Echinococcus granulosus tropomyosin isoforms: from gene structure to expression analysis.
Tropomyosins (Trps) constitute a family of actin filament-binding proteins found in all eukaryotic cells. In muscle cells, they play a central role in contraction by regulating calcium-sensitive interaction of actin and myosin. In non-muscle cells, tropomyosins regulate actin filament organization and dynamics. Trps genes exhibit extensive cell type-specific isoform diversity generated by alternative splicing. Here, we report the characterization of tropomyosin gene transcribed sequences from the parasitic platyhelminth Echinococcus granulosus. Using RT-PCR approach we isolated three isoforms (egtrpA, egtrpB and egtrpC), which display significant homologies to know tropomyosins of different phylogenetic origin. The corresponding gene, egtrp (5656 bp), contains eight introns and nine exons. Southern blot hybridization studies showed that egtrp is present as single copy locus in E. granulosus. We demonstrated that egtrp expresses three different transcripts which differ in alternatively spliced exon 4 and intron VI. Interestingly, intron VI suffers intron retention and contains an internal stop codon in frame. Three major bands are also detected by Western blot analysis using a specific anti-rEgTrp antiserum. Immune-localization and in situ hybridization studies showed that egtrp transcription and translation is mostly localized at the protoscoleces suckers. This is the first report of alternative splicing in this parasite.
Alvite
Gabriela
G
Biochemistry Section, Cellular and Molecular Biology Department, Faculty of Sciences, University of the Republic, Montevideo, Uruguay. gabial@fcien.edu.uy
Esteves
Adriana
A
D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D015153:Blotting, Western; D017931:DNA Primers; D048209:Echinococcus granulosus / Q000235:genetics*; D005091:Exons; D015870:Gene Expression; D017403:In Situ Hybridization; D007438:Introns; D008969:Molecular Sequence Data; D020033:Protein Isoforms / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014335:Tropomyosin / Q000235:genetics*
D017931:DNA Primers; D020033:Protein Isoforms; D014335:Tropomyosin