The molecular basis for alternative splicing of the CABP 1 transcripts inDictyostelium discoideum

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Abstract
We have determined the nucleotide sequence of the CABP 1 gene from Dictyostelium discoideum. Together with previous data on cDNA sequences, we establish that alternative splicing of transcripts derived from this gene is responsible for the production of the two CABP 1 subunits. RNA blot analysis suggested that alternative splicing of the CABP 1 transcripts occurs during growth and throughout development. In addition, we have compiled the intron sequences of Dictyostelium pre-mRNAs and observed that the GUAAGU hexanucleotide at the 5′ splice site is highly conserved. The 5′ splice site of CABP 1 deviates from the consensus hexanucleotide in having a sequence of GUAAUA. To assess the role of the modified 5′ splice on differential splicing, we have constructed an actin-CABP1 fusion gene and transformed it into Dictyostelium cells. Analysis by immunoprecipitation with anti-CABP 1 antibody and amplification of specific cDNAs by polymerase chain reaction show that the transcripts generated by the fusion gene are alternatively spliced. When the 5′ splice site of the fusion gene is mutated to conform to the consensus sequence, the resulting transcripts are constitutively spliced. These observations suggest that changes in positions 5 and 6 of the donor splice site are involved in the alternative splicing of the CABP1 transcripts.