Human Vam6p promotes lysosome clustering and fusion in vivo.
Regulated fusion of mammalian lysosomes is critical to their ability to acquire both internalized and biosynthetic materials. Here, we report the identification of a novel human protein, hVam6p, that promotes lysosome clustering and fusion in vivo. Although hVam6p exhibits homology to the Saccharomyces cerevisiae vacuolar protein sorting gene product Vam6p/Vps39p, the presence of a citron homology (CNH) domain at the NH(2) terminus is unique to the human protein. Overexpression of hVam6p results in massive clustering and fusion of lysosomes and late endosomes into large (2-3 microm) juxtanuclear structures. This effect is reminiscent of that caused by expression of a constitutively activated Rab7. However, hVam6p exerts its effect even in the presence of a dominant-negative Rab7, suggesting that it functions either downstream of, or in parallel to, Rab7. Data from gradient fractionation, two-hybrid, and coimmunoprecipitation analyses suggest that hVam6p is a homooligomer, and that its self-assembly is mediated by a clathrin heavy chain repeat domain in the middle of the protein. Both the CNH and clathrin heavy chain repeat domains are required for induction of lysosome clustering and fusion. This study implicates hVam6p as a mammalian tethering/docking factor characterized with intrinsic ability to promote lysosome fusion in vivo.
Caplan
S
S
Cell Biology and Metabolism Branch at the National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA.
Hartnell
L M
LM
Aguilar
R C
RC
Naslavsky
N
N
Bonifacino
J S
JS
D033942:Adaptor Proteins, Vesicular Transport; D000595:Amino Acid Sequence; D000818:Animals; D000071183:Autophagy-Related Proteins; D019556:COS Cells; D003001:Cloning, Molecular; D011992:Endosomes / Q000378:metabolism; D005799:Genes, Dominant; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008247:Lysosomes / Q000378:metabolism* / Q000648:ultrastructure; D008565:Membrane Proteins / Q000502:physiology*; D051379:Mice; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D008957:Models, Genetic; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D014018:Tissue Distribution; D020798:Two-Hybrid System Techniques; D033921:Vesicular Transport Proteins; D020691:rab GTP-Binding Proteins / Q000378:metabolism; D000090802:rab7 GTP-Binding Proteins
D033942:Adaptor Proteins, Vesicular Transport; D000071183:Autophagy-Related Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D029701:Saccharomyces cerevisiae Proteins; C406817:VAM6 protein, S cerevisiae; C485791:VPS39 protein, human; D033921:Vesicular Transport Proteins; D000090802:rab7 GTP-Binding Proteins; C000716463:rab7 GTP-binding proteins, human; C000716465:rab7 GTP-binding proteins, mouse; D020691:rab GTP-Binding Proteins