Characterization of Arabidopsis thaliana SR protein genes: mutations, alternative splicing, and ESE selection

Abstract

RNA processing in eukaryotes is a highly complex process requiring numerous steps and factors that can play roles in the regulation of functional protein production. SR proteins are a well-defined family of splicing factors identified by a conserved RNA Recognition Motif (RRM) and carboxyl-terminal arginine/serine (RS) repeats. SR proteins are known to bind to mRNA precursors via Exonic Splicing Enhancers, and to recruit U2AF and the U1 snRNP to promote splicing.

I have identified mutations in five Arabidopsis thaliana SR protein genes that result in altered phenotypes. Two (scl28-1 and srp31-1) result in embryonic lethal phenotypes, while three others (sc35-1, sr45-1, and srp30-1) result in viable and fertile plants with a range of phenotypes.

I have also found that mutations in individual SR protein genes can effect the ability of a specific sequence to act as an ESE and hence affect splicing efficiency. Because 16 of the 20 Arabidopsis thaliana SR proteins themselves are alternatively spliced, I have looked for cross regulation using RT-PCR analysis of isoform accumulation in alternatively spliced SR protein genes. I found that SR proteins do, in fact, regulate the alternative splicing of gene targets and do so in both a gene and a tissue specific manner.

In order to begin to fully understand the relationship between individual SR proteins it is essential to know when and where they are expressed throughout development. I have studied the expression pattern of 16 of the 20 SR proteins in the roots of wild-type plants as well as sc35-1, srp30-1, and sr45-1 mutants. I have identified both spatial and temporal expression patterns for these 16 proteins relative to specific tissues that compose the root.