We have identified gas5 (growth arrest-specific transcript 5) as a non-protein-coding multiple small nucleolar RNA (snoRNA) host gene similar to UHG (U22 host gene). Encoded within the 11 introns of the mouse gas5 gene are nine (10 in human) box C/D snoRNAs predicted to function in the 2-O-methylation of rRNA. The only regions of conservation between mouse and human gas5 genes are their snoRNAs and 5-end sequences. Mapping the 5 end of the mouse gas5 transcript demonstrates that it possesses an oligopyrimidine tract characteristic of the 5-terminal oligopyrimidine (5TOP) class of genes. Arrest of cell growth or inhibition of translation by cycloheximide, pactamycin, or rapamycin-which specifically inhibits the translation of 5TOP mRNAs-results in accumulation of the gas5 spliced RNA. Classification of gas5 as a 5TOP gene provides an explanation for why it is a growth arrest specific transcript: while the spliced gas5 RNA is normally associated with ribosomes and rapidly degraded, during arrested cell growth it accumulates in mRNP particles, as has been reported for other 5TOP messages. Strikingly, inspection of the 5-end sequences of currently known snoRNA host gene transcripts reveals that they all exhibit features of the 5TOP gene family.In the nucleolus of eukaryotic cells, ribosomal DNA is transcribed by RNA polymerase I into long precursor (pre-rRNA) transcripts, which are modified by methylation and pseudouridylation, cleaved to yield 18S, 5.8S, and 28S rRNAs, and then assembled into the mature large and small ribosomal subunits prior to export to the cytoplasm (for reviews see references 22, 26, and 77). A large number of small nucleolar ribonucleoprotein (snoRNP) particles have emerged as key players in this biosynthetic process. Currently more than 70 snoRNA species have been identified (for reviews see references 51, 75, and 82). All snoRNAs, with the exception of MRP RNA, can be divided into two classes: those that possess boxes C (RUGAUGA) and D (CUGA), which are required for association with the abundant nucleolar autoantigen fibrillarin (see reference 51), and those that possess boxes H (ANANNA) and ACA, which mediate the binding of Gar1 protein (4,8,24,37). Only a few snoRNAs have been found to be required for growth in yeast (U3 [6,29] Recently, box C/D snoRNAs and box H/ACA snoRNAs were found to target specific sites in pre-rRNA for 2Ј-O-methylation and pseudouridylation, respectively (for reviews see references 46, 48, 62, 75, and 82). These modification reactions are mediated by extensive regions (10 to 21 nt) of complementarity between the so-called "antisense" snoRNAs and sequences flanking the rRNA sites to be modified. Specifically, U24, U20, and U25 were shown to direct site-specific ribose methylation of pre-rRNA in HeLa cells (39), yeast (13), and Xenopus oocytes (87), respectively, and snR8, snR3, snR33, and snR5 (among others) were demonstrated to target prerRNA for pseudouridylation in yeast (23,57). The presence of ϳ200 modified nucleotides in vertebrate rRNA (47) suggests that more...

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The molecular chaperone and cytoprotective activities of the Hsp70 and Hsp40 chaperones represent therapeutic targets for human diseases such as cancer and those that arise from defects in protein folding; however, very few Hsp70 and no Hsp40 modulators have been described. Using an assay for ATP hydrolysis, we identified and screened small molecules with structural similarity to 15-deoxyspergualin and NSC 630668-R/1 for their effects on endogenous and Hsp40-stimulated Hsp70 ATPase activity. Several of these compounds

trimethyl guanosine (TMG) cap structure. Phylogenetic comparison of U3 snoRNAs from various species re-Department of Molecular Biophysics and vealed conserved sequence elements called boxes C Biochemistry (UGAUGA/U) and D (CUGA), which are also present in Howard Hughes Medical Institute the next vertebrate snoRNAs (U8 and U13) to be identi-Yale University fied. Boxes C and D direct binding of the abundant New Haven, Connecticut 06536nucleolar protein fibrillarin, a target of autoantibodies, and are necessary for the accumulation and stability of these snoRNAs. Yet, as more snoRNAs were discov-Biologists have known for decades that the nucleolus ered, not all exhibited the hallmarks of U3, U8, and U13: is the compartment of the eukaryotic cell most densely some possessed an unmodified terminal 5Ј monophospacked with RNA. But few would have guessed that, in phate instead of the TMG cap and some lacked the addition to the ribosomal RNAs (rRNAs) at various characteristic box C and D sequences (Table 1). Progstages of maturation, the nucleolus contains a multitude ress in assigning functions was made by deletion/rescue of discrete small RNA molecules. These small nucleolar experiments in yeast and in Xenopus oocytes. Certain RNAs, or snoRNAs, are responsible not only for orches-snoRNAs were found to be essential for growth (U3, trating the cleavage events that cut the long pre-rRNA U14, snR10 [temperature sensitive], and snR30) in yeast into 18S, 5.8S, and 28S molecules, but also for adding and/or for specific cleavage steps in pre-rRNA profinishing touches to rRNAs as they assemble into the cessing (U3, U8, and U22) in Xenopus. Curiously, deleultimate products of the nucleolus, the ribosomal subtion of other yeast snoRNA genes had no detectable units. These finishing touches include remodeling of cereffect on growth rate or rRNA maturation (reviewed by tain rRNA uridines into pseudouridines and tagging of Maxwell and Fournier, 1995). numerous ribose moieties with methyl groups. The A completely unanticipated mode of biogenesis for amazing recent realization is that each of these modifithe particularly small (60-90 nucleotides) and less abuncations is directed by its own specific snoRNA that is dant ‫01ف(‬ 4 copies per cell) members of the vertebrate packaged (as are all small nuclear RNAs) into a ribobox C/D snoRNA family then emerged (reviewed by nucleoprotein (snoRNP) particle. Maxwell and Fournier, 1995). Rather than being tran-Suggesting some important function, methylated and scribed from their own genes, these snoRNAs are intronpseudouridylated residues are confined to the most encoded. Liberated by exonucleolytic processing of exhighly conserved portions of rRNA sequences and are cised introns, these stable intronic fragments possess absent from the discarded regions of pre-rRNA. Verte-5Ј monophosphates instead of TMG caps. SnoRNA host brate rRNAs contain approximately 100 methylated suggenes most commonly specify proteins involved in ars, 95 pseudourdines, and 10 methylated bases (Matranslation or ribosome bi...