PACSINs bind to the TRPV4 cation channel. PACSIN 3 modulates the subcellular localization of TRPV4.
TRPV4 is a cation channel that responds to a variety of stimuli including mechanical forces, temperature, and ligand binding. We set out to identify TRPV4-interacting proteins by performing yeast two-hybrid screens, and we isolated with the avian TRPV4 amino terminus the chicken orthologues of mammalian PACSINs 1 and 3. The PACSINs are a protein family consisting of three members that have been implicated in synaptic vesicular membrane trafficking and regulation of dynamin-mediated endocytotic processes. In biochemical interaction assays we found that all three murine PACSIN isoforms can bind to the amino terminus of rodent TRPV4. No member of the PACSIN protein family was able to biochemically interact with TRPV1 and TRPV2. Co-expression of PACSIN 3, but not PACSINs 1 and 2, shifted the ratio of plasma membrane-associated versus cytosolic TRPV4 toward an apparent increase of plasma membrane-associated TRPV4 protein. A similar shift was also observable when we blocked dynamin-mediated endocytotic processes, suggesting that PACSIN 3 specifically affects the endocytosis of TRPV4, thereby modulating the subcellular localization of the ion channel. Mutational analysis shows that the interaction of the two proteins requires both a TRPV4-specific proline-rich domain upstream of the ankyrin repeats of the channel and the carboxyl-terminal Src homology 3 domain of PACSIN 3. Such a functional interaction could be important in cell types that show distribution of both proteins to the same subcellular regions such as renal tubule cells where the proteins are associated with the luminal plasma membrane.
Cuajungco
Math P
MP
Department of Otolaryngology, Stanford University School of Medicine, Stanford, California 94305, USA.
Grimm
Christian
C
Oshima
Kazuo
K
D'hoedt
Dieter
D
Nilius
Bernd
B
Mensenkamp
Arjen R
AR
Bindels
René J M
RJ
Plomann
Markus
M
Heller
Stefan
S
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002451:Cell Compartmentation; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D002645:Chickens; D003598:Cytoskeletal Proteins; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D009479:Neuropeptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D021381:Protein Transport; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D051381:Rats; D050916:TRPV Cation Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D003598:Cytoskeletal Proteins; D047908:Intracellular Signaling Peptides and Proteins; D009479:Neuropeptides; C114726:Pacsin1 protein, mouse; C485644:Pacsin2 protein, mouse; C486019:Pacsin3 protein, mouse; D010750:Phosphoproteins; D011506:Proteins; D050916:TRPV Cation Channels; C486576:Trpv4 protein, mouse