Heterogeneous Nuclear Ribonucleoproteins: Role in RNA Splicing

• DOI: 10.1126/science.3952495
• Publication Date: 2006-10-05T20:28:52Z
• Authors: Yang Do Choi; Paula J. Grabowski; Phillip A. Sharp; Gideon Dreyfuss
• Keywords: 0301 basic medicine; 0303 health sciences; Macromolecular Substances; RNA Splicing; Antibodies, Monoclonal; In Vitro Techniques; Heterogeneous-Nuclear Ribonucleoproteins; 3. Good health; 03 medical and health sciences; Adenosine Triphosphate; Ribonucleoproteins; Humans; RNA, Heterogeneous Nuclear; HeLa Cells

Splicing in vitro of a messenger RNA (mRNA) precursor (pre-mRNA) is inhibited by a monoclonal antibody to the C proteins (anti-C) of the heterogeneous nuclear RNA (hnRNA)-ribonucleoprotein (hnRNP) particles. This antibody, 4F4, inhibits an early step of the reaction: cleavage at the 3′ end of the upstream exon and the formation of the intron lariat. In contrast, boiled 4F4, or a different monoclonal antibody (designated 2B12) to the C proteins, or antibodies to other hnRNP proteins (120 and 68 kilodaltons) and nonimmune mouse antibodies have no inhibitory effect. The 4F4 antibody does not prevent the adenosine triphosphate-dependent formation of a 60 S splicing complex (spliceosome). Furthermore, the 60 S splicing complex contains C proteins, and it can be immunoprecipitated with 4F4. Depletion of C proteins from the splicing extract by immunoadsorption with either of the two monoclonal antibodies to the C proteins (4F4 or 2B12) results in the loss of splicing activity, whereas mock-depletion with nonimmune mouse antibodies has no effect. A 60 S splicing complex does not form in a C protein-depleted nuclear extract. These results indicate an essential role for proteins of the hnRNP complex in the splicing of mRNA precursors.