The yeast dynamin-like protein Vps1:vps1 mutations perturb the internalization and the motility of endocytic vesicles and endosomes via disorganization of the actin cytoskeleton.
Mammalian dynamin is responsible for scission of endocytic vesicles from the plasma membrane. A previous study showed that Vps1, a yeast dynamin-like protein, plays an important role in pheromone receptor internalization (Yu and Cai, 2004; J. Cell Sci. 117, 3839-3853). However, the details of how Vps1 acts in various phases of endocytosis including early internalization of the endocytic vesicle are poorly understood. To investigate the potential roles of Vps1 in both endocytic vesicle formation/maturation on the plasma membrane and endocytic vesicle internalization, time-lapse fluorescent images of GFP-tagged endocytic markers in live cells were analyzed using a particle tracking software. The loss of Vps1 leads to a robust increase in the lifespan of newly forming cortical endocytic vesicles carrying Las17-GFP, Ede1-GFP, Sla1-GFP, and Abp1-GFP, indicating that Vps1 is required for the proper assembly and maturation of endocytic vesicles. Particle track analysis revealed that Abp1-GFP vesicles in vps1 null cells moved a relatively short distance away from the cell membrane due to their non-directional movement. Furthermore, we found that the GTPase and the GED domains of Vps1 are required for the proper endocytic function of Vps1. Our tracking analysis data also revealed that the post-internalized vesicle motility en route to the vacuole was decreased significantly, perhaps due to severe disruption of the actin cables in Vps1 mutant cells.
Nannapaneni
Srikant
S
Department of Biology, Missouri State University, 901 South National Ave., Springfield, MO 65897, USA.
Wang
Daobing
D
Jain
Sandhya
S
Schroeder
Blake
B
Highfill
Chad
C
Reustle
Lindsay
L
Pittsley
Delilah
D
Maysent
Adam
A
Moulder
Shawn
S
McDowell
Ryan
R
Kim
Kyoungtae
K
D000199:Actins / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism; D004705:Endocytosis / Q000235:genetics / Q000502:physiology*; D011992:Endosomes / Q000378:metabolism*; D020558:GTP Phosphohydrolases / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D005838:Genotype; D007700:Kinetics; D008856:Microscopy, Fluorescence; D009154:Mutation; D011726:Pyridinium Compounds / Q000378:metabolism; D000644:Quaternary Ammonium Compounds / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D022161:Transport Vesicles / Q000378:metabolism*; D033921:Vesicular Transport Proteins / Q000235:genetics* / Q000378:metabolism*
D000199:Actins; C092350:FM 4-64; D011726:Pyridinium Compounds; D000644:Quaternary Ammonium Compounds; D033921:Vesicular Transport Proteins; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; C064553:VPS1 protein, S cerevisiae