Structure and rearrangements of rRNA genes in chloroplast DNA in two strains of Euglena gracilis

Abstract

The organisation of the rRNA genes in the chloroplast genomes of two strains of Euglena gracilis were analyzed and compared. It was previously shown that the bacillaris strain contains three complete rrn (rRNA) operons (7) and that the Z-S strain contains one operon (21). Using heteroduplex analysis it was found that the bacillaris strain contains, apart from the three complete rrn operons, an extra 16S rRNA gene, an extra partial 23S rRNA gene sequence and an inverted duplication of a stretch within the 5S-16S spacer. In addition a short (<100 bp) inverted repeat sequence (13) which forms a stem/loop structure in single-stranded cpDNA was located between the 3'-end of the extra 16S rRNA gene and the partial 23 S rRNA sequence.The Z-S strain differs from the bacillaris strain by a deletion of two units of the complete rrn operons. The region upstream of the single complete rrn operon, including the inverted repeats, the partial 23S and the extra 16S rRNA sequences is identical with the bacillaris strain.The only non-homology found in heteroduplexes between the SalI fragments of B of the two strains is the deletion-insertion loop which represents the two rrn operons. A small deletion loop was found occasionally in hetero-and in homoduplexes of both strands in the region of variable size. Apart from the deletion/insertion of two rrn operons the two genomes appear to be colinear as can be seen from partial denaturation mapping. The organisation of the rRNA genes of the two strains is compared with those of the Z strain and the bacillaris-ATCC strain.

A chimeric transcript containing a 16 S rRNA and a potential mRNA in chloroplasts of Euglena gracilis

“…In one strain which apparently has only one rrn operon some restriction sites characteristic for this region are missing, but this does not exclude that the ORF406 is still present. In a similar analysis of the wild type strain Z-S, the s16 S rrn gene was not detected by gel hybridizations, but was later identified by electron microscopy [35,13]. The ORF406 seems to be present in most if not all strains of Euglena gracilis, although it could easily be deleted by homologous recombination between the s16 S rrn gene and one of the other 16 S rrn genes with a simi-345 lar frequency as the complete operons.…”

“…In four naturally occurring strains [7,9,12,24,35] and in 35 antibiotic-resistant mutants [19] which were obtained by UV irradiation the number of the repeated units of the rrn operons fluctuate frequently between one and five. The s16 S rrn gene and the ORF406 were identified in four wild type strains [10,12,13]. Restriction patterns of the antibiotic-resistant mutants indicate that these genes are also present in 34 of the 35 strains analysed.…”

“…The number of these units can fluctuate between one and five in different strains of the alga [7,9,12,24,35]. Supplementary 16 S and 5 S rRNA genes (sl6 S and s5 S rrn genes) and in some strains also a partial 23 S rRNA sequence were found upstream of the complete rrn operons [10,12,13]. In the strain Z of Euglenagracilis an inverted repeat of about 100 bp and an open reading frame with the coding capacity for 406 amino acids (ORF406) were found between the 3'-end of the s16 S and the first complete rrn operon [11,28,29].…”

Initiation of rrn transcription in chloroplasts of Euglena gracilis bacillaris

“…Chloroplast rRNA genes show marked structural similarities to prokaryotic rrn genes from which they are thought to have originated (Gray 1983;Palmer 1985), but relatively little is known about the control of their expression. The chloroplast genomes of the alga Euglena gracilis strain B (bacillaris) and Z each contain three complete and one incomplete rrn operons arranged in tandem (Koller and Delius 1982b;Koller et al 1984). Like three of the seven E. coIi rrn operons and the rrn operons of higher plants, the complete operons contain (in order of transcription) genes for 16S rRNA, tRNA ne, tRNA Ala, 23S rRNA, and 5S rRNA (Fig.…”

Comparison between the organization of nuclear ribosomal DNA unit of Euglena gracilis Z and var. Bacillaris

“…Depending on the strain Abbreviations kb: kilo base kpb: kilo base pair plasmids: pRH 59 and pRH 57 contain a Hind III-Hind III nuclear DNA fragment from W3BUL of 5.9 and 5.7 kbp respectively, pRB 48 and pRB 35 contain a Bam H1-Bam H1 nuclear DNA fragment from wildtype Z of 4.8 and 3.5 kbp respectively SDS: sodium dodecyl sulfate UV: ultra-violet analyzed, there are 1 (Z-S, 31), 2 (Y3BUD, 20), 3 (most of wild-type Z and bacillaris, 12,13) or 5 (ATCC 10616, 15) complete ribosomal operons. When searched, an extra copy of the 16 S rRNA gene has been found upstream from the first operon (12,16). While this work was in progress, Curtis and Rawson (4) described the organization of the nuclear rDNA unit from Euglena strain Z.…”