LIP5 interacts with aquaporin 2 and facilitates its lysosomal degradation.
Vasopressin binding to the V2 receptor in renal principal cells leads to activation of protein kinase A, phosphorylation of aquaporin 2 (AQP2) at Ser256, and the translocation of AQP2 to the apical membrane, resulting in concentration of the urine. In contrast, phorbol ester-induced activation of protein kinase C pathway leads to ubiquitination of AQP2 at Lys270 and its internalization to multivesicular bodies, where it is targeted for lysosomal degradation or stored for recycling. Because little is known about the regulation of AQP2 trafficking, we used the carboxy-terminal tail of constitutively nonphosphorylated AQP2 (S256A) as a bait for interacting proteins in a yeast two-hybrid assay. We isolated lysosomal trafficking regulator-interacting protein 5 (LIP5) and found that LIP5 interacted with the proximal carboxy-terminal tail (L230-D243) of AQP2 in vitro but not with AQP3 or AQP4, which are also expressed in principal cells. Immunohistochemistry revealed that LIP5 co-localized with AQP2 in principal cells. LIP5 binding occurred independent of the state of Ser256 phosphorylation or Lys270 ubiquitination. LIP5 has been shown to facilitate degradation of the EGF receptor; here, LIP5 seemed to bind this receptor. Knockdown of LIP5 in mouse renal cells (mpkCCD) reduced the phorbol ester-induced degradation of AQP2 approximately two-fold. In summary, LIP5 binds cargo proteins and, considering the role of LIP5 in protein sorting to multivesicular bodies, plays a role in the degradation of AQP2, possibly by reducing the formation of late endosomes.
Department of Physiology, Nijmegen Center of Molecular Sciences, 6500 HB Nijmegen, The Netherlands.
Bas W M
BW
van Balkom
Michelle
M
Boone
Giel
G
Hendriks
Erik-Jan
EJ
Kamsteeg
Joris H
JH
Robben
H Christiaan
HC
Stronks
Anne
A
van der Voorde
Francois
F
van Herp
Peter
P
van der Sluijs
Peter M T
PM
Deen
D000818:Animals; D051399:Aquaporin 2 / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D056827:Endosomal Sorting Complexes Required for Transport; D066246:ErbB Receptors / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D007668:Kidney / Q000502:physiology*; D051379:Mice; D008822:Mice, Transgenic; D012441:Saccharomyces cerevisiae / Q000235:genetics; D014162:Transfection; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
C496024:Aqp2 protein, mouse; D051399:Aquaporin 2; D002352:Carrier Proteins; D056827:Endosomal Sorting Complexes Required for Transport; C499837:VTA1 protein, human; D066246:ErbB Receptors; D001616:beta-Galactosidase