A Possible Role for Short Introns in the Acquisition of Stroma-Targeting Peptides in the Flagellate Euglena gracilis

Abstract

The chloroplasts of Euglena gracilis bounded by three membranes arose via secondary endosymbiosis of a green alga in a heterotrophic euglenozoan host. Many genes were transferred from symbiont to the host nucleus. A subset of Euglena nuclear genes of predominately symbiont, but also host, or other origin have obtained complex presequences required for chloroplast targeting. This study has revealed the presence of short introns (41–93 bp) either in the second half of presequence-encoding regions or shortly downstream of them in nine nucleus-encoded E. gracilis genes for chloroplast proteins (Eno29, GapA, PetA, PetF, PetJ, PsaF, PsbM, PsbO, and PsbW). In addition, the E. gracilis Pbgd gene contains two introns in the second half of presequence-encoding region and one at the border of presequence-mature peptide-encoding region. Ten of 12 introns present within presequence-encoding regions or shortly downstream of them identified in this study have typical eukaryotic GT/AG borders, are T-rich, 45–50 bp long, and pairwise sequence identities range from 27 to 61%. Thus single recombination events might have been mediated via these cis-spliced introns. A double crossing over between these cis-spliced introns and trans-spliced introns present in 5′-UTRs of Euglena nuclear genes is also likely to have occurred. Thus introns and exon-shuffling could have had an important role in the acquisition of chloroplast targeting signals in E. gracilis. The results are consistent with a late origin of photosynthetic euglenids.

Euglenoid flagellates: A multifaceted biotechnology platform

“…Euglena can be also used to purify polluted waste streams converting environmental pollutants into nontoxic biomass which can be used as a nutritional supplement in animal feed and aquaculture, and also as a viable source for biofuel production. Montandon et al (1986), Moreira et al (2004), Vacula et al (1999), Vesteg et al (2010), and Vlček et al (2011).…”

A small portion of plastid transcripts is polyadenylated in the flagellate Euglena gracilis

“…Nucleic acid isolation and purification, cDNA synthesis E. gracilis wild type strains Z and bacillaris (hereafter abbreviated as EgZ and EgB, respectively) were cultivated under constant illumination or in the dark in Hutner medium [28]. Genomic DNA was isolated as in the protocol described earlier using phenolchloroform method [29]. RNA extraction and mRNA isolation were carried out as described previously [29,30].…”

“…Genomic DNA was isolated as in the protocol described earlier using phenolchloroform method [29]. RNA extraction and mRNA isolation were carried out as described previously [29,30]. cDNA synthesis was performed with oligo(dT) and random hexanucleotide primers using ImProm-II Reverse Transcription System (Promega, Madison, WI, USA).…”

“…PCR was performed as described earlier [29,30] except the annealing temperature of 60°C instead of 58°C. In the first set of PCR reactions, the 26 bp-long SL sequence present at the 5 0 -end of E. gracilis nuclear mRNAs was used as a forward primer (SL).…”

Selective forces for the origin of spliceosomes

“…An intermediary evolutionary stage analogous to that leading to the origin prokaryotic operonal gene structure from eukaryotic-like one can be exemplified by trypanosomatids. In contrast to their relatives-free-living euglenids (Vesteg et al 2010), trypanosomatid parasites are almost completely devoid of cis-spliceosomal introns. Nevertheless, trypanosomatid premRNAs are polycistronic containing transcripts of genes separated by trans-introns and the individual mRNAs are resolved and capped by spliceosome-dependent SL-RNA-mediated trans-splicing (for review see Martínez-Calvillo et al 2010).…”