It is increasingly clear that pre-mRNA-protein complexes (also known as heterogenous nuclear RNA-protein complexes or hnRNP) play an important role in RNA splicing. We propose that one of the functions of hnRNP in RNA splicing is sequestration of RNA splicing center and stabilization of the quaternary structure in which the splice junctions are in a close proximity during RNA splicing. Certain pre-mRNA structures in which the splice junctions are artificially brought into a close proximity appear to be spliced in vitro in the absence of hnRNP formation and perhaps mimic the conformation of hnRNP involving the splicing reaction center. We believe that careful analysis of various pre-mRNA structures will provide important information on the role of hnRNP in RNA splicing. In this application we will construct various pre-mRNA structures in which the splice junctions are in a close proximity by hybridization of chicken b-globin and mouse metallothionein-I pre-mRNAs with cRNA and also by forming a stem structure with complementary exon sequences. Such pre- mRNAs will be spliced in HeLa nuclear extracts and the spliced products and intermediates will be analyzed by high resolution polyacrylamide gel electrophoresis. In addition, we will investigate binding of hnRNP proteins, assembly of hnRNP, requirement of snRNPs for splicing, and possible sequestration of splicing reaction in the absence of hnRNP formation.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM036975-03
Application #
3291746
Study Section
Physiological Chemistry Study Section (PC)
Project Start
1987-04-01
Project End
1990-03-31
Budget Start
1989-04-01
Budget End
1990-03-31
Support Year
3
Fiscal Year
1989
Total Cost
Indirect Cost
Name
University of North Carolina Chapel Hill
Department
Type
Schools of Medicine
DUNS #
078861598
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599