Association of T2/S-RNase With Self-Incompatibility of Japanese Citrus Accessions Examined by Transcriptomic, Phylogenetic, and Genetic Approaches

Abstract

Several citrus varieties show gametophytic self-incompatibility (GSI), which can contribute to seedless fruit production in several cultivars. This study investigated the genes regulating this trait through RNA-seq performed using styles collected from the flowers of Japanese citrus cultivars ‘Hyuganatsu,' ‘Tosabuntan,' ‘Hassaku,' ‘Banpeiyu,' and ‘Sweet Spring'. We screened the transcripts of putative T2 RNases, i.e., the protein family including all S-RNases from S-RNase-based GSI plants, and constructed a phylogenetic tree using the screened T2 RNases and S-RNases retrieved from citrus genome databases and a public database. Three major clusters (class I–III) were formed, among which, the class III cluster contained family specific subclusters formed by S-RNase and a citrus-specific cluster monophyletic to the S-RNase clusters. From the citrus class III cluster, six transcripts were consistent with the S haplotypes previously determined in Japanese citrus accessions, sharing characteristics such as isoelectric point, extracellular localization, molecular weight, intron number and position, and tissue-specific expression with S-RNases. One T2 RNase gene in self-incompatible Hyuganatsu was significantly down-regulated in the styles of a self-compatible mutant of Hyuganatsu in RNA-seq and qPCR analyses. In addition, the inheritance pattern of some T2 RNase genes was consistent with the pattern of the S haplotype in the progeny population of Hyuganatsu and Tosabuntan. As all results supported citrus self-incompatibility being based on S-RNase, we believe that six T2 RNase genes were S-RNases. The homology comparison between the six T2 RNases and S-RNases recently reported in Chinese citrus revealed that three out of six T2 RNases were identical to S-RNases from Chinese citrus. Thus, the other three T2 RNases were finally concluded to be novel citrus S-RNases involved in self-incompatibility.

Segregation Distortion for Male Parents in High Density Genetic Maps from Reciprocal Crosses between Two Self-Incompatible Cultivars Confirms a Gametophytic System for Self-Incompatibility in Citrus

“…They located the corresponding SI locus that also included F-box genes, at the beginning of the pseudo-chromosome 7 of the clementine reference genome. The involvement of T2/S-RNase in self-incompatibility of citrus was also proposed by Honsho et al [47] on the basis of transcriptomic, phylogenetic and genetic approaches. Both studies [46,47] analyzed the segregation of markers of the S-RNase genes on controlled progenies and found, for some of them, segregations in agreement with the gametophytic model for selfincompatibility.…”

“…The involvement of T2/S-RNase in self-incompatibility of citrus was also proposed by Honsho et al [47] on the basis of transcriptomic, phylogenetic and genetic approaches. Both studies [46,47] analyzed the segregation of markers of the S-RNase genes on controlled progenies and found, for some of them, segregations in agreement with the gametophytic model for selfincompatibility. However no genetic studies based on whole genome segregation analysis definitively validated the location of the SI locus and its unicity.…”

Self-incompatibility Related to Seedless Fruit Production in <i>Citrus</i> Plants

“…Fourteen self-incompatible S-RNase genes, S 1 -S 14 -RNase, and one self-compatible S-RNase gene, S m -RNase, coding a truncated protein owing to a frameshift with a gap insertion into a functional sequence, were reported by Liang et al (2020). In addition, the S-RNase genes of some Japanese citrus varieties, including self-incompatible 'Hyuganatsu', 'Nomabeni Hassaku', 'Tosa Buntan', and 'Banpeiyu' varieties, and probably self-compatible Sweet Spring variety, were identified, and the three novel S-RNase genes, S 15 -S 17 -RNase, were reported (Honsho et al, 2021). To date, 18 S-RNase genes, S 1 -S 17 -RNase and S m -RNase, have been identified (Table 2).…”

Breakdown of Self-Incompatibility in Citrus by Temperature Stress, Bud Pollination and Polyploidization

“…Later novel genes associated with self-pollen rejection in citrus have been identified [32]. Recent studies [33,34] indicate that GSI is based on S-RNase, which acts as a pistil S determinant by inhibiting pollen in an S-specific manner. Both studies analyzed the segregation of markers of S-RNase genes in controlled progenies and found segregations in agreement with this system for some.…”