Dataset Information

Ubiquitin-specific protease 4 inhibits mono-ubiquitination of the master growth factor signaling kinase PDK1.

ABSTRACT:

Background

Phosphorylation by the phospho-inositide-dependent kinase 1 (PDK1) is essential for many growth factor-activated kinases and thus plays a critical role in various processes such as cell proliferation and metabolism. However, the mechanisms that control PDK1 have not been fully explored and this is of great importance as interfering with PDK1 signaling may be useful to treat diseases, including cancer and diabetes.

Methodology/principal findings

In human cells, few mono-ubiquitinated proteins have been described but in all cases this post-translational modification has a key regulatory function. Unexpectedly, we find that PDK1 is mono-ubiquitinated in a variety of human cell lines, indicating that PDK1 ubiquitination is a common and regulated process. Ubiquitination occurs in the kinase domain of PDK1 yet is independent of its kinase activity. By screening a library of ubiquitin proteases, we further identify the Ubiquitin-Specific Protease 4 (USP4) as an enzyme that removes ubiquitin from PDK1 in vivo and in vitro and co-localizes with PDK1 at the plasma membrane when the two proteins are overexpressed, indicating direct deubiquitination.

Conclusions

The regulated mono-ubiquitination of PDK1 provides an unanticipated layer of complexity in this central signaling network and offers potential novel avenues for drug discovery.

SUBMITTER: Uras IZ

PROVIDER: S-EPMC3274522 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Publications

Ubiquitin-specific protease 4 inhibits mono-ubiquitination of the master growth factor signaling kinase PDK1.

Uras Iris Z IZ List Thomas T Nijman Sebastian M B SM

PloS one 20120207 2

<h4>Background</h4>Phosphorylation by the phospho-inositide-dependent kinase 1 (PDK1) is essential for many growth factor-activated kinases and thus plays a critical role in various processes such as cell proliferation and metabolism. However, the mechanisms that control PDK1 have not been fully explored and this is of great importance as interfering with PDK1 signaling may be useful to treat diseases, including cancer and diabetes.<h4>Methodology/principal findings</h4>In human cells, few mono- ...[more]

PMID: 22347420

Similar Datasets

Project description:Background3-phosphoinositide-dependent protein kinase-1 (PDK1) acts as a master kinase of protein kinase A, G, and C family (AGC) kinase to predominantly govern cell survival, proliferation, and metabolic homeostasis. Although the regulations to PDK1 downstream substrates such as protein kinase B (AKT) and ribosomal protein S6 kinase beta (S6K) have been well established, the upstream regulators of PDK1, especially its degrader, has not been defined yet.MethodA clustered regularly interspaced short palindromic repeats (CRISPR)-based E3 ligase screening approach was employed to identify the E3 ubiquitin ligase for degrading PDK1. Western blotting, immunoprecipitation assays and immunofluorescence (IF) staining were performed to detect the interaction or location of PDK1 with speckle-type POZ protein (SPOP). Immunohistochemistry (IHC) staining was used to study the expression of PDK1 and SPOP in prostate cancer tissues. In vivo and in vitro ubiquitination assays were performed to measure the ubiquitination conjugation of PDK1 by SPOP. In vitro kinase assays and mass spectrometry approach were carried out to identify casein kinase 1 (CK1) and glycogen synthase kinase 3 (GSK3)-mediated PDK1 phosphorylation. The biological effects of PDK1 mutations and correlation with SPOP mutations were performed with colony formation, soft agar assays and in vivo xenograft mouse models.ResultsWe identified that PDK1 underwent SPOP-mediated ubiquitination and subsequent proteasome-dependent degradation. Specifically, SPOP directly bound PDK1 by the consensus degron in a CK1/GSK3β-mediated phosphorylation dependent manner. Pathologically, prostate cancer patients associated mutations of SPOP impaired PDK1 degradation and thus activated the AKT kinase, resulting in tumor malignancies. Meanwhile, mutations that occurred around or within the PDK1 degron, by either blocking SPOP to bind the degron or inhibiting CK1 or GSK3β-mediated PDK1 phosphorylation, could markedly evade SPOP-mediated PDK1 degradation, and played potently oncogenic roles via activating the AKT kinase.ConclusionsOur results not only reveal a physiological regulation of PDK1 by E3 ligase SPOP, but also highlight the oncogenic roles of loss-of-function mutations of SPOP or gain-of-function mutations of PDK1 in tumorigenesis through activating the AKT kinase.

Dataset Information

Ubiquitin-specific protease 4 inhibits mono-ubiquitination of the master growth factor signaling kinase PDK1.

Background

Methodology/principal findings

Conclusions

Publications

Ubiquitin-specific protease 4 inhibits mono-ubiquitination of the master growth factor signaling kinase PDK1.

Similar Datasets

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