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Vol. 19, Issue 12, 5559-5578, December 2008

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The Yeast LATS/Ndr Kinase Cbk1 Regulates Growth via Golgi-dependent Glycosylation and Secretion

Cornelia Kurischko^*, Venkata K. Kuravi^*, Nattha Wannissorn^*, Pavel A. Nazarov^*, Michelle Husain, Chao Zhang, Kevan M. Shokat, J. Michael McCaffery, and Francis C. Luca^*

*Department of Animal Biology and the Mari Lowe Center for Comparative Oncology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA 19104; Integrated Imaging Center, Johns Hopkins University, Baltimore, MD 21218; and Department of Cellular and Molecular Pharmacology and Howard Hughes Medical Institute, University of California, San Francisco, CA 94143

Submitted May 5, 2008; Revised September 19, 2008; Accepted September 29, 2008
Monitoring Editor: David G. Drubin

Saccharomyces cerevisiae Cbk1 is a LATS/Ndr protein kinase and a downstream component of the regulation of Ace2 and morphogenesis (RAM) signaling network. Cbk1 and the RAM network are required for cellular morphogenesis, cell separation, and maintenance of cell integrity. Here, we examine the phenotypes of conditional cbk1 mutants to determine the essential function of Cbk1. Cbk1 inhibition severely disrupts growth and protein secretion, and triggers the Swe1-dependent morphogenesis checkpoint. Cbk1 inhibition also delays the polarity establishment of the exocytosis regulators Rab-GTPase Sec4 and its exchange factor Sec2, but it does not interfere with actin polarity establishment. Cbk1 binds to and phosphorylates Sec2, suggesting that it regulates Sec4-dependent exocytosis. Intriguingly, Cbk1 inhibition causes a >30% decrease in post-Golgi vesicle accumulation in late secretion mutants, indicating that Cbk1 also functions upstream of Sec2-Sec4, perhaps at the level of the Golgi. In agreement, conditional cbk1 mutants mislocalize the cis-Golgi mannosyltransferase Och1, are hypersensitive to the aminoglycoside hygromycin B, and exhibit diminished invertase and Sim1 glycosylation. Significantly, the conditional lethality and hygromycin B sensitivity of cbk1 mutants are suppressed by moderate overexpression of several Golgi mannosyltransferases. These data suggest that an important function for Cbk1 and the RAM signaling network is to regulate growth and secretion via Golgi and Sec2/Sec4-dependent processes.

Address correspondence to: Francis C. Luca (fluca{at}vet.upenn.edu).

Abbreviations used: COG, conserved oligomeric Golgi complex; EM, electron microscopy; ER, endoplasmic reticulum; GEF, guanine nucleotide exchange factor; RAM, regulation of Ace2 and morphogenesis; SNARE, soluble N-ethylmaleimide-sensitive factor attachment protein receptor.