Intermediate introns in nuclear genes of euglenids – are they a distinct type?

Abstract

BackgroundNuclear genes of euglenids contain two major types of introns: conventional spliceosomal and nonconventional introns. The latter are characterized by variable non-canonical borders, RNA secondary structure that brings intron ends together, and an unknown mechanism of removal. Some researchers also distinguish intermediate introns, which combine features of both types. They form a stable RNA secondary structure and are classified into two subtypes depending on whether they contain one (intermediate/nonconventional subtype) or both (conventional/intermediate subtype) canonical spliceosomal borders. However, it has been also postulated that most introns classified as intermediate could simply be special cases of conventional or nonconventional introns.ResultsSequences of tubB, hsp90 and gapC genes from six strains of Euglena agilis were obtained. They contain four, six, and two or three introns, respectively (the third intron in the gapC gene is unique for just one strain). Conventional introns were present at three positions: two in the tubB gene (at one position conventional/intermediate introns were also found) and one in the gapC gene. Nonconventional introns are present at ten positions: two in the tubB gene (at one position intermediate/nonconventional introns were also found), six in hsp90 (at four positions intermediate/nonconventional introns were also found), and two in the gapC gene.ConclusionsSequence and RNA secondary structure analyses of nonconventional introns confirmed that their most strongly conserved elements are base pairing nucleotides at positions +4, +5 and +6/ -8, −7 and −6 (in most introns CAG/CTG nucleotides were observed). It was also confirmed that the presence of the 5' GT/C end in intermediate/nonconventional introns is not the result of kinship with conventional introns, but is due to evolutionary pressure to preserve the purine at the 5' end. However, an example of a nonconventional intron with GC-AG ends was shown, suggesting the possibility of intron type conversion between nonconventional and conventional. Furthermore, an analysis of conventional introns revealed that the ability to form a stable RNA secondary structure by some introns is probably not a result of their relationship with nonconventional introns. It was also shown that acquisition of new nonconventional introns is an ongoing process and can be observed at the level of a single species. In the recently acquired intron in the gapC gene an extended direct repeats at the intron-exon junctions are present, suggesting that double-strand break repair process could be the source of new nonconventional introns.Electronic supplementary materialThe online version of this article (doi:10.1186/s12862-016-0620-5) contains supplementary material, which is available to authorized users.

Euglena gracilis Genome and Transcriptome: Organelles, Nuclear Genome Assembly Strategies and Initial Features

“…7.5) (for details on Euglenoid introns see Milanowski et al 2014, 2016, andMcWaters andRussell, this volume). Overall gene lengths in the transcriptome are similar to those previously reported (Schantz and Schantz 1989;Jackson et al 2006;Levasseur et al 1994;Milanowski et al 2014Milanowski et al , 2016Canaday et al 2001), but are considerably shorter in the genome owing to truncation and fragmentation.…”

Euglena Transcript Processing

“…Position of insertion of the group II intron in domain VI of the group III intron is indicated by the small box. Relative positions of branch point A nucleotides for both introns are indicated nucleotide in non-conventional introns has also been observed (Milanowski et al 2016) that perhaps affects splicing efficiency, thus explaining the frequent observation of non-conventional introns starting with the sequence 5′GT/C (i.e. spliceosomal-like) but also containing all other typical features of non-canonical introns.…”

“…These could potentially be excised using components of both the spliceosome and trans-acting non-conventional intron splicing factors. Milanowski et al have recently examined the conservation of intron position and class in conserved nuclear genes in different Euglena species to shed light on such questions (Milanowski et al 2014(Milanowski et al , 2016. These studies have further refined the limited conserved sequence and structural features of non-conventional introns (as described above) and revealed that non-conventional intron gain/loss appears to occur much more frequently than observed for euglenid spliceosomal introns.…”

“…The non-conventional introns are defined as containing extensive secondary structural potential via base-pairing of intron 5′ and 3′ end proximal sequences, but little overall intron sequence conservation (Tessier et al 1991;Canaday et al 2001;Russell et al 2005;Milanowski et al 2014Milanowski et al , 2016Schwartzbach 1992, 1994) (Fig. 8.1b).…”

“…Milanowski et al have noted that these features are reminiscent of MITE-like transposon elements (Milanowski et al 2014); therefore, if the non-conventional introns have been derived from such elements then perhaps trans-acting factors that associate with them may have been co-opted to be involved in the splicing mechanism. While there is no apparent conservation of extended sequence elements in these introns, intron sequence comparisons have revealed a preference for intron 5′ end proximal nucleotide positions +4 to +6 to be 'CAG' and the complementary sequence (CTG) starting 6 nucleotides from the intron 3′ end (Milanowski et al 2014(Milanowski et al , 2016 (Fig. 8.1b).…”