# Splice site diversity and abundance of noncanonical introns in diplonemids (Diplonemea, Euglenozoa)

## TL;DR

This study explores the unique intron structures in diplonemids, revealing noncanonical introns that challenge typical splicing patterns.

## Contribution

The research identifies noncanonical introns in diplonemids with diverse splice sites, expanding understanding of splicing mechanisms in euglenozoans.

## Key findings

- Diplonemids possess a nearly complete U2-type spliceosome but have mostly noncanonical introns.
- Noncanonical introns in Artemidia motanka and Namystynia karyoxenos lack GT-AG dinucleotides but can form extensive base-pairing.
- Splice sites show high variability among diplonemid species despite a conserved splicing apparatus.

## Abstract

Noncoding introns are a unifying feature of protein-coding genes in virtually all extant eukaryotes, with most lineages following the canonical intron structure. However, euglenozoans, unicellular flagellates that include free-living euglenids, human pathogenic kinetoplastids, and highly diverse and abundant marine diplonemids, are a notable exception. Euglenozoan genomes range from extremely intron-poor kinetoplastids to euglenid genomes containing both canonical and noncanonical introns. Here, we present a comprehensive analysis of splice sites and spliceosomal components in six species of understudied diplonemids. All diplonemids examined contain a nearly complete set of spliceosomal snRNP components, indicating the presence of a functional U2-type spliceosome. However, the majority of introns in the hemistasiid diplonemids Artemidia motanka and Namystynia karyoxenos are noncanonical and lack conserved GT-AG terminal dinucleotides typical for U2-type introns. These noncanonical introns are capable of extensive base-pairing, which brings intron ends into close proximity. Thus, while the splicing apparatus is conserved in diplonemids, the splice sites are highly variable among individual species.