A Degenerate Cohort of Yeast Membrane Trafficking DUBs Mediates Cell Polarity and Survival*

Janelr Beckley,Jun-Song Chen,Yanling Yang,Junmin Peng,Kathleenl Gould

doi:10.1074/mcp.m115.050039

Janelr Beckley, Jun-Song Chen + Show 3 more

Open Access

https://doi.org/10.1074/mcp.m115.050039

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Abstract

Deubiquitinating enzymes (DUBs), cysteine or metallo- proteases that cleave ubiquitin chains or protein conjugates, are present in nearly every cellular compartment, with overlapping protein domain structure, localization, and functions. We discovered a cohort of DUBs that are involved in membrane trafficking (ubp4, ubp5, ubp9, ubp15, and sst2) and found that loss of all five of these DUBs but not loss of any combination of four, significantly impacted cell viability in the fission yeast Schizosaccharomyces pombe (1). Here, we delineate the collective and individual functions and activities of these five conserved DUBs using comparative proteomics, biochemistry, and microscopy. We find these five DUBs are degenerate rather than redundant at the levels of cell morphology, substrate selectivity, ubiquitin chain specificity, and cell viability under stress. These studies reveal the complexity of interplay among these enzymes, providing a foundation for understanding DUB biology and providing another example of how cells utilize degeneracy to improve survival.

Highlights

Eukaryotic cells integrate signaling pathways to modulate their response to environmental changes, predominately through dynamic protein posttranslational modifications like ubiquitination (Ub’n) [2, 3]
Images were processed in Image J (Fiji) [25], vacuolar volume was calculated using the 3D object counter plugin [26], and final figures were assembled in Adobe Creative Suite (CS6). These five membrane trafficking Deubiquitinating enzymes (DUBs)—Ubp4, Ubp5, Ubp9, Ubp15, and Sst2—span two of the four Ub protease domain families found in S. pombe—Ub-specific protease (USP: Ubp4, Ubp5, Ubp9, and Ubp15) and Jab1/MPN domain-associated metallo-isopeptidase (JAMM: Sst2) (Fig. 1A)
Loss of cell polarity is detected by light microscopy of live cells [33], and the ratio of cell length divided by cell width at septation is a simple way to quantitate changes in cell polarity and morphology

Summary

Introduction

Eukaryotic cells integrate signaling pathways to modulate their response to environmental changes, predominately through dynamic protein posttranslational modifications like ubiquitination (Ub’n) [2, 3]. Proteins missing from WT but present in the DUB delete were included as putative substrates (indicated by a ratio of “ND” in Supplemental Table S6).

Results

Conclusion