Department of Biochemistry and Molecular Pharmacology
*3' Untranslated Regions; Alternative Splicing; Antigens, Polyomavirus Transforming; Base Sequence; Biological Transport; Cell Line, Transformed; Cell Nucleus; Humans; Molecular Sequence Data; *Mutagenesis; Nucleic Acid Conformation; *RNA Precursors; *RNA Processing, Post-Transcriptional; *RNA, Small Nuclear; Ribonuclease T1; Ribonuclease, Pancreatic
Life Sciences | Medicine and Health Sciences
The biosynthesis of U1, U2, U4 and U5 spliceosomal small nuclear RNAs (snRNAs) involves the nuclear export of precursor molecules extended at their 3' ends, followed by a cytoplasmic phase during which the pre-snRNAs assemble into ribonucleoprotein particles and undergo hypermethylation of their 5' caps and 3' end processing prior to nuclear import. Previous studies have demonstrated that the assembly of pre-snRNAs into ribonucleoprotein particles containing the Sm core proteins is essential for nuclear import in mammalian cells but that 5' cap hypermethylation is not. In the present investigation we have asked whether or not 3' end processing is required for nuclear import of U2 RNA. We designed human pre-U2 RNAs that carried modified 3' tails, and identified one that was stalled (or greatly slowed) in 3' end processing, leading to its accumulation in the cytoplasm of human cells. Nonetheless, this 3' processing arrested pre-U2 RNA molecule was found to undergo cytoplasmic assembly into Sm protein-containing complexes to the same extent as normal pre-U2 RNA. The Sm protein-associated, unprocessed mutant pre-U2 RNA was not observed in the nuclear fraction. Using an assay based on suppression of a genetically blocked SV40 pre-mRNA splicing pathway, we found that the 3' processing deficient U2 RNA was significantly reduced in its ability to rescue splicing, consistent with its impaired nuclear import.
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Citation: Nucleic Acids Res. 1999 Feb 15;27(4):1025-31.
Nucleic acids research
Huang, Qian and Pederson, Thoru, "A human U2 RNA mutant stalled in 3' end processing is impaired in nuclear import" (1999). Open Access Articles. 1700.