The JAMM motif of human deubiquitinase Poh1 is essential for cell viability

Millennium Pharmaceuticals, Inc., 40 Landsdowne Street, Cambridge, MA 02139, USA.
Molecular Cancer Therapeutics (Impact Factor: 5.68). 02/2007; 6(1):262-8. DOI: 10.1158/1535-7163.MCT-06-0542
Source: PubMed

ABSTRACT Poh1 deubiquitinase activity is required for proteolytic processing of polyubiquitinated substrates by the 26S proteasome, linking deubiquitination to complete substrate degradation. Poh1 RNA interference (RNAi) in HeLa cells resulted in a reduction in cell viability and an increase in polyubiquitinated protein levels, supporting the link between Poh1 and the ubiquitin proteasome pathway. To more specifically test for any requirement of the zinc metalloproteinase motif of Poh1 to support cell viability and proteasome function, we developed a RNAi complementation strategy. Effects on cell viability and proteasome activity were assessed in cells with RNAi of endogenous Poh1 and induced expression of wild-type Poh1 or a mutant form of Poh1, in which two conserved histidines of the proposed catalytic site were replaced with alanines. We show that an intact zinc metalloproteinase motif is essential for cell viability and 26S proteasome function. As a required enzymatic component of the proteasome, Poh1 is an intriguing therapeutic drug target for cancer.

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    • "In particular, targeting proteasome-associated deubiquitination has the potential of increasing the levels of ubiquitin conjugated proteins, hence triggering proteotoxic stress and apoptosis similar to that observed with inhibitors of proteolytic activity. The metalloprotease POH1 is an obvious target since it is absolutely required for cell viability and is overexpressed in a variety of tumours (Gallery et al., 2007). "
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    • "PSMD14 is also expressed in many tissues and functions as a component of the 26S proteasome, which plays a role in protein degradation via deubiquitination [Verma et al., 2002; Yao and Cohen, 2002]. Knock-down of PSMD14 expression in human cell lines 842 AMERICAN JOURNAL OF MEDICAL GENETICS PART A revealed decreased cell viability [Gallery et al., 2007] and in carcinoma cells it leads to cell cycle arrest [Byrne et al., 2010]. Furthermore, knock-down of this gene in postmitotic neurons resulted in apoptosis [Staropoli and Abeliovich, 2005]. "
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    • "Indeed, a single point mutation on the conserved zinc-coordinating histidine 113 abolished DUB activity. These results are in agreement with mutational studies on Psmd14 function in human protein orthologues (Gallery et al., 2007). In contrast, we found cysteine 120 to be dispensable for Psmd14 function in ES cells, consistent with the fact that it belongs to a family distinct from the cysteine proteases. "
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