Nucleus–Vacuole Junctions inSaccharomyces cerevisiaeAre Formed Through the Direct Interaction of Vac8p with Nvj1p

Abstract

Vac8p is a vacuolar membrane protein that is required for efficient vacuole inheritance and fusion, cytosol-to-vacuole targeting, and sporulation. By analogy to other armadillo domain proteins, including β-catenin and importin α, we hypothesize that Vac8p docks various factors at the vacuole membrane. Two-hybrid and copurfication assays demonstrated that Vac8p does form complexes with multiple binding partners, including Apg13p, Vab2p, and Nvj1p. Here we describe the surprising role of Vac8p-Nvj1p complexes in the formation of nucleus–vacuole (NV) junctions. Nvj1p is an integral membrane protein of the nuclear envelope and interacts with Vac8p in the cytosol through its C-terminal 40–60 amino acids (aa). Nvj1p green fluorescent protein (GFP) concentrated in small patches or rafts at sites of close contact between the nucleus and one or more vacuoles. Previously, we showed that Vac8p-GFP concentrated in intervacuole rafts, where is it likely to facilitate vacuole-vacuole fusion, and in “orphan” rafts at the edges of vacuole clusters. Orphan rafts of Vac8p red-sifted GFP (YFP) colocalize at sites of NV junctions with Nvj1p blue-sifted GFP (CFP). GFP-tagged nuclear pore complexes (NPCs) were excluded from NV junctions. In vac8-Δ cells, Nvj1p-GFP generally failed to concentrate into rafts and, instead, encircled the nucleus. NV junctions were absent in both nvj1-Δ andvac8-Δ cells. Overexpression of Nvj1p caused the profound proliferation of NV junctions. We conclude that Vac8p and Nvj1p are necessary components of a novel interorganelle junction apparatus.

Endoplasmic Reticulum Structure and Interconnections with Other Organelles

“…In the first reported example of ER-organelle membrane contact sites, the nucleus-vacuole junction in yeast cells was identified; these membrane contact sites are mediated by the direct interaction between the ER membrane protein Nvj1 and the vacuole protein Vac8 (Pan et al 2000). The nucleus-vacuole junction is not present in mammalian cells, but other endosome compartments have been observed interacting with the ER; high-resolution immuno-EM studies showed the colocalization of the ER localized phophatase PTP1B with the endocytic cargo EGFR (Fig.…”

Glycerolipid transfer for the building of membranes in plant cells

“…To date, only one protein "bridge" between two organelles has been characterized between the nucleus and the vacuole in yeast [94]. This "bridge" contains one nuclear envelope outer [95].…”

Protein evolution: structure‐function relationships of the oncogene beta‐catenin in the evolution of multicellular animals

“…In the slime mold Dictyostelium discoideum, a betacatenin like gene, Aardvark, seems to fulfill beta-catenin like properties in adhesion and cell specification (Grimson et al, 2000). In yeast, the closest match to metazoan beta-catenin is the vacuolar membrane protein Vac8p (Wang et al, '96;Pan and Goldfarb, '98;Wang et al, '98;Pan et al, 2000) that forms complexes with several proteins. However, the proteins that are known to complex with Vac8p in yeast are not homologous to the proteins that complex with beta-catenin in metazoans.…”