Pre-meiotic, 24-nt reproductive phasiRNAs are abundant in anthers of wheat and barley but not rice and maize

Bélanger, Sébastien; Pokhrel, Suresh; Czymmek, Kirk; Meyers, Blake C.

Two classes of pre-meiotic (21-nt) and meiotic (24-nt) phasiRNAs and their patterns of accumulation have been described in maize and rice anthers. Their precise function remains unclear, but some studies have shown that they support male fertility. The important role of phasiRNAs in anthers underpins our current study to their characterization in wheat and barley anthers. In this study, we staged anthers at every 0.2 mm of development for one wheat and two barley varieties. We isolated pre-meiotic (0.2 mm, 0.4 mm and 0.6 mm), meiotic (0.8 mm, 1.0 mm and 1.4 mm) and post-meiotic (1.8 mm) anthers for which we then investigated accumulation patterns of RNAs, including reproductive phasiRNAs. We annotated a total 12,821 and 2,897 PHAS loci in the wheat and barley genomes, respectively. When comparing the total number of PHAS loci in genomes of maize, rice, barley and wheat, we characterized an expansion of reproductive PHAS loci in the genomes of Poaceae subfamilies from Panicoideae to Oryzoideae and to Poideae. In addition to the two classes of pre-meiotic (21-nt) and meiotic (24-nt) phasiRNAs, previously described in maize and rice anthers, we described a group of 24-nt phasiRNAs that accumulate in pre-meiotic anthers. The absence of pre-meiotic 24-nt phasiRNAs in maize and rice suggests a divergence in grass species of the Poideae subfamily. Additionally, we performed a co-expression gene analysis describing the regulation of phasiRNA biogenesis in wheat and barley anthers. We highlight AGO9 and AGO6 as candidate binding partners of pre-meiotic and meiotic 24-nt phasiRNAs, respectively. One sentence summary In wheat and barley anthers, 24-nt reproductive phasiRNAs are abundant in both meiotic and pre-meiotic stages.