Evidence that mating by the Saccharomyces cerevisiae gpa1Val50 mutant occurs through the default mating pathway and a suggestion of a role for ubiquitin-mediated proteolysis.

Xu, Bing-e; Kurjan, Janet

doi:10.1091/mbc.8.9.1649

Cited by 12 publications

(22 citation statements)

References 51 publications

(84 reference statements)

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“…Early studies have shown that inhibition of the proteasome activity leads to stabilization of Rpn4 and subsequent upregulation of the proteasome genes, which may compensate the impairment imposed to the proteasome London et al 2004). Consistently, deletion of RPN4 or inactivation of Rpn4 exhibits a strong synthetic lethality with mutations of several proteasome subunits (Xu and Kurjan 1997;Fujimoro et al 1998;. Recently, Hanna et al (2007) suggested that depletion of ubiquitin (ubiquitin stress) may trigger an Rpn4-dependent upregulation of the proteasome genes, which, along with an increased expression of the polyubiquitin gene UBI4 and the gene encoding deubiquitylating enzyme Ubp6, enable the cell to restore a normal ubiquitin level.…”

Section: Discussionmentioning

confidence: 89%

Disruption of Rpn4-Induced Proteasome Expression in Saccharomyces cerevisiae Reduces Cell Viability Under Stressed Conditions

Wang

et al. 2008

Genetics

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The proteasome homeostasis in Saccharomyces cerevisiae is regulated by a negative feedback circuit in which the transcription activator Rpn4 upregulates the proteasome genes and is rapidly degraded by the assembled proteasome. Previous studies have shown that rpn4D cells are sensitive to a variety of stresses. However, the contribution of the loss of Rpn4-induced proteasome expression to the rpn4D phenotypes remains unclear because Rpn4 controls numerous genes other than the proteasome genes. Here we construct a yeast strain in which one of the essential proteasome genes, PRE1, is no longer induced by Rpn4. We show that the active proteasome level is lower in this strain than in the wild-type counterpart. Moreover, we demonstrate that loss of Rpn4-induced proteasome expression leads to cell-cycle delay in G 2 /M and sensitizes cells to various stresses. To our knowledge, this is the first report that explicitly reveals the physiological function of Rpn4-induced proteasome expression. This study also provides a tool for understanding the interactions between proteasome homeostasis and other cellular processes.

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Section: Discussionmentioning

confidence: 89%

Disruption of Rpn4-Induced Proteasome Expression in Saccharomyces cerevisiae Reduces Cell Viability Under Stressed Conditions

Wang

et al. 2008

Genetics

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“…We have proposed that a protein turnover mechanism leads to low levels of the G␣ proteins in the ⌬gnb-1 and ⌬gng-1 backgrounds, as (i) mRNA levels for the three genes are normal in ⌬gnb-1 mutants (34, 60), (ii) proteolysis of G␣ proteins is a regulatory mechanism observed in other fungi (8,28,55,58), and (iii) results from preliminary experiments analyzing chemicals that influence the proteasomal or vacuolar protein turnover pathways are consistent with proteolysis of at least one G␣ protein (GNA-1 [data not shown]). Importantly, we were able to restore levels of the three G␣ proteins through overexpression of the activated alleles.…”

Section: Figmentioning

confidence: 99%

Genetic and Physical Interactions between Gα Subunits and Components of the Gβγ Dimer of Heterotrimeric G Proteins in Neurospora crassa

et al. 2012

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Heterotrimeric G proteins are critical regulators of growth and asexual and sexual development in the filamentous fungus Neurospora crassa. Three G␣ subunits (GNA-1, GNA-2, and GNA-3), one G␤ subunit (GNB-1), and one G␥ subunit (GNG-1) have been functionally characterized, but genetic epistasis relationships between G␤ and G␣ subunit genes have not been determined. Physical association between GNB-1 and FLAG-tagged GNG-1 has been previously demonstrated by coimmunoprecipitation, but knowledge of the G␣ binding partners for the G␤␥ dimer is currently lacking. In this study, the three N. crassa G␣ subunits are analyzed for genetic epistasis with gnb-1 and for physical interaction with the G␤␥ dimer. We created double mutants lacking one G␣ gene and gnb-1 and introduced constitutively active, GTPase-deficient alleles for each G␣ gene into the ⌬gnb-1 background. Genetic analysis revealed that gna-3 is epistatic to gnb-1 with regard to negative control of submerged conidiation. gnb-1 is epistatic to gna-2 and gna-3 for aerial hyphal height, while gnb-1 appears to act upstream of gna-1 and gna-2 during aerial conidiation. None of the activated G␣ alleles restored female fertility to ⌬gnb-1 mutants, and the gna-3 Q208L allele inhibited formation of female reproductive structures, consistent with a need for G␣ proteins to cycle through the inactive GDP-bound form for these processes. Coimmunoprecipitation experiments using extracts from the gng-1-FLAG strain demonstrated that the three G␣ proteins interact with the G␤␥ dimer. The finding that the G␤␥ dimer interacts with all three G␣ proteins is supported by epistasis between gnb-1 and gna-1, gna-2, and gna-3 for at least one function.

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“…Given these results, we expressed either a C-terminally tagged RPN4-flag or an N-terminally tagged ha-RPN4 from the P CUP1 promoter and determined the levels of RPN4 protein at different concentrations of the promoter-inducing CuSO 4 . Neither of these tagged derivatives of RPN4 could be detected by immunoblotting even on the induction of P CUP1 (Fig.…”

Section: Rpn4 Is a Short-lived Protein Degraded By The 26s Proteasomementioning

confidence: 99%

“…The phenotypes of AVY302 and the previously described strain (36) were similar as well (data not shown). Two other studies reported that rpn2⌬ cells were inviable (4,56). The disruption of RPN2 in these works was carried out with either TRP1 or ADE2, by using the RPN2 BglII site at codon 38.…”

Section: Rpn4mentioning

confidence: 99%

“…More recent studies have shown that mutations in RPN4 inhibit the degradation of normally short-lived proteins that are targeted by the N-end rule pathway, by the ubiquitin͞fusion͞degradation (UFD) pathway, and apparently also by other pathways of the ubiquitin (Ub)-proteasome system (3,4). These findings suggested that the ability of rpn4 mutations to suppress the conditional lethality of sec63-101 may stem from stabilization of the mutant but partially active SEC63-101 against degradation at nonpermissive temperature.…”

mentioning

confidence: 99%

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RPN4 is a ligand, substrate, and transcriptional regulator of the 26S proteasome: A negative feedback circuit

Xie

Varshavsky

2001

Proc. Natl. Acad. Sci. U.S.A.

413

396

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The RPN4 (SON1, UFD5) protein of the yeast Saccharomyces cerevisiae is required for normal levels of intracellular proteolysis. RPN4 is a transcriptional activator of genes encoding proteasomal subunits. Here we show that RPN4 is required for normal levels of these subunits. Further, we demonstrate that RPN4 is extremely short-lived (t 1/2 Ϸ2 min), that it directly interacts with RPN2, a subunit of the 26S proteasome, and that rpn4⌬ cells are perturbed in their cell cycle. The degradation signal of RPN4 was mapped to its N-terminal region, outside the transcription-activation domains of RPN4. The ability of RPN4 to augment the synthesis of proteasomal subunits while being metabolically unstable yields a negative feedback circuit in which the same protein up-regulates the proteasome production and is destroyed by the assembled active proteasome.proteolysis ͉ ubiquitin ͉ N-end rule ͉ UFD pathway ͉ cell cycle T he Saccharomyces cerevisiae RPN4 gene (its earlier names are SON1 and UFD5) (1) was originally identified through mutant rpn4 alleles that suppressed the growth defect of sec63-101 cells, which bore a temperature-sensitive (ts) variant of SEC63, an essential component of the protein translocation channel in the endoplasmic reticulum membrane (2). More recent studies have shown that mutations in RPN4 inhibit the degradation of normally short-lived proteins that are targeted by the N-end rule pathway, by the ubiquitin͞fusion͞degradation (UFD) pathway, and apparently also by other pathways of the ubiquitin (Ub)-proteasome system (3, 4). These findings suggested that the ability of rpn4 mutations to suppress the conditional lethality of sec63-101 may stem from stabilization of the mutant but partially active SEC63-101 against degradation at nonpermissive temperature.Regulated proteolysis by the Ub͞proteasome system plays essential roles in the cell cycle, differentiation, stress responses, and many other processes (5-7). Ub is a 76-residue protein whose covalent conjugation to other proteins marks these proteins for degradation by the 26S proteasome, an ATP-dependent multisubunit protease. Ub conjugation involves the formation of a thioester between the C terminus of Ub and a specific cysteine of the Ub-activating (E1) enzyme. The Ub moiety of E1ϳUb thioester is transesterified to a cysteine in one of several Ubconjugating (E2) enzymes. The Ub moiety of E2ϳUb thioester is conjugated via the isopeptide bond to the -amino group of either a substrate's Lys residue or a Lys residue of another Ub moiety, the latter reaction resulting in a substrate-linked multi-Ub chain (7,8). Most E2 enzymes function in complexes with proteins called E3 (9-11). The functions of E3s include the initial recognition of degradation signals (degrons) in substrate proteins, with different E3s recognizing different classes of degrons (12)(13)(14). The E2-E3 complexes, referred to as Ub ligases (this term is also used to denote E3s alone), mediate the formation of substrate-linked multi-Ub chains (15, 16). Ubiquitylated substrates are proce...

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Evidence that mating by the Saccharomyces cerevisiae gpa1Val50 mutant occurs through the default mating pathway and a suggestion of a role for ubiquitin-mediated proteolysis.

Cited by 12 publications

References 51 publications

Disruption of Rpn4-Induced Proteasome Expression in Saccharomyces cerevisiae Reduces Cell Viability Under Stressed Conditions

Disruption of Rpn4-Induced Proteasome Expression in Saccharomyces cerevisiae Reduces Cell Viability Under Stressed Conditions

Genetic and Physical Interactions between Gα Subunits and Components of the Gβγ Dimer of Heterotrimeric G Proteins in Neurospora crassa

RPN4 is a ligand, substrate, and transcriptional regulator of the 26S proteasome: A negative feedback circuit

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