Research:Genetics of MS

Evsyukova I, Bradrick SS, Gregory SG, Garcia-Blanco MA. Cleavage and polyadenylation specificity factor 1 (CPSF1) regulates alternative splicing of interleukin 7 receptor (IL7R) exon 6. RNA. 2012 Nov 14. [Epub ahead of print]

Interleukin 7 receptor, IL7R, is expressed exclusively on cells of the lymphoid lineage, and its expression is crucial for the development and maintenance of T cells. Alternative splicing of IL7R exon 6 results in membrane-bound (exon 6 included) and soluble (exon 6 skipped) IL7R isoforms. Interestingly, the inclusion of exon 6 is affected by a single-nucleotide polymorphism associated with the risk of developing multiple sclerosis. Given the potential association of exon 6 inclusion with multiple sclerosis, we investigated the elements that regulate exon 6 splicing. We identified multiple exonic (protein coding) and intronic (protein non-coding) elements that impact inclusion of exon 6. Moreover, we utilized RNA affinity chromatography followed by mass spectrometry to identify factors that bind exon 6 and regulate its splicing. These experiments identified cleavage and polyadenylation specificity factor 1 (CPSF1) among protein-binding candidates. A consensus polyadenylation signal is present in intron 6 of IL7R directly downstream from the 5' (DNA part upstream of the gene) splice site. Mutations to this site and CPSF1 knockdown both resulted in an increase in exon 6 inclusion. We found no evidence that this site is used to produce cleaved and polyadenylated mRNAs, suggesting that CPSF1 interaction with intronic IL7R pre-mRNA interferes with spliceosome binding to the exon 6 5' splice site. Our results suggest that competing mRNA splicing and polyadenylation regulate exon 6 inclusion and consequently determine the ratios of soluble to membrane-bound IL7R. This may be relevant for both T cell ontogeny and function and development of multiple sclerosis.

The Interleukin 7 receptor (IL-7R) gene variants are part of the genetic susceptibility to MS. Interleukin-7 is a cytokine that stimulates immune cell development. Genes contain coding (Exons) and non-coding (introns) element in the DNA. When the gene is made in to RNA the introns are removed (sliced away) to make a coding copy of RNA. In the IL-7R the inclusion of exon 6 makes variants made that put the receptor in the cell membrane or result in its secretion outside the cell.

Polyadenylation is the addition of a poly(A) tail to an RNA molecule. The poly(A) tail is a stretch of RNA that has only adenine bases. This is part of the process that produces mature messenger RNA (mRNA) for translation intomaking amino acids and proteins It, therefore, forms part of the larger process of gene expression.The process of polyadenylation begins as the transcription of a gene finishes, or terminates. The poly(A) tail is important for the nuclear export, translation, and stability of mRNA. The tail is shortened over time, and, when it is short enough, the mRNA is enzymatically degraded.

In this study polyadenylation specificity factor 1 (CPSF1) was found to interact with the exon 6 of IL-7R and blockade of CPSF1 increased the inclusion of exon 6 so it would more likely to be membrane bound and signal cell functions. RNAs that influence splicing of the gene and the polyadenylation site will determine whether IL-7R is secreted or cell bound which will influence immune cell development, which may influence whther MS occurs or not.

The MS-associated variant leads to the shed version of the receptor, which can mop IL-7 up as shown recently

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