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NOG1 is a nucleolar GTP-binding protein present in eukaryotes ranging from trypanosomes to humans. In this report we demonstrate that NOG1 is functionally linked to ribosome biogenesis. In sucrose density gradients Trypanosoma brucei NOG1 co-sediments with 60 S ribosomal subunits but not with monosomes. 60 S precursor RNAs are co-precipitated with NOG1. Together with the nucleolar localization of NOG1, these data indicate that NOG1 is associated with a precursor particle to the 60 S subunit. Disruption of NOG1 function through RNA interference led to a dramatic decrease in the levels of free 60 S particles and the appearance of an atypical rRNA intermediate in which ITS2 was not cleaved. Overexpression of mutant nog1 with a defect in its GTP binding motif on a wild type background caused a modest defect in 60 S biogenesis and a relative decrease in processing of the large subunit rRNAs. In contrast to the mutant protein, neither the N-terminal half of NOG1, which contains the GTP binding motifs, nor the C-terminal half of NOG1 associated with pre-ribosomal particles, although both localized to the nucleolus.

Objective: To evaluate elderly women's knowledge of their skeletal status, assess adequacy of calcium intake, determine the prevalence of low bone density, and determine whether peripheral bone density testing led to medical interventions in a group of rural, elderly Wisconsin women recruited in community pharmacies., Design: Recruiting notices were posted in each pharmacy, and eligible women were enrolled in the order in which they volunteered. Each completed a fracture-risk questionnaire. Calcaneal bone density was measured within the following 6 weeks, using peripheral dual-energy X-ray absorptiometry. Mail surveys were used to assess interventions subsequent to the womens' study participation., Setting: The study was conducted at 5 community pharmacies in rural Wisconsin., Results: Of 133 women, 20% had calcaneal osteoporosis, defined as a T score < or =2.5 (calcaneal bone density <2.5 SDs below the young reference database). Thirty percent of women met National Osteoporosis Foundation (NOF) treatment criteria based on heel bone density and NOF-designated risk factors. Of those meeting treatment criteria, 75% were unaware of their low bone mass. Half of the women received <1200 mg/d of calcium, the recommended dose for osteoporosis prevention. Those who were taking a calcium supplement were much more likely to receive the recommended amount. Women who had discussed bone density test results with their physicians were more likely to receive central dual energy X-ray absorptiometry (DXA) measurements and/or start antiresorptive therapy than women who did not., Conclusions: Rural, elderly Wisconsin women are at substantial risk for osteoporosis, based on calcaneal bone density, but most are unaware of their risk. Compounding this risk is low calcium intake. Community screening of rural, elderly women by peripheral bone density measurement can lead to medical interventions in such individuals.

Objectives: To assess the feasibility of establishing an osteoporosis screening program in rural community pharmacies based on information and resources provided by the National Osteoporosis Foundation (NOF), to survey primary care providers regarding the usefulness of this screening program, and to recommend strategies for pharmacists interested in working with patients at risk for osteoporosis., Design and Participants: Pharmacists and/or nurses enrolled women 65 years of age and older into the study, measured calcaneal bone density, administered a questionnaire to ascertain subjects' osteoporosis risk factors, and provided NOF literature to subjects. With their agreement, women's bone mass data and risk factor assessments were provided to primary care providers along with NOF's Physician's Guide to Prevention and Treatment of Osteoporosis. These providers were surveyed as to whether they found this information useful., Setting: Five independent community pharmacies in rural Wisconsin., Results: We enrolled and tested 133 women. Of these, 122 (92%) agreed to have information mailed to their primary health care providers. These 57 providers were surveyed and 24 (42%) responded; of these 24, 20 (83%) found the information they received useful., Conclusion: A community pharmacy-based osteoporosis screening program using NOF materials was well accepted by physicians. NOF resources and recommendations can provide a strong foundation for such programs.

We describe here a novel, evolutionarily conserved set of predicted G-proteins. The founding member of this family, TbNOG1, was identified in a two-hybrid screen as a protein that interacts with NOPP44/46, a nucleolar phosphoprotein of Trypanosoma brucei. The biological relevance of the interaction was verified by co-localization and co-immunoprecipitation. TbNOG1 localized to the trypanosome nucleolus and interacted with domains of NOPP44/46 that are found in several other nucleolar proteins. Genes encoding proteins highly related to TbNOG1 are present in yeast and metazoa, and related G domains are found in bacteria. We show that NOG1 proteins in humans and Saccharomyces cerevisae are also nucleolar. The S. cerevisae NOG1 gene is essential for cell viability, and mutations in the predicted G motifs abrogate function. Together these data suggest that NOG1 may play an important role in nucleolar functions. The GTP-binding region of TbNOG1 is similar to those of Obg and DRG proteins, which, together with NOG, form a newly recognized family of G-proteins, herein named ODN. The ODN family differs significantly from other G-protein families, and shows several diagnostic sequence characteristics. All organisms appear to possess an ODN gene, pointing to the biological significance of this family of G-proteins.

Methylation of cytosine residues in eukaryotic DNA is common, but poorly understood. Typically several percent of the cytosines are methylated; however, it is unclear what governs which sequences eventually become modified. Neurospora crassa DNA containing the "zeta-eta" (zeta-eta) region, which is a region of unusually heavy methylation, was tested for its ability to direct DNA methylation de novo. DNA stripped of its methylation by propagation in Escherichia coli was reintroduced into Neurospora crassa by transformation. The zeta-eta region reproducibly became "properly" methylated whether inserted at its native chromosomal position or at ectopic sites. Adjacent Neurospora and bacterial sequences in the transforming DNA rarely became methylated. A model is presented that accounts for position-independent faithful methylation as observed in the zeta-eta region, as well as position-dependent methylation, as occasionally observed, especially with sequences not native to Neurospora.

In the Neurospora genome duplicate sequences are detected and altered in the sexual phase. Both copies of duplicate genes are inactivated at high frequency, whether or not they are linked. Restriction sites change, and affected sequences typically become heavily methylated. To characterize the alterations of the DNA, duplicated sequences were isolated before and after one or more sexual cycles. DNA sequencing and heteroduplex analyses demonstrated that the process (termed RIP) produces exclusively G-C to A-T mutations. Changes occur principally at sites where adenine is 3' of the changed cytosine. A sequence duplicated at a distant site in the genome lost approximately 10 percent of its G-C pairs in one passage through a cross. A closely linked duplication of the same sequence that was passed twice through a cross lost about half of its G-C pairs. The results suggest a mechanism for the RIP process.

Introduction of DNA into Neurospora crassa can lead to sequence instability in the sexual phase of the life cycle. Sequence instability was investigated by using a set of strains transformed with single copies of a plasmid including host sequences, Neurospora sequences deleted from the host genome, and foreign sequences. The sequences already represented in the host were rearranged at high frequency in a cross. In general, both elements of the duplication, that from the plasmid and that from the host, became rearranged, whether or not they were linked. Unique sequences were left unaltered. Cytosine residues in the rearranged sequences typically became methylated de novo. Results from tetrad analyses indicated that the rearrangements occur before meiosis, during a stage between fertilization and karyogamy. We suggest that this previously unrecognized genetic process, RIP (rearrangement induced premeiotically), may contribute diversity for evolution and also maintain the gross organization of the genome.