Global alternative splicing analysis of cryptic splicing suppressors in C. elegans
Ontology highlight
ABSTRACT:
ORGANISM(S): Caenorhabditis elegans
PROVIDER: GSE113275 | GEO | 2018/07/17
REPOSITORIES: GEO
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Project description:The spliceosome undergoes extensive rearrangements as it assembles in multiple steps onto the precursor messenger RNA. In the earliest assembly step, U1snRNA identifies the 5' splice site through base-pairing interactions. However, U1snRNA leaves the spliceosome relatively early in the assembly process. The 5' splice site identity is subsequently maintained through interactions with U6snRNA, protein factor PRP8, and other components of the spliceosome during the complex assembly and rearrangements that build the catalytic site. Using a forward genetic screen in C. elegans, we have identified splicing suppressors of a locomotion defect caused by a 5'ss mutation. Here we report three new extragenic suppressor alleles from this screen, two in PRP8 and one in SNRNP200/Brr2. mRNASeq studies of these suppressor strains indicates that there are specific native targets with alternative 5' and alternative 3' splicing events affected by these suppressors, especially for the suppressor PRP8 D 1549N (position D1556 in humans). The strong suppressor at the unstructured N-terminus of SNRP200, N18K, indicates a potential regulatory role for this region. By examining distinct changes in the splicing of native genes, and by mapping these conserved suppressor residues onto cryoEM structural models of assembling human spliceosomes, we conclude that there are multiple interactions in the spliceosome that are required to ensure that the initial 5'ss identified by U1snRNA early in spliceosome assembly is the one that gets loaded into the catalytic core.The spliceosome undergoes extensive rearrangements as it assembles in multiple steps onto the precursor messenger RNA. In the earliest assembly step, U1snRNA identifies the 5' splice site through base-pairing interactions. However, U1snRNA leaves the spliceosome relatively early in the assembly process. The 5' splice site identity is subsequently maintained through interactions with U6snRNA, protein factor PRP8, and other components of the spliceosome during the complex assembly and rearrangements that build the catalytic site. Using a forward genetic screen in C. elegans, we have identified splicing suppressors of a locomotion defect caused by a 5'ss mutation. Here we report three new extragenic suppressor alleles from this screen, two in PRP8 and one in SNRNP200/Brr2. mRNASeq studies of these suppressor strains indicates that there are specific native targets with alternative 5' and alternative 3' splicing events affected by these suppressors, especially for the suppressor PRP8 D 1549N (position D1556 in humans). The strong suppressor at the unstructured N-terminus of SNRP200, N18K, indicates a potential regulatory role for this region. By examining distinct changes in the splicing of native genes, and by mapping these conserved suppressor residues onto cryoEM structural models of assembling human spliceosomes, we conclude that there are multiple interactions in the spliceosome that are required to ensure that the initial 5'ss identified by U1snRNA early in spliceosome assembly is the one that gets loaded into the catalytic core.
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