R-SNARE ykt6 resides in membrane-associated protease-resistant protein particles and modulates cell cycle progression when over-expressed.

Authors:
Nandhakumar Thayanidhi
Nandhakumar Thayanidhi
University of Montana
United States
Yingjian Liang
Yingjian Liang
The First Affiliated Hospital of Harbin Medical University
China
Haruki Hasegawa, PhD
Haruki Hasegawa, PhD
Amgen Inc.
Principal Scientist
Protein trafficking Protein biosynthesis
South San Francisco, CA | United States
Viola Oorschot
Viola Oorschot
Monash University
Australia
Judith Klumperman
Judith Klumperman
Center for Molecular Medicine
Athens | United States
Jesse C Hay
Jesse C Hay
The University of Montana
Missoula | United States

Biol Cell 2012 Jul 29;104(7):397-417. Epub 2012 May 29.

Division of Biological Sciences and Centre for Structural and Functional Neuroscience, The University of Montana, Missoula, MT 59812, USA.

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, a lipidated C-terminus, localization to punctae that do not correspond with known endomembrane markers, a potent ability to protect the secretory pathway from alpha-synuclein over-expression and specific up-regulation 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 endoplasmic reticulum/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. 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 normal rat kidney 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 represent 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.

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http://dx.doi.org/10.1111/boc.201100048DOI Listing

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July 2012
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