Members of the RSC chromatin-remodeling complex are required for maintaining proper nuclear envelope structure and pore complex localization.
The assembly, distribution, and functional integrity of nuclear pore complexes (NPCs) in the nuclear envelope (NE) are key determinants in the nuclear periphery architecture. However, the mechanisms controlling proper NPC and NE structure are not fully defined. We used two different genetic screening approaches to identify Saccharomyces cerevisiae mutants with defects in NPC localization. The first approach examined green fluorescent protein (GFP)-Nic96 in 531 strains from the yeast Tet-promoters Hughes Collection with individual essential genes expressed from a doxycycline-regulated promoter (TetO(7)-orf). Under repressive conditions, depletion of the protein encoded by 44 TetO(7)-orf strains resulted in mislocalized GFP-Nic96. These included STH1, RSC4, RSC8, RSC9, RSC58, ARP7, and ARP9, each encoding components of the RSC chromatin remodeling complex. Second, a temperature-sensitive sth1-F793S (npa18-1) mutant was identified in an independent genetic screen for NPC assembly (npa) mutants. NPC mislocalization in the RSC mutants required new protein synthesis and ongoing transcription, confirming that lack of global transcription did not underlie the phenotypes. Electron microscopy studies showed significantly altered NEs and nuclear morphology, with coincident cytoplasmic membrane sheet accumulation. Strikingly, increasing membrane fluidity with benzyl alcohol treatment prevented the sth1-F793S NE structural defects and NPC mislocalization. We speculate that NE structure is functionally linked to proper chromatin architecture.
Titus
Laura C
LC
Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232-8240, USA.
Dawson
T Renee
TR
Rexer
Deborah J
DJ
Ryan
Kathryn J
KJ
Wente
Susan R
SR
D042002:Chromatin Assembly and Disassembly; D046911:Macromolecular Substances / Q000378:metabolism*; D009154:Mutation; D009685:Nuclear Envelope / Q000378:metabolism / Q000648:ultrastructure*; D022022:Nuclear Pore / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure*; D028861:Nuclear Pore Complex Proteins / Q000235:genetics / Q000378:metabolism; D010641:Phenotype; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism
D046911:Macromolecular Substances; C082533:NIC96 protein, S cerevisiae; D028861:Nuclear Pore Complex Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins