Abstract

BackgroundGenome-wide computational analysis of alternative splicing (AS) in several flowering plants has revealed that pre-mRNAs from about 30% of genes undergo AS. Chlamydomonas, a simple unicellular green alga, is part of the lineage that includes land plants. However, it diverged from land plants about one billion years ago. Hence, it serves as a good model system to study alternative splicing in early photosynthetic eukaryotes, to obtain insights into the evolution of this process in plants, and to compare splicing in simple unicellular photosynthetic and non-photosynthetic eukaryotes. We performed a global analysis of alternative splicing in Chlamydomonas reinhardtii using its recently completed genome sequence and all available ESTs and cDNAs.ResultsOur analysis of AS using BLAT and a modified version of the Sircah tool revealed AS of 498 transcriptional units with 611 events, representing about 3% of the total number of genes. As in land plants, intron retention is the most prevalent form of AS. Retained introns and skipped exons tend to be shorter than their counterparts in constitutively spliced genes. The splice site signals in all types of AS events are weaker than those in constitutively spliced genes. Furthermore, in alternatively spliced genes, the prevalent splice form has a stronger splice site signal than the non-prevalent form. Analysis of constitutively spliced introns revealed an over-abundance of motifs with simple repetitive elements in comparison to introns involved in intron retention. In almost all cases, AS results in a truncated ORF, leading to a coding sequence that is around 50% shorter than the prevalent splice form. Using RT-PCR we verified AS of two genes and show that they produce more isoforms than indicated by EST data. All cDNA/EST alignments and splice graphs are provided in a website at http://combi.cs.colostate.edu/as/chlamy.ConclusionsThe extent of AS in Chlamydomonas that we observed is much smaller than observed in land plants, but is much higher than in simple unicellular heterotrophic eukaryotes. The percentage of different alternative splicing events is similar to flowering plants. Prevalence of constitutive and alternative splicing in Chlamydomonas, together with its simplicity, many available public resources, and well developed genetic and molecular tools for this organism make it an excellent model system to elucidate the mechanisms involved in regulated splicing in photosynthetic eukaryotes.

Highlights

  • Genome-wide computational analysis of alternative splicing (AS) in several flowering plants has revealed that pre-mRNAs from about 30% of genes undergo AS

  • Plant introns are rich in T or T/A, which is necessary for the recognition of splice sites and efficient splicing of pre-mRNAs [25,26]

  • During the last six years, the estimates of the extent of alternative splicing in flowering plants has increased from 5% to 30% [3] due to an increase in available EST and full-length cDNA sequences

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Summary

Introduction

Genome-wide computational analysis of alternative splicing (AS) in several flowering plants has revealed that pre-mRNAs from about 30% of genes undergo AS. We performed a global analysis of alternative splicing in Chlamydomonas reinhardtii using its recently completed genome sequence and all available ESTs and cDNAs. The coding regions (exons) of most eukaryotic genes are interrupted by non-coding sequences (introns). The availability of the complete genome sequences of many multicellular eukaryotic organisms and large sets of full-length cDNAs and ESTs has permitted comprehensive analysis of alternative splicing. Analysis of alternative splicing in flowering plants by aligning the available cDNAs/ESTs to genome sequences has shown that pre-mRNAs from ~ 30% of genes are alternatively spliced [10,11]. Recent completion of the Chlamydomonas genome and the availability of a fairly large number of ESTs [18,19,20] permit global analysis of post-transcriptional events including alternative splicing in a unicellular photosynthetic eukaryote

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