The RNA-binding protein hnRNPLL induces a T cell alternative splicing program delineated by differential intron retention in polyadenylated RNA

Background

Retention of a subset of introns in spliced polyadenylated mRNA is emerging as a frequent, unexplained finding from RNA deep sequencing in mammalian cells.

Results

Here we analyze intron retention in T lymphocytes by deep sequencing polyadenylated RNA. We show a developmentally regulated RNA-binding protein, hnRNPLL, induces retention of specific introns by sequencing RNA from T cells with an inactivating Hnrpll mutation and from B lymphocytes that physiologically downregulate Hnrpll during their differentiation. In Ptprc mRNA encoding the tyrosine phosphatase CD45, hnRNPLL induces selective retention of introns flanking exons 4 to 6; these correspond to the cassette exons containing hnRNPLL binding sites that are skipped in cells with normal, but not mutant or low, hnRNPLL. We identify similar patterns of hnRNPLL-induced differential intron retention flanking alternative exons in 14 other genes, representing novel elements of the hnRNPLL-induced splicing program in T cells. Retroviral expression of a normally spliced cDNA for one of these targets, Senp2, partially corrects the survival defect of Hnrpll-mutant T cells. We find that integrating a number of computational methods to detect genes with differentially retained introns provides a strategy to enrich for alternatively spliced exons in mammalian RNA-seq data, when complemented by RNA-seq analysis of purified cells with experimentally perturbed RNA-binding proteins.

Conclusions

Our findings demonstrate that intron retention in mRNA is induced by specific RNA-binding proteins and suggest a biological significance for this process in marking exons that are poised for alternative splicing.     

Intriguing balancing selection on the intron 5 region of LMBR1 in human population

Background

The intron 5 of gene LMBR1 is the cis-acting regulatory module for the sonic hedgehog (SHH) gene. Mutation in this non-coding region is associated with preaxial polydactyly, and may play crucial roles in the evolution of limb and skeletal system.

Methodology/Principal Findings

We sequenced a region of the LMBR1 gene intron 5 in East Asian human population, and found a significant deviation of Tajima''s D statistics from neutrality taking human population growth into account. Data from HapMap also demonstrated extended linkage disequilibrium in the region in East Asian and European population, and significantly low degree of genetic differentiation among human populations.

Conclusion/Significance

We proposed that the intron 5 of LMBR1 was presumably subject to balancing selection during the evolution of modern human.