Abstract #50

Outer nuclear membrane targeting and partner-binding domains of Nvj1p reveal a role in sterol-dependent protein trafficking. David S. Goldfarb, Erik Kvam. Department of Biology, University of Rochester, Rochester, NY 14627.
   Nvj1p is an outer nuclear membrane (ONM) protein that interacts with the vacuole membrane protein Vac8p to form nucleus-vacuole (NV) junctions. NV junctions mediate piecemeal microautophagy of the nucleus (PMN), during which portions of the nucleus are pinched-off into the vacuole lumen and degraded. The up-regulation of Nvj1p under starvation conditions results in the expansion of NV junctions and the sequestration of two additional binding partners, Tsc13p and Osh1p. Tsc13p is an integral membrane protein required for very-long-chain fatty acid biosynthesis. Osh1p is implicated in sterol homeostasis by homology to mammalian oxysterol-binding protein. We have mapped the domains of Nvj1p responsible for ONM targeting and partner binding. ONM targeting requires both an N-terminal signal anchor-like sequence and the membrane-spanning domain of Nvj1p. In addition to linking the vacuole membrane to the ONM, Nvj1p physically connects the outer and inner nuclear membranes by bridging the perinuclear lumen. A region encompassing the membrane-spanning domain is sufficient to bind Tsc13p. Osh1p and Vac8p bind to distinct regions in the cytoplasmic tail of Nvj1p. Over-expression of Nvj1p in trp1 cells caused a growth defect in low tryptophan that is likely related to the sterol-dependent sorting of the tryptophan permease, Tat2p, to the plasma membrane. Deletion of the Osh1p-binding domain of Nvj1p abrogates this tryptophan transport-related growth phenotype. Moreover, nvj1-D trp1 cells grow faster than NVJ1 trp1 cells in limiting tryptophan. These results suggest that Osh1p sequestration by Nvj1p regulates protein trafficking to the plasma membrane in response to nutrient depletion.

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