Integrated microRNA and mRNA expression profiling reveals a complex network regulating pomegranate (Punica granatum L.) seed hardness

Abstract

The breeding of new soft-seeded pomegranate cultivars provides new products for the market and increases farmers’ incomes, yet the genetic architecture mediating seed hardness is largely unknown. Here, the seed hardness and hundred-seed weights of 26 cultivars were determined in 2 successive years. We conducted miRNA and mRNA sequencing to analyse the seeds of two varieties of Punica granatum: soft-seeded Tunisia and hard-seeded Sanbai, at 60 and 120 d after flowering. Seed hardness was strongly positively correlated with hundred-seed weight. We detected 25 and 12 differentially expressed miRNA–mRNA pairs with negative regulatory relationships between the two genotypes at 60 and 120 d after flowering, respectively. These miRNA–mRNA pairs mainly regulated seed hardness by altering cell wall structure. Transcription factors including NAC1, WRKY and MYC, which are involved in seed hardness, were targeted by differentially expressed mdm-miR164e and mdm-miR172b. Thus, seed hardness is the result of a complex biological process regulated by a miRNA–mRNA network in pomegranate. These results will help us understand the complexity of seed hardness and help to elucidate the miRNA-mediated molecular mechanisms that contribute to seed hardness in pomegranate.

The pomegranate (Punica granatum L.) draft genome dissects genetic divergence between soft‐ and hard‐seeded cultivars

“…These genes participate in several biological pathways, including cell division, cell wall biogenesis, signal transduction, transcriptional regulation, product transport and metabolism (Figure S8). The pathways related to cell division and the cell wall structure were proved to control pomegranate seed hardness (Luo et al ., ; Niu et al ., ). We also identified many transcription factor families, such as MYB (16 genes), WRKY (five genes), AP2‐like (16 genes), MYC (two genes) and NAC (nine genes), that contained different proportions of SNPs and InDels (Figure b).…”

“…We also identified many transcription factor families, such as MYB (16 genes), WRKY (five genes), AP2‐like (16 genes), MYC (two genes) and NAC (nine genes), that contained different proportions of SNPs and InDels (Figure b). Most of these transcription factors have been reported to play roles in regulating the seed hardness of pomegranates (Luo et al ., ; Xue et al ., ) and hawthorns (Dai et al ., ). In the comparison between ‘Tunisia’ and ‘Sanbai’, we detected a SNP (T‐C) at the 166‐bp position of the NAC ( PgL0137670 ) transcription factor coding sequence.…”

“…fruit setting to ripening stages) (Xue et al, 2017;Zarei et al, 2016). Quantitative proteomics and microRNA sequencing results have suggested that genes altering the cell wall structure contribute to the differences in seed hardness between soft-and hard-seeded pomegranates (Luo et al, 2018;Niu et al, 2018). Four seed hardness-related quantitative trait loci, potentially explaining 15% to 30% of the observed phenotypic variation, were identified by Harel-Beja et al (2015).…”

Small RNA and mRNA Sequencing Reveal the Roles of microRNAs Involved in Pomegranate Female Sterility

“…Several conserved and novel miRNAs in pomegranate have been identified in previous studies through high-throughput small RNA sequencing [28][29][30]. Compared with the miRNAs expressed in fruit [30], a total of 28 known miRNAs identified in this study were conserved in pistil and fruit, while 67 known miRNAs were newly identified in pistil. Of the 28 conserved miRNAs, pg-miR160a-5p, pg-miR166a-3p, pg-miR167h, pg-miR168a, pg-miR159a, pg-miR171b, and pg-miR319a-3p were abundantly enriched in both pistil and fruit, which indicated their pleiotropy in pistil and fruit development.…”

“…The study of small RNAs in pomegranate has been reported previously [28][29][30]. However, the involvement of small RNAs in pomegranate FMFs' ovule development cessation has not yet been determined.…”