Ruk is ubiquitinated but not degraded by the proteasome

Verdier, Frédérique; Valovka, Taras; Zhyvoloup, Alexander; Дробот, Л. Б.; Buchman, Vladimir L.; Waterfield, Mike; Gout, Ivan

doi:10.1046/j.1432-1033.2002.03031.x

Cited by 30 publications

(34 citation statements)

References 37 publications

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“…3). While it has previously been demonstrated that the coiled-coil domain of SETA can mediate multimerization Verdier et al, 2002) this is the first instance to our knowledge, of a SETA-binding partner showing a preference for SETA dimers.…”

Section: Discussionmentioning

confidence: 62%

SETA/CIN85/Ruk and its binding partner AIP1 associate with diverse cytoskeletal elements, including FAKs, and modulate cell adhesion

Schmidt

Chen

Randazzo

et al. 2003

Journal of Cell Science

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The adaptor protein SETA/CIN85/Ruk is involved in regulating diverse signal transduction pathways, including the internalization of tyrosine kinase receptors via the Cbl ubiquitin ligases, and attenuating PI3K activity by interaction with its regulatory subunit. Here we present evidence for a new aspect of SETA function, based on the initial observation that it co-localizes with actin in microfilaments and at focal adhesions, and with microtubules. Although there was no evidence for direct molecular interactions between SETA and cytoskeletal proteins, the SETA-interacting protein AIP1, which is a rat ortholog of the Xenopus src substrate Xp95, strongly interacted with structural proteins of the cytoskeleton, including actin and tubulins. Both SETA and AIP1 interacted with focal adhesion kinase (FAK) and proline rich tyrosine kinase 2 (PYK-2), and c-Cbl interacted with PYK-2. AIP1, which interacted more strongly than either SETA or c-Cbl, required an intact consensus tyrosine kinase phosphorylation sequence at Y319 to bind to focal adhesion kinases, which suggests that phosphorylation is an important mediator of this complex. SETA, which interacted as a dimer with focal adhesion kinases, promoted the interaction between PYK-2 and AIP1. Direct analysis of the impact of these proteins on cell adhesion, by use of an electrical cell-substrate impedance sensor (ECIS), showed that SETA promoted cell adhesion while AIP1 and c-Cbl reduced it. Furthermore, the ability of AIP1 and AIP1 mutants to decrease cell adhesion in ECIS analysis correlated with their presence in PYK-2 complexes, providing a direct link between AIP1-mediated molecular interactions and cellular behavior. Transfection of AIP1 also reduced the level of phosphorylation of endogenous PYK-2 and FAK, suggesting that this protein may directly regulate focal adhesion kinases, and thereby cell adhesion. These data are the first to implicate the adaptor protein SETA and its binding partner AIP1 as being involved with the cytoskeleton and in the regulation of cell adhesion, and suggest that they may be part of the focal adhesion kinase regulatory complex.

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Section: Discussionmentioning

confidence: 62%

SETA/CIN85/Ruk and its binding partner AIP1 associate with diverse cytoskeletal elements, including FAKs, and modulate cell adhesion

Schmidt

Chen

Randazzo

et al. 2003

Journal of Cell Science

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“…However, neither IP of SETA and antiubiquitin Western blot, nor cotransfection of an HA-tagged polyubiquitin provided evidence that SETA itself is polyubiquitinated (data not shown). We did not investigate the monoubiquitination of SETA, because it has been established that the protein is modified in this way (32,37).…”

Section: Seta Can Modulate the Polyubiquitination Of Egfr By Cblsmentioning

confidence: 99%

Epidermal growth factor receptor signaling intensity determines intracellular protein interactions, ubiquitination, and internalization

Schmidt

Furnari²,

Cavenee

et al. 2003

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

142

116

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Ligand activation of the epidermal growth factor receptor (EGFR) causes the binding of Cbls, which leads to EGFR polyubiquitination and internalization through endophilin complexes that contain the adaptor protein SH3-domain encoding, expressed in tumorigenic astrocytes͞Cbl-interacting protein of 85 kDa͞regulator of ubiquitous kinase (SETA͞CIN85͞Ruk). In cells grown at high density, high levels of SETA interfered in the recruitment of Casitas B-lineage (Cbl) proteins to the EGFR and reduced its polyubiquitination, suggesting that SETA has a regulatory function in the formation of the EGFR-Cbl-endophilin complex and in EGFR down-regulation. In a situation where there is EGFR signaling but no internalization or down-regulation, as is the case with the EGFR with exons 2-7 deleted (⌬EGFR) oncogene, these proteins were absent altogether. By using mAb 806, which recognizes an EGFR-activation state and preferentially immunoprecipitates ⌬EGFR, we show that ⌬EGFR did not interact with Cbls, SETA, or endophilin A1, providing a mechanistic explanation for its lack of internalization. As would be expected by the absence of Cbl proteins in the ⌬EGFR complex, the mutant receptor was also not polyubiquitinated. The intracellular C terminus and tyrosine autophosphorylation pattern of ⌬EGFR are similar to wild-type EGFR, but it signals at a lower intensity as determined by levels of EGFR phosphotyrosine. To test the implication that the lack of interaction with the Cbl-SETA-endophilin complex is because of differences in signal intensity, EGFR-expressing cells were treated with tyrphostin AG1478 EGFR inhibitor. Attenuation of wild-type EGFR signal to levels similar to that found in ⌬EGFR resulted in the dissociation of SETA and Cbl proteins and a concomitant attenuation of receptor internalization.

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“…Of these proteins, epsin, Eps15, ␤-arrestin, and CIN85 are known to be ubiquitinated. Specifically, they are modified with monoubiquitin (4,7,(15)(16)(17)(18)(19).…”

mentioning

confidence: 99%

“…The Cbl ubiquitin ligase was recently found to recruit a complex of CIN85 with another endocytic protein, endophilin, to facilitate the internalization of activated, ubiquitinated growth factor receptors (5,23). Furthermore, CIN85 is monoubiquitinated at an unknown site(s) by both Nedd4 and Cbl (7,18).…”

mentioning

confidence: 99%

The Rsp5 Ubiquitin Ligase Binds to and Ubiquitinates Members of the Yeast CIN85-Endophilin Complex, Sla1-Rvs167

Stamenova¹,

Dunn²,

Adler

et al. 2004

Journal of Biological Chemistry

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Sla1 and Rvs167 are yeast proteins required for receptor internalization and organization of the actin cytoskeleton. Here we provide evidence that Sla1 and Rvs167 are orthologues of the mammalian CIN85 and endophilin proteins, respectively, which are required for ligand-stimulated growth factor receptor internalization. Sla1 is similar in domain structure to CIN85 and binds directly to the endophilin-like Rvs167. Akin to CIN85, Sla1 interacts with synaptojanins and a ubiquitin ligase that regulates endocytosis. This ubiquitin ligase, Rsp5, binds directly to both Sla1 and Rvs167. The interaction between Rsp5 and Rvs167 is mediated through Rsp5 WW domains and PXY motifs in the central Gly-Pro-Ala-rich domain of Rvs167. Rvs167 PXY motifs are required for Rsp5-dependent monoubiquitination of Rvs167 on Lys 481 in the Src homology 3 (SH3) domain. Mutation of Lys 481 3 Arg causes cells to grow slowly on medium containing 1 M NaCl, although this phenotype is not due to the defect in ubiquitination caused by the K481R mutation. We propose that Rsp5 interaction with Sla1-Rvs167 promotes Rvs167 ubiquitination and regulates activity of this protein complex. Rvs167 ubiquitination is not required for general function of Rvs167, but may control specific Rvs167 SH3 domain-protein interactions or negatively regulate SH3 domain activity.

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Ruk is ubiquitinated but not degraded by the proteasome

Cited by 30 publications

References 37 publications

SETA/CIN85/Ruk and its binding partner AIP1 associate with diverse cytoskeletal elements, including FAKs, and modulate cell adhesion

SETA/CIN85/Ruk and its binding partner AIP1 associate with diverse cytoskeletal elements, including FAKs, and modulate cell adhesion

Epidermal growth factor receptor signaling intensity determines intracellular protein interactions, ubiquitination, and internalization

The Rsp5 Ubiquitin Ligase Binds to and Ubiquitinates Members of the Yeast CIN85-Endophilin Complex, Sla1-Rvs167

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