Although the La autoantigen binds to the 3' ends of all nascent polymerase III transcripts, its function in vivo has long been unclear. Although S. cerevisiae cells lacking the La protein homolog Lhp1p are viable, cells containing a mutation that disrupts the anticodon stem of tRNA(Ser)CGA require Lhp1p for growth. We demonstrate that for the wild-type pre-tRNA(Ser)CGA and other pre-tRNAs, Lhp1p is required for the normal endonucleolytic removal of the 3' trailer sequence. In cells lacking Lhp1p, the 3' trailer is removed by exonuclease(s). Although maturation of the mutant pre-tRNA(Ser)CGA requires Lhp1p, introduction of a second mutation that restores base pairing eliminates the requirement. We propose that binding by Lhp1p stabilizes pre-tRNAs in conformations that allow the 3' endonucleolytic cleavage to occur.

Double‐stranded RNA (dsRNA) unwinding/modifying activity, which is present in a wide range of eukaryotic cells, has been previously shown to convert up to 50% of adenosine residues to inosines within intermolecular dsRNA. In the present study, we report that this activity also modifies, though slightly less efficiently, intramolecular double‐stranded regions of synthetic RNAs. Our results widen the range of the possible biological substrates for the activity since many stem and loop type RNA secondary structures (intramolecular dsRNA), present in eukaryotic as well as viral transcripts, can potentially serve as substrates. In addition, we have found that the dsRNA unwinding/modifying activity requires a double‐stranded region of at least 15–20 base pairs (bp) for substrate recognition. Furthermore, modification efficiency was found to be critically dependent on the length of the double‐stranded region; as the size decreased below 100 bp, it dropped precipitously. Our results suggest that efficient modification may occur only with relatively long (greater than 100 bp) dsRNA, perhaps because multiple copies of the enzyme must be bound.

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et al., 1998). Binding by Lhp1p to pre-tRNAs is required for the normal pathway of tRNA maturation (Yoo and In all eukaryotic nuclei, the La autoantigen binds Wolin, 1997). While Lhp1p is not essential in wild-type nascent RNA polymerase III transcripts, stabilizing cells, it becomes required when cells contain a mutation these RNAs against exonucleases. Here we report that that disrupts the anticodon stem of an essential tRNA Ser . the La protein also functions in the assembly of certainAs restoration of base pairing in the stem eliminates the RNA polymerase II-transcribed RNAs into RNPs. A requirement for LHP1, Lhp1p may stabilize the mutant mutation in a core protein of the spliceosomal snRNPs, pre-tRNA in the correct structure (Yoo and Wolin, 1997). Smd1p, causes yeast cells to require the La proteinLhp1p also stabilizes certain pre-tRNAs in yeast strains that fail to carry out the 1-methyladenosine modification Lhp1p for growth at low temperatures. Precursors to (Calvo et al., 1999). U1, U2, U4 and U5 RNAs are bound by Lhp1p inTo learn more about the function of Lhp1p, we carried both wild-type and mutant cells. At the permissive out genetic screens to identify mutations that cause yeast temperature, smd1-1 cells contain higher levels of stable cells to require LHP1. Cells containing a mutation in U1 and U5 snRNPs when Lhp1p is present. At low SMD1, which encodes an Sm core protein of the spliceotemperatures, Lhp1p becomes essential for the accumusomal U1, U2, U4 and U5 snRNPs (Rymond, 1993), lation of U4/U6 snRNPs and for cell viability. When require Lhp1p for growth at low temperature. We demon-U4 RNA is added to extracts, the pre-U4 RNA, but strate that Lhp1p functions in the assembly of the RNA not the mature RNA, is bound by Smd1p. These results polymerase II-transcribed U RNAs into snRNPs. In versuggest that, by stabilizing a 3Ј-extended form of U4 tebrate cells, newly transcribed U RNAs are exported to RNA, Lhp1p facilitates efficient Sm protein binding, the cytoplasm, where they assemble with Sm proteins and thus assisting formation of the U4/U6 snRNP.undergo hypermethylation of the 5Ј cap (Mattaj, 1988).