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Delineation of the mechanisms of aberrant splicing caused by two unusual intronic mutations in the RSK2 gene involved in Coffin-Lowry syndrome.

ABSTRACT: Coffin-Lowry syndrome (CLS) is caused by mutations in the RSK2 gene encoding a protein kinase of the Ras signalling pathway. We have studied two point mutations which cause aberrant splicing but do not concern the invariant GT or AG nucleotides of splice sites. The first, an A-->G transition at position +3 of the 5' splice site of exon 6, results in vivo and in vitro in exon skipping and premature translation termination. The natural 5' splice site, although intrinsically weak, is not transactivated under normal conditions. Consequently, replacement of an A/U by a G/U base pairing with U1 snRNA reduces its strength below a critical threshold. The second mutation, an A-->G transition 11 nt upstream of exon 5, creates a new AG near the natural 3' splice site. In vitro this synthetic 3' AG is used exclusively by the splicing machinery. In vivo this splicing event is also observed, but is underestimated because the resulting RSK2 mRNA contains premature stop codons which trigger the nonsense-mediated decay process. We show that a particular mechanism is involved in the aberrant splicing of exon 5, implying involvement of the natural 3' AG during the first catalytic step and the new 3' AG during the second step. Thus, our results explain how these mutations cause severe forms of CLS.

SUBMITTER: Zeniou M

PROVIDER: S-EPMC373406 | biostudies-other | 2004

REPOSITORIES: biostudies-other

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Delineation of the mechanisms of aberrant splicing caused by two unusual intronic mutations in the RSK2 gene involved in Coffin-Lowry syndrome.

Zeniou Maria M Gattoni Renata R Hanauer André A Stévenin James J

Nucleic acids research 20040218 3

Coffin-Lowry syndrome (CLS) is caused by mutations in the RSK2 gene encoding a protein kinase of the Ras signalling pathway. We have studied two point mutations which cause aberrant splicing but do not concern the invariant GT or AG nucleotides of splice sites. The first, an A-->G transition at position +3 of the 5' splice site of exon 6, results in vivo and in vitro in exon skipping and premature translation termination. The natural 5' splice site, although intrinsically weak, is not transactiv ...[more]

PMID: 14973203

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Project description:Coffin-Lowry syndrome (CLS) is a syndromic form of X-linked mental retardation that is characterized, in male patients, by psychomotor and growth retardation and various skeletal anomalies. Typical facial changes and specific clinical and radiological hand aspects exhibited by patients are essential clues for the diagnosis. CLS is caused by mutations in a gene that is located in Xp22.2 and that encodes RSK2, a growth-factor-regulated protein kinase. RSK2 mutations are extremely heterogeneous and lead to premature termination of translation and/or loss of phosphotransferase activity. Surprisingly, among a series of 250 patients screened by single-strand conformation polymorphism (SSCP) analysis, in whom a clinical diagnosis of CLS was made, no mutations were detected in 66% (165) of the patients. To determine what proportion of these latter patients have a RSK2 mutation that has not been detected and what proportion have different disorders that are phenotypically similar to CLS, we have, in the present article, investigated, by western blot analysis and in vitro kinase assay, cell lines from 26 patients in whom no mutation was previously identified by SSCP analysis. This approach allowed us to identify seven novel RSK2 mutations: two changes in the coding sequence of RSK2, one intragenic deletion, and four unusual intronic nucleotide substitutions that do not affect the consensus GT or AG splice sites. We have also determined the nucleotide sequence of the promoter region of the RSK2 gene, and we have screened it for mutations. No disease-causing nucleotide change was identified, suggesting that mutations affecting the promoter region are unlikely to account for a large number of patients with CLS. Finally, our results provide evidence that some patients have a disease that is phenotypically very similar to CLS, which is not caused by RSK2 defects. This suggests that there are defects in either additional genes or combinations of genes that may result in a CLS-like phenotype.

Dataset Information

Delineation of the mechanisms of aberrant splicing caused by two unusual intronic mutations in the RSK2 gene involved in Coffin-Lowry syndrome.

Publications

Delineation of the mechanisms of aberrant splicing caused by two unusual intronic mutations in the RSK2 gene involved in Coffin-Lowry syndrome.

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