Kar5p is required for multiple functions in both inner and outer nuclear envelope fusion in Saccharomyces cerevisiae.

TitleKar5p is required for multiple functions in both inner and outer nuclear envelope fusion in Saccharomyces cerevisiae.
Publication TypeJournal Article
Year of Publication2014
AuthorsRogers, JV, Rose, MD
JournalG3 (Bethesda)
Volume5
Issue1
Pagination111-21
Date Published2014 Dec 02
ISSN2160-1836
KeywordsMembrane Fusion, Membrane Proteins, Nuclear Envelope, Nuclear Proteins, Protein Structure, Tertiary, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins
Abstract

<p>During mating in the budding yeast Saccharomyces cerevisiae, two haploid nuclei fuse via two sequential membrane fusion steps. SNAREs (i.e., soluble N-ethylmaleimide-sensitive factor attachment protein receptors) and Prm3p mediate outer nuclear membrane fusion, but the inner membrane fusogen remains unknown. Kar5p is a highly conserved transmembrane protein that localizes adjacent to the spindle pole body (SPB), mediates nuclear envelope fusion, and recruits Prm3p adjacent to the SPB. To separate Kar5p's functions, we tested localization, Prm3p recruitment, and nuclear fusion efficiency in various kar5 mutants. All domains and the conserved cysteine residues were essential for nuclear fusion. Several kar5 mutant proteins localized properly but did not mediate Prm3p recruitment; other kar5 mutant proteins localized and recruited Prm3p but were nevertheless defective for nuclear fusion, demonstrating additional functions beyond Prm3p recruitment. We identified one Kar5p domain required for SPB localization, which is dependent on the half-bridge protein Mps3p. Electron microscopy revealed a kar5 mutant that arrests with expanded nuclear envelope bridges, suggesting that Kar5p is required after outer nuclear envelope fusion. Finally, a split-GFP assay demonstrated that Kar5p localizes to both the inner and outer nuclear envelope. These insights suggest a mechanism by which Kar5p mediates inner nuclear membrane fusion. </p>

DOI10.1534/g3.114.015800
Alternate JournalG3 (Bethesda)
PubMed ID25467943
PubMed Central IDPMC4291462
Grant ListR01 GM037739 / GM / NIGMS NIH HHS / United States
T32 GM007388 / GM / NIGMS NIH HHS / United States
GM007388 / GM / NIGMS NIH HHS / United States
GM037739 / GM / NIGMS NIH HHS / United States