Details

Autor(en) / Beteiligte

• Kallay, Laura M.
• Brett, Christopher L.
• Tukaye, Deepali N.
• Wemmer, Megan A.
• Chyou, Anthony
• Odorizzi, Greg
• Rao, Rajini

Titel

Endosomal Na+ (K+)/H+ Exchanger Nhx1/Vps44 Functions Independently and Downstream of Multivesicular Body Formation

Ist Teil von

• The Journal of biological chemistry, 2011-12, Vol.286 (51), p.44067-44077

Ort / Verlag

United States: Elsevier Inc

Erscheinungsjahr

2011

Link zum Volltext

Quelle

Free E-Journal (出版社公開部分のみ）

Beschreibungen/Notizen

• The multivesicular body (MVB) is an endosomal intermediate containing intralumenal vesicles destined for membrane protein degradation in the lysosome. In Saccharomyces cerevisiae, the MVB pathway is composed of 17 evolutionarily conserved ESCRT (endosomal sorting complex required for transport) genes grouped by their vacuole protein sorting Class E mutant phenotypes. Only one integral membrane protein, the endosomal Na+ (K+)/H+ exchanger Nhx1/Vps44, has been assigned to this class, but its role in the MVB pathway has not been directly tested. Herein, we first evaluated the link between Nhx1 and the ESCRT proteins and then used an unbiased phenomics approach to probe the cellular role of Nhx1. Select ESCRT mutants (vps36Δ, vps20Δ, snf7Δ, and bro1Δ) with defects in cargo packaging and intralumenal vesicle formation shared multiple growth phenotypes with nhx1Δ. However, analysis of cellular trafficking and ultrastructural examination by electron microscopy revealed that nhx1Δ cells retain the ability to sort cargo into intralumenal vesicles. In addition, we excluded a role for Nhx1 in Snf7/Bro1-mediated cargo deubiquitylation and Rim101 response to pH stress. Genetic epistasis experiments provided evidence that NHX1 and ESCRT genes function in parallel. A genome-wide screen for single gene deletion mutants that phenocopy nhx1Δ yielded a limited gene set enriched for endosome fusion function, including Rab signaling and actin cytoskeleton reorganization. In light of these findings and the absence of the so-called Class E compartment in nhx1Δ, we eliminated a requirement for Nhx1 in MVB formation and suggest an alternative post-ESCRT role in endosomal membrane fusion. Background: Nhx1/Vps44 is proposed to be a Class E gene involved in formation of the multivesicular body (MVB). However, this hypothesis has not been tested. Results: Nhx1 is not required for cargo sorting or MVB formation and shows synthetic phenotypes with select ESCRT mutants. Conclusion: Nhx1 functions independently of the ESCRT pathway. Significance: Nhx1 may have a post-ESCRT role in endosomal membrane fusion.