The rpl23 gene and pseudogene are hotspots of illegitimate recombination in barley chloroplast mutator seedlings

Abstract

Previously, through a TILLING (Targeting Induced Local Lesions in Genomes) approach applied on barley chloroplast mutator ( cpm ) seedlings a high frequency of polymorphisms in the rpl23 gene was detected. All the polymorphisms corresponded to five differences already known to exist in nature between the rpl23 gene located in the inverted repeats (IRs) and the rpl23 pseudogene located in the large single copy region (LSC). In this investigation, polymorphisms in the rpl23 gene were verified and besides, a similar situation was found for the pseudogene in cpm seedlings. On the other hand, no polymorphisms were found in any of those loci in 40 wild type barley seedlings. Those facts and the independent occurrence of polymorphisms in the gene and pseudogene in individual seedlings suggest that the detected polymorphisms initially arose from gene conversion between gene and pseudogene. Moreover, an additional recombination process involving small recombinant segments seems to occur between the two gene copies as a consequence of their location in the IRs. These and previous results support the hypothesis that the CPM protein is a component of the plastome mismatch repair (MMR) system, whose failure of the anti-recombination activity results in increased illegitimate recombination between the rpl23 gene and pseudogene.

Four large indels detected by cpTILLING in barley chloroplast mutator seedlings

“…The presence of short perfect or imperfect direct repeats in the borders of the four large indels identified in the plastome of cpm seedlings is in agreement with the hypothesis that they originated as a consequence of increased rates of illegitimate recombination between these short repeats. Previously, illegitimate recombination was also postulated as the cause of the high frequencies of polymorphisms observed in the rpl23 gene and pseudogene in cpm seedlings (Lencina et al, 2019). These observations together with the increased rates of substitutions and small indels in microsatellites previously observed in cpm seedlings (Landau et al, 2016) Figure 1.…”

“…The vast majority of those polymorphisms were due to substitutions and small indels (insertions/deletions) in microsatellites. However, a peculiar spectrum that included several combinations of five single nucleotide polymorphisms was observed in the rpl23 gene, which was later explained as arisen not from mutations, but from increased illegitimate recombination between the rpl23 gene and its pseudogene (Lencina et al, 2019). Besides the polymorphisms mentioned above, four large indels were found, which are the subject of analysis of the present work.…”

“…imperfect direct repeats were identified at both ends of the four large indels. This result suggests that, as in the case of the polymorphisms observed in the rpl23 gene and its pseudogene (Lencina et al, 2019), they also may be due to increased illegitimate recombination caused by malfunction of the Cpm gene product. Furthermore, we investigated the consequences of protein expression of two of the large indels located in coding regions.…”

Variation in Plastome Sizes Accompanied by Evolutionary History in Monogenomic Triticeae (Poaceae: Triticeae)

“…In the grass plastome, the rpl23 gene is originally located in the IR region and encodes the functional ribosomal protein L23. Present studies have indicated that rpl23 has been non-reciprocally translocated to a region downstream from rbcL (Bowman et al, 1988;Ogihara et al, 1988;He et al, 2017;Lencina et al, 2019). A non-reciprocal translocation of the rpl23 gene occurred during the differentiation of Poaceae from its unknown ancestor (Katayama and Ogihara, 1996).…”

Induced mutations in crop plants at Instituto de Genética "Ewald A. Favret": mutants of scientific and/or agronomic interest: Induced mutations in crop plants at Instituto de Genética "Ewald A. Favret"

“…At least 61 different mutational events were detected; most of them were transitions and mono nucleotide indels [29]. This strategy also allowed the discovery of a high frequency of seedlings carrying different combinations of five polymorphisms, which already exist in nature, between the rpl23 gene and its pseudogene, which are postulated to originate in an increased rate of illegitimate recombination between these two loci [30]. All the results suggest that the failure of the cpm gene corresponds to a malfunction of a mismatch repair (MMR) gene involved in maintaining the stability of the plastome and that the cpm mutant constitutes an exceptional experimental material to obtain genetic variability in the otherwise highly conserved plastid genome.…”