RESUMO
AAA ATPase VCP and its yeast ortholog Cdc48, in a complex with the Ufd1-Npl4 heterodimer as an adaptor, play an essential role in endoplasmic reticulum-associated degradation (ERAD). Several UBX domain-containing proteins function to recruit ubiquitylated substrates to VCP/Cdc48 by binding both VCP/Cdc48 and other ERAD components such as ubiquitin ligases. Here we show that mammalian UBXD1 is an additional UBX domain-containing protein involved in the ERAD process. UBXD1 is a cytosolic protein that interacts with VCP and Derlin-1. Overexpression of UBXD1 in cells causes selective dissociation of Ufd1 from VCP, resulting in inhibition of mutant cystic fibrosis transmembrane conductance regulator (CFTR) degradation by ERAD. Additionally, depletion of endogenous UBXD1 protein by RNA interference also results in a defect in CFTR degradation. Collectively, these findings suggest that UBXD1 is a regulatory component of ERAD that may modulate the adaptor binding to VCP.
Assuntos
Adenosina Trifosfatases/metabolismo , Proteínas de Transporte/metabolismo , Proteínas de Ciclo Celular/metabolismo , Retículo Endoplasmático/metabolismo , Proteínas de Membrana/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Proteínas Adaptadoras de Transporte Vesicular , Proteínas Relacionadas à Autofagia , Proteínas de Transporte/genética , Células HeLa , Humanos , Estrutura Terciária de Proteína , Proteína com ValosinaRESUMO
NVL (nuclear VCP-like protein), a member of the AAA-ATPase family, is known to exist in two forms with N-terminal extensions of different lengths in mammalian cells. Here, we show that they are localized differently in the nucleus; NVL2, the major species, is mainly present in the nucleolus, whereas NVL1 is nucleoplasmic. Mutational analysis demonstrated the presence of two nuclear localization signals in NVL2, one of which is shared with NVL1. In addition, a nucleolar localization signal was found to exist in the N-terminal extra region of NVL2. The nucleolar localization signal is critical for interaction with ribosomal protein L5, which was identified as a specific interaction partner of NVL2 on yeast two-hybrid screening. The interaction of NVL2 with L5 is ATP-dependent and likely contributes to the nucleolar translocation of NVL2. The physiological implication of this interaction was suggested by the finding that a dominant negative NVL2 mutant inhibits ribosome biosynthesis, which is known to take place in the nucleolus.
