Diplonema spp. possess spliced leader RNA genes similar to the kinetoplastida

The phylogenetic placement of the genus Diplonema in relation to fellow phy lum members Euglena and Trypanosoma has been uncertain. The spliced leader RNA gene, present in the euglenids and kinetoplastids in distinct forms. wa s a potential target for resolving this question. The first indication supp orting a closer relationship to the kinetoplastids was the recognition of p otential spliced leader RNA exon sequences in the genomic DNA of two Diplon ema isolates. Examination of total cell RNA revealed transcripts in the ant icipated size range at approximately 120 and 130 nt. Specific PCR amplifica tion of a spliced leader RNA gene repeat was performed. The hallmark featur es of the kinetoplastid-type spliced leader RNA, specifically the 39-nt exo n, splice-donor site. Sm-binding sire and poly-T tract and the potential to form the requisite stem-loop structures, were found. Diplonema spp. are di fferent from the kinetoplastids by virtue of C residues at positions 4 and 18 in the exon. While the intergenic spacer regions varied in size, each co ntained the complete sequence or remnants of a 5S ribosomal RNA gene. Posse ssion of a functional spliced leader RNA gene of the kinetoplastid variety in Diplonema supports a closer evolutionary relationship with this group th an with the euglenids.