Limited complementarity between U1 snRNA and a retroviral 5' splice site permits its attenuation via RNA secondary structure.
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.
Institute for Virology, Hannover Medical School, 30625 Hannover, Germany.
Daniela
D
Zychlinski
Steffen
S
Erkelenz
Vanessa
V
Melhorn
Christopher
C
Baum
Heiner
H
Schaal
Jens
J
Bohne
D017398:Alternative Splicing; D001483:Base Sequence; D002460:Cell Line; D006801:Humans; D009052:Leukemia Virus, Murine / Q000235:genetics* / Q000502:physiology; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry; D009690:Nucleic Acid Conformation; D017434:Protein Structure, Tertiary; D011533:Proviruses / Q000235:genetics / Q000502:physiology; D022821:RNA Splice Sites; D012342:RNA, Small Nuclear / Q000737:chemistry* / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry; D000068103:Serine-Arginine Splicing Factors; D013489:Suppression, Genetic; D014779:Virus Replication
D009687:Nuclear Proteins; D022821:RNA Splice Sites; D012342:RNA, Small Nuclear; D016601:RNA-Binding Proteins; C405483:U1 small nuclear RNA; D000068103:Serine-Arginine Splicing Factors