSE4. Overexpression of UBI4 Figure 2 Role of Doa1 in proteolysis of Cse4 in a Psh1-independent manner.…”

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confidence: 99%

A Novel Role of the N Terminus of Budding Yeast Histone H3 Variant Cse4 in Ubiquitin-Mediated Proteolysis

Dawson

Rawson

et al. 2013

Genetics

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Regulating levels of centromeric histone H3 (CenH3) variant is crucial for genome stability. Interaction of Psh1, an E3 ligase, with the C terminus of Cse4 has been shown to contribute to its proteolysis. Here, we demonstrate a role for ubiquitination of the N terminus of Cse4 in regulating Cse4 proteolysis for faithful chromosome segregation and a role for Doa1 in ubiquitination of Cse4.C ENTROMERIC histone H3 (CenH3), an evolutionarily conserved histone H3 variant, is essential for chromosome segregation (Stoler et al. 1995;Meluh et al. 1998;Blower and Karpen 2001;Maddox et al. 2012). Mis-localization and overexpression of CenH3 has been observed in cancers and associated with aneuploidy in Drosophila melanogaster (Tomonaga et al. 2003;Heun et al. 2006;Moreno-Moreno et al. 2006). Studies from budding yeast and fruit flies have shown that proteolysis of CenH3 plays an important role in preventing its mis-localization MorenoMoreno et al. 2006 MorenoMoreno et al. , 2011. Psh1, an E3 ligase, interacts with Cse4, the budding yeast CenH3, and mediates its ubiquitination for proteolysis (Hewawasam et al. 2010;Ranjitkar et al. 2010). Despite ubiquitination of Cse4 by Psh1, Cse4 is only partially stabilized when overexpressed in psh1Δ strains (Hewawasam et al. 2010;Ranjitkar et al. 2010), suggesting that additional factors regulate Cse4 protein stability.In this study, we investigated the role of Cse4 domains in directing Cse4 proteolysis and used a genome-wide screen to identify pathways that regulate Cse4 stability. Since subcellular levels of Cse4 are stringently regulated, strains overexpressing CSE4 or cse4 mutants have been used to identify proteolytic pathways and factors such as Psh1 that mediate its degradation (Hewawasam et al. 2010;Ranjitkar et al. 2010). We initiated our studies with a mutant cse416KR (16KR) and 16KR fusion mutants in which lysines (K) are mutated to arginines (R) (Figure 1, A-C; Supporting Information, Table S1 and File S1). Overexpression of 16KR leads to defects in chromosome segregation, and this correlates with enrichment in chromatin and increased protein stability Au et al. 2008) . Our results show that the stability of CK (fusion of Cse4 N terminus amino acids 1-135 and 16KR C terminus) is low (t 1/2 = 26 6 6 min) and is comparable to that of Cse4 (t 1/2 = 34 6 3 min), indicating that the lack of ubiquitination of the C terminus due to K-to-R mutations does not increase protein stability. In contrast, the stability of KC (a fusion of 16KR N terminus and Cse4 C terminus) is high (t 1/2 . 120 min) and similar to 16KR (Figure 1, B and C), showing that K-to-R mutations in the N terminus of Cse4 increase protein stability. Consistent with these results are previous observations showing that Cse4 lacking the N terminus (Δ129) is fivefold more stable than full-length Cse4 (Morey et al. 2004). Increased Cse4 stability correlates with chromosome loss, as KC and 16KR strains exhibit fourfold higher chromosome loss than strains expressing CK or wild-type Cse4 ( Figure 1G). Based on ...

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Role of Doa1 in the Saccharomyces cerevisiae DNA Damage Response

Cited by 35 publications

References 52 publications

Cellular Functions of Ufd2 and Ufd3 in Proteasomal Protein Degradation Depend on Cdc48 Binding

Cellular Functions of Ufd2 and Ufd3 in Proteasomal Protein Degradation Depend on Cdc48 Binding

Structural Basis for Ubiquitin Recognition by a Novel Domain from Human Phospholipase A2-activating Protein

A Novel Role of the N Terminus of Budding Yeast Histone H3 Variant Cse4 in Ubiquitin-Mediated Proteolysis

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