Ribonucleoprotein organization of eukaryotic RNA. XXXII. U2 small nuclear RNA precursors and their accurate 3′ processing in vitro as ribonucleoprotein particles

Eric D Wieben, Joan M. Nenninger, Thoru Pederson

Research output: Contribution to journalArticle

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Abstract

Biosynthetic precursors of U2 small nuclear RNA have been identified in cultured human cells by hybrid-selection of pulse-labeled RNA with cloned U2 DNA. These precursor molecules are one to approximately 16 nucleotides longer than mature U2 RNA and contain 2,2,7-trimethylguanosine "caps". The U2 RNA precursors are associated with proteins that react with a monoclonal antibody for antigens characteristic of small nuclear ribonucleoprotein particles. Like previously described precursors of U1 and U4 small nuclear RNAs, the pre-U2 RNAs are recovered in cytoplasmic fractions, although it is not known if this is their location in vivo. The precursors are processed to mature-size U2 RNA when cytoplasmic extracts are incubated in vitro at 37 °C. Mg2+ is required but ATP is not. The ribonucleoprotein structure of the pre-U2 RNA is maintained during the processing reaction in vitro, as are the 2,2,7-trimethylguanosine caps. The ribonucleoprotein organization is of major importance, as exogenous, protein-free U2 RNA precursors are degraded rapidly in the in vitro system. Two lines of evidence indicate that the conversion of U2 precursors to mature-size U2 RNA involves a 3′ processing reaction. First, the reaction is unaffected by a large excess of mature U2 small nuclear RNP, whose 5′ trimethylguanosine caps would be expected to compete for a 5′ processing activity. Second, when pre-U2 RNA precursors are first stoichiometrically decorated with an antibody specific for 2,2,7-trimethylguanosine, the extent of subsequent processing in vitro is unaffected. These results provide the first demonstration of a eukaryotic RNA processing reaction in vitro occurring within a ribonucleoprotein particle.

Original languageEnglish (US)
Pages (from-to)69-78
Number of pages10
JournalJournal of Molecular Biology
Volume183
Issue number1
DOIs
StatePublished - May 5 1985
Externally publishedYes

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Ribonucleoproteins
RNA Precursors
RNA
Small Nuclear Ribonucleoproteins
U2 small nuclear RNA
In Vitro Techniques
Cultured Cells
Proteins
Nucleotides
Adenosine Triphosphate
Monoclonal Antibodies
Antigens
Antibodies
DNA

ASJC Scopus subject areas

  • Virology

Cite this

Ribonucleoprotein organization of eukaryotic RNA. XXXII. U2 small nuclear RNA precursors and their accurate 3′ processing in vitro as ribonucleoprotein particles. / Wieben, Eric D; Nenninger, Joan M.; Pederson, Thoru.

In: Journal of Molecular Biology, Vol. 183, No. 1, 05.05.1985, p. 69-78.

Research output: Contribution to journalArticle

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