Group I introns and associated homing endonuclease genes reveals a clinal structure for Porphyra spiralis var. amplifolia (Bangiales, Rhodophyta) along the Eastern coast of South America

Abstract

Background: Group I introns are found in the nuclear small subunit ribosomal RNA gene (SSU rDNA) of some species of the genus Porphyra (Bangiales, Rhodophyta). Size polymorphisms in group I introns has been interpreted as the result of the degeneration of homing endonuclease genes (HEG) inserted in peripheral loops of intron paired elements. In this study, intron size polymorphisms were characterized for different Porphyra spiralis var. amplifolia (PSA) populations on the Southern Brazilian coast, and were used to infer genetic relationships and genetic structure of these PSA populations, in addition to cox2-3 and rbcL-S regions. Introns of different sizes were tested qualitatively for in vitro self-splicing.

Results: Five intron size polymorphisms within 17 haplotypes were obtained from 80 individuals representing eight localities along the distribution of PSA in the Eastern coast of South America. In order to infer genetic structure and genetic relationships of PSA, these polymorphisms and haplotypes were used as markers for pairwise Fst analyses, Mantel's test and median joining network. The five cox2-3 haplotypes and the unique rbcL-S haplotype were used as markers for summary statistics, neutrality tests Tajima's D and Fu's Fs and for median joining network analyses. An event of demographic expansion from a population with low effective number, followed by a pattern of isolation by distance was obtained for PSA populations with the three analyses. In vitro experiments have shown that introns of different lengths were able to self-splice from pre-RNA transcripts.

Conclusion: The findings indicated that degenerated HEGs are reminiscent of the presence of a full-length and functional HEG, once fixed for PSA populations. The cline of HEG degeneration determined the pattern of isolation by distance. Analyses with the other markers indicated an event of demographic expansion from a population with low effective number. The different degrees of degeneration of the HEG do not refrain intron self-splicing. To our knowledge, this was the first study to address intraspecific evolutionary history of a nuclear group I intron; to use nuclear, mitochondrial and chloroplast DNA for population level analyses of Porphyra; and intron size polymorphism as a marker for population genetics.

Figure 1

Map of South America highlighting Porphyra spiralis var. amplifolia (PSA) collection sites. 1- PSA-A; 2- PSA-L; 3- PSA-R*; 4- PSA-I; 5- PSA-S; 6- PSA-T; 7- PSA-G; 8- PSA-C; 9- PSA-D*; 10- PSA-B*; 11- PSA-V. Additional information on collections sites are presented on Table 1. * Data obtained from Oliveira and Ragan [2].

Figure 2

Translation of homing endonuclease gene open reading frames of the five size polymorphisms. The His-Cys box motif is underlined and zinc binding sites and active sites found in I-PpoI endonuclease are indicated by line above amino acids. Stop codon is indicated by asterisk. End of sequence without stop codon is indicated by +. H – haplotype.

Figure 3

Median-joining networks: A – Introns + homing endonuclease gene (HEG); B – and introns without HEG; C – Cox2-3 region. Circles indicate sequence types and circle sizes are proportional to haplotype frequency. Lines indicate substitutions (not on scale). MV- median vector, H- haplotype. Haplotypes were not plotted on intron without HEG network.