YEAST NOP2 ENCODES AN ESSENTIAL NUCLEOLAR PROTEIN WITH HOMOLOGY TO A HUMAN PROLIFERATION MARKER

We have isolated a gene (NOP2) encoding a nucleolar protein during a s earch for previously unidentified nuclear proteins in the yeast Saccha romyces cerevisiae. The protein encoded by NOP2 (Nop2p) has a predicte d molecular mass of 70 kD, migrates at 90 kD by SDS-PAGE, and is essen tial for cell viability. Nop2p shows significant amino acid sequence h omology to a human proliferation-associated nucleolar protein, p120. A pproximately half of Nop2p exhibits 67% amino acid sequence identity t o p120. Analysis of subcellular fractions indicates that Nop2p is loca ted primarily in the nucleus, and nuclear fractionation studies sugges t that Nop2p is associated with the nucleolus. Indirect immunofluoresc ence localization of Nop2p shows a nucleolar-staining pattern, which i s heterogeneous in appearance, and a faint staining of the cytoplasm. The expression of NOP2 during the transition from stationary phase gro wth arrest to rapid growth was measured, and compared to the expressio n of TCM1, which encodes the ribosomal protein L3. Nop2p protein level s are markedly upregulated during the onset of growth, compared to the levels of ribosomal protein L3, which remain relatively constant. NOP 2 mRNA levels also increase during the onset of growth, accompanied by a similar increase in the levels of TCM1 mRNA. The consequences of ov erexpressing NOP2 from the GAL10 promoter on a multicopy plasmid were investigated. Although NOP2 overexpression produced no discernible gro wth phenotype and had no effect on ribosome subunit synthesis, overexp ression was found to influence the morphology of the nucleolus, as jud ged by electron microscopy. Overexpression caused the nucleolus to bec ome detached from the nuclear envelope and to become more rounded and/ or fragmented in appearance. These findings suggest roles for NOP2 in nucleolar function during the onset of growth, and in the maintenance of nucleolar structure.