Trans-splicing and cis-splicing in the colourless Euglenoid, Entosiphon sulcatum

Area of Science:

• Molecular Biology
• Eukaryotic Gene Expression
• Euglenoid Genomics

Background:

• Entosiphon sulcatum is a colorless euglenoid, potentially diverging before the evolution of photosynthetic euglenoids like Euglena gracilis.
• Understanding gene expression mechanisms in early-diverging euglenoids provides insights into eukaryotic evolution.

Purpose of the Study:

• To investigate the RNA splicing mechanisms in Entosiphon sulcatum.
• To identify genes involved in RNA processing, including spliced leader-sequence RNA (SL-RNA) and small nuclear RNAs (snRNAs).
• To compare splicing mechanisms between E. sulcatum and Euglena gracilis.

Main Methods:

• Isolation and characterization of genes encoding SL-RNA, 5S rRNA, and snRNAs (U1, U2, U5) from E. sulcatum.
• Analysis of pre-mRNA maturation through trans-splicing.
• Identification of intron-exon boundaries in the beta-tubulin gene.

Main Results:

• E. sulcatum utilizes trans-splicing for pre-mRNA maturation, confirmed by the presence of SL-RNA.
• A repeated DNA fragment in E. sulcatum encodes SL-RNA, 5S rRNA, and snRNAs U1, U2, and U5.
• The presence of snRNA U1 suggests the existence of classical cis-splicing.
• The beta-tubulin gene in E. sulcatum exhibits GU-AG intron borders, characteristic of spliceosome-matured introns.

Conclusions:

• E. sulcatum employs both trans-splicing and cis-splicing mechanisms for gene expression.
• The findings suggest that both splicing pathways were present early in euglenoid evolution.
• The unique intron features in E. sulcatum offer further avenues for studying spliceosome evolution.