NUP145 ENCODES A NOVEL YEAST GLYCINE-LEUCINE-PHENYLALANINE-GLYCINE (GLFG) NUCLEOPORIN REQUIRED FOR NUCLEAR-ENVELOPE STRUCTURE

We have isolated and characterized the gene encoding a fourth yeast gl ycine-leucine-phenylalanine-glycine (GLFG) repeat nucleoporin with a c alculated molecular mass of 145.3 kD, and therefore termed NUP145. The amino-terminal half of Nup145p is similar to two previously identifie d GLFG nucleoporins, Nup116p and Nup100p (Wente, S. R., M. P. Rout, an d G. Blobel 1992. J. Cell Biol. 119:705-723). A deletion/disruption in the aminoterminal half of NUP145 (nupl45 Delta N) had only a slight e ffect on cell growth at temperatures between 17 and 37 degrees C. Howe ver, immunofluorescence microscopy of nul45 Delta N cells with antinuc leoporin antibodies showed that the characteristic punctate nuclear st aining normally seen in wild-type yeast cells was reduced, with the ma jority of the signal located in one or two intense spots at the nuclea r periphery. Thin section electron microscopy analysis revealed the pr esence of what appeared to be successive herniations of the nuclear en velope forming grape-like structures at primarily one site on the nupl 45 Delta N nuclei. These successive herniations contained numerous NPC -like structures, correlating to the limited bright patches of antinuc leoporin immunofluorescence signal. In some cases the successive herni ations were small. Occasionally however, multi-lobulated nuclei were s een. We suggest that the ultrastructural phenotype of nupl45 Delta N c ells is due to a defective interaction of nupl45 Delta N NPCs with the surrounding pore membrane domain of the nuclear envelope. We have als o analyzed the synthetic lethal phenotypes among GLFG nucleoporin muta nt alleles, and found that strains harboring nup116 and either nup100 or nup145 mutations were not viable. This, in combination with the mor phological analysis, may reflect overlapping yet distinct roles for th ese three GLFG nucleoporins in NPC-nuclear envelope interactions.