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R-SNARE YKT6 Resides in Membrane-Associated Protease-Resistant Protein Particles and Modulates Cell Cycle Progression When Over-Expressed.
MedLine Citation:
PMID:  22443861     Owner:  NLM     Status:  Publisher    
Abstract/OtherAbstract:
BACKGROUND INFORMATION: The arginine-type soluble N-ethylmaleimide-sensitive factor attachment protein receptor (R-SNARE) ykt6 possesses several atypical properties including selective high expression in neurons and neuroendocrine cells, a lipidated C-terminus rather than a transmembrane anchor, localization to punctae that do not correspond with known endomembrane markers, a potent ability to protect the secretory pathway from insults such as alpha-synuclein over-expression, and a propensity for specific upregulation in tumors. We have followed up on several of these features that together suggest nontraditional SNARE structures and functions. RESULTS: A significant portion of ykt6 in PC12 cells was found in a protease-resistant state suggestive of a large complex or aggregate. Other ER/Golgi SNAREs were not protease resistant, demonstrating that SNARE complexes per se did not cause protease resistance. Mutagenesis indicated that lipidation of the ykt6 C-terminus was also not involved, implicating its longin domain in particle formation. Gel filtration demonstrated that a smaller fraction of particulate ykt6 participated in SNARE complexes than other ER/Golgi SNAREs, suggesting that protease resistant ykt6 may not be available for SNARE interactions. Previously observed ykt6 fluorescent punctae were validated as bona fide particulate structures. Immunogold electron microscopy revealed ykt6 labeling of ~100 nm electron densities associated with diverse membranes. Density gradient analysis of the protease resistant structures confirmed their tight association with membranes. Since excess ykt6 has been correlated with tumorigenesis, we tested whether ykt6 over-expression in NRK cells that normally express little ykt6 affected the cell cycle. Ykt6 over-expression was found to result in altered cell division cycles as evidenced by significantly smaller cells, a higher mitotic index, and increased DNA synthesis. Mutagenesis studies dis-correlated SNARE function with the cell cycle effects; instead, the cell cycle effects correlated better with ykt6 properties related to the longin domain or particle formation. CONCLUSIONS: The ykt6 particles/aggregates may represent ykt6 engaged in a non-SNARE function(s) or else nonfunctional, stored, and/or excess ykt6. Whether the particulate ykt6 structures represents a means of buffering the apparent proliferative activity or are in fact mechanistically related to this activity will be of future interest in neuroscience and cancer biology.
Authors:
Nandhakumar Thayanidhi; Yingjian Liang; Haruki Hasegawa; Deborah C Nycz; Viola Oorschot; Judith Klumperman; Jesse C Hay
Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2012-3-22
Journal Detail:
Title:  Biology of the cell / under the auspices of the European Cell Biology Organization     Volume:  -     ISSN:  1768-322X     ISO Abbreviation:  -     Publication Date:  2012 Mar 
Date Detail:
Created Date:  2012-3-26     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  8108529     Medline TA:  Biol Cell     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
Copyright Information:
Copyright © 2012 Soçiété Francaise des Microscopies and Société de Biologie Cellulaire de France.
Affiliation:
From the Division of Biological Sciences and Center for Structural & Functional Neuroscience The University of Montana, Missoula Montana.
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