Folding of the td pre-RNA with the help of the RNA chaperone StpA.
The td group I intron is inserted in the reading frame of the thymidylate synthase gene, which is mainly devoid of structural elements. In vivo, translation of the pre-mRNA is required for efficient folding of the intron into its splicing-competent structure. The ribosome probably resolves exon-intron interactions that interfere with splicing. Uncoupling splicing from translation, by introducing a non-sense codon into the upstream exon, reduces the splicing efficiency of the mutant pre-mRNA. Alternatively to the ribosome, co-expression of genes that encode proteins with RNA chaperone activity promote folding of the td pre-mRNA in vivo. These proteins also efficiently accelerate folding of the td pre-mRNA in vitro. In order to understand the mechanism of action of RNA chaperones, we probed the impact of the RNA chaperone StpA on the structure of the td intron in vivo and compared it with that of the well characterized Cyt-18 protein, which is a group-I-intron-specific splicing factor. We found that the two proteins have opposite effects on the structure of the td intron. While StpA loosens the three-dimensional structure, Cyt-18 tightens it up. Furthermore, mutations that destabilize the intron structure render the mutants sensitive to StpA, whereas splicing of these mutants is rescued by Cyt-18. Our results provide direct evidence for protein-induced conformational changes within a catalytic RNA in vivo. Whereas StpA resolves tertiary contacts enabling the RNA to refold, Cyt-18 contributes to the stabilization of the native three-dimensional structure.
Mayer
O
O
Institute of Microbiology and Genetics, Vienna Biocenter, Dr. Bohrgasse 9/4, A-1030 Vienna, Austria.
Waldsich
C
C
Grossberger
R
R
Schroeder
R
R
D001483:Base Sequence; D004268:DNA-Binding Proteins / Q000737:chemistry*; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry*; D007438:Introns; D018832:Molecular Chaperones / Q000737:chemistry*; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D012313:RNA / Q000737:chemistry*; D012326:RNA Splicing; D016337:RNA, Catalytic; D012333:RNA, Messenger / Q000378:metabolism; D013997:Time Factors
D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D018832:Molecular Chaperones; D016337:RNA, Catalytic; D012333:RNA, Messenger; C090104:StpA protein, E coli; D012313:RNA