Nucleic Acids Research Advance Access published online on October 23, 2009
Nucleic Acids Research, doi:10.1093/nar/gkp694
Gene Regulation, Chromatin and Epigenetics |
Limited complementarity between U1 snRNA and a retroviral 5' splice site permits its attenuation via RNA secondary structure
1Institute for Virology, Hannover Medical School, 30625 Hannover, 2Institute for Virology, Heinrich-Heine-University, 40225 Düsseldorf and 3Department of Experimental Hematology, Hannover Medical School, 30625 Hannover, Germany
*To whom correspondence should be addressed. Email: bohne.jens{at}mh-hannover.de The authors wish it to be known that, in their opinion, the second and third authors should be regarded as joint Second Authors.
Received April 1, 2009. Revised June 26, 2009. Accepted August 6, 2009.
Multiple types of regulation are used by cells and viruses to control alternative splicing. In murine leukemia virus, accessibility of the 5' splice site (ss) is regulated by an upstream region, which can fold into a complex RNA stem–loop structure. The underlying sequence of the structure itself is negligible, since most of it could be functionally replaced by a simple heterologous RNA stem–loop preserving the wild-type splicing pattern. Increasing the RNA duplex formation between U1 snRNA and the 5'ss by a compensatory mutation in position +6 led to enhanced splicing. Interestingly, this mutation affects splicing only in the context of the secondary structure, arguing for a dynamic interplay between structure and primary 5'ss sequence. The reduced 5'ss accessibility could also be counteracted by recruiting a splicing enhancer domain via a modified MS2 phage coat protein to a single binding site at the tip of the simple RNA stem–loop. The mechanism of 5'ss attenuation was revealed using hyperstable U1 snRNA mutants, showing that restricted U1 snRNP access is the cause of retroviral alternative splicing.