Nedd4-interacting protein 2, a short half-life membrane protein degraded in lysosomes, negatively controls down-regulation of connexin43.
Nedd4-interacting protein 2 (NDFIP2) has three transmembrane domains and interacts with multiple Nedd4 family ubiquitin ligases through polyprolinetyrosine (PY) motifs located in its N-terminal cytoplasmic domain. It has been postulated that NDFIP2 acts as an adaptor for the ubiquitylation of substrates with Nedd4 ubiquitin ligase. However, whether NDFIP2 promotes or inhibits the ubiquitylation of Nedd4 substrates is still under debate. We show here that although NDFIP2 is detected in the Golgi/trans-Golgi network (TGN) area, it is rapidly delivered to and degraded in lysosomes with its half-life ca. 1.5 h. Intriguingly, knockdown (KD) of NDFIP2 with small interfering RNA (siRNA) impaired both the formation and function of gap junctions. Indeed, KD of NDFIP2 destabilized the gap junction protein connexin43 that contains PY motif. In support of this, overexpression of NDFIP2 stabilized connexin43 and enhanced the formation of gap junctions. Furthermore, the PY motifs of NDFIP2, which are required for its interaction with Nedd4, Atrophin-1 interacting protein (AIP) 4 (AIP4)/Itch, and AIP2/WWP2, were necessary for the targeting of NDFIP2 to lysosomes and/or the stability of connexin43 and gap junctions. Collectively these findings suggest that NDFIP2 may inhibit the Nedd4-dependent ubiquitylation of membrane proteins containing PY motifs, such as connexin43, in a competitive manner.
Division of Pharmaceutical Cell Biology, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka 812-8582, Japan.
Chiho
C
Ohzono
Sachise
S
Etoh
Masaki
M
Matsumoto
Keiichi I
KI
Nakayama
Yuko
Y
Hirota
Yoshitaka
Y
Tanaka
Hideaki
H
Fujita
D000818:Animals; D019556:COS Cells; D002352:Carrier Proteins / Q000378:metabolism*; D002522:Chlorocebus aethiops; D018031:Connexin 43 / Q000378:metabolism*; D045643:DNA Repair Enzymes; D015536:Down-Regulation / Q000187:drug effects; D056827:Endosomal Sorting Complexes Required for Transport / Q000378:metabolism*; D017629:Gap Junctions / Q000502:physiology*; D006056:Golgi Apparatus / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008247:Lysosomes / Q000378:metabolism*; D000075702:Nedd4 Ubiquitin Protein Ligases; D010455:Peptides / Q000378:metabolism; D034741:RNA, Small Interfering; D051381:Rats; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D054875:Ubiquitination; D021601:trans-Golgi Network / Q000378:metabolism
D002352:Carrier Proteins; D018031:Connexin 43; D056827:Endosomal Sorting Complexes Required for Transport; C476254:N4BP2 protein, human; D010455:Peptides; D034741:RNA, Small Interfering; C000618479:NEDD4L protein, rat; D000075702:Nedd4 Ubiquitin Protein Ligases; C000618483:Nedd4 protein, human; C000618484:Nedd4 protein, rat; D044767:Ubiquitin-Protein Ligases; D045643:DNA Repair Enzymes