BELL1 interacts with CRABS CLAW and INNER NO OUTER to regulate ovule and seed development in pomegranate

Abstract

Pomegranate (Punica granatum) flowers are classified as bisexual flowers and functional male flowers. Functional male flowers have sterile pistils that show abnormal ovule development. In previous studies, we identified INNER NO OUTER (INO), CRABS CLAW (CRC) and BELL1 (BEL1), which were specifically expressed in bisexual and functional male flowers. However, the functions of ovule identity genes and the mechanism underlying ovule sterility in pomegranate remain unknown. Here, we found that the integument primordia formed and then ceased developing in the ovules of functional male flowers with a vertical diameter of 8.1–13.0 mm. Megaspore mother cells were observed in bisexual flowers when the vertical diameters of flowers were 10.1–13.0 mm, but not in functional male flowers. We analyzed the expression patterns of ovule-related genes in pomegranate ovule sterility and found that PgCRC mRNA was highly expressed at a critical stage of ovule development in bisexual flowers. Ectopic expression of PgCRC and PgINO was sufficient to increase seed number in transgenic lines. PgCRC partially complemented the Arabidopsis (Arabidopsis thaliana) crc mutant, and PgINO successfully rescued the seeds set in the Arabidopsis ino mutant. The results of Y2H assays, BiFC assays and genetic data analyses showed that PgCRC and PgINO directly interact with PgBEL1. Our results also showed that PgCRC and PgINO could not interact directly with MADS-box proteins and that PgBEL1 interacted with SEPALLATA (PgSEP) proteins. We report the function of PgCRC and PgINO in ovule and seed development and show that PgCRC and PgINO interact with PgBEL1. Thus, our results provide understanding of the genetic regulatory networks underlying ovule development in pomegranate.

Exploring MicroRNAs Associated with Pomegranate Pistil Development: An Identification and Analysis Study

“…Functional male flowers have sterile pistils that show abnormal ovule development. This result indicates that the vertical diameter of 8.1-13.0 mm is a critical stage for pomegranate ovule development [16]. In our study, pomegranate miRNAs of 'Taishanhong' bisexual flowers and functional male flowers were sequenced at the critical stages of pomegranate ovule development.…”

“…According to Zhao's study [16], pomegranate ovule development was divided into three stages (initial ovule development stage: 5.0-10.0 mm; critical ovule abortion stage: 10.1-13.0 mm; ovule maturity stage: 13.1-18.0 mm). The pistils of bisexual and functional male flowers with vertical bud diameters of 5.0-10.0 mm (I), 10.1-13.0 mm (II), and 13.1-18.0 mm (III) were used as the test materials for miRNAs sequencing.…”

“…The calyx, petal, and stamen tissues were removed, and only the female organs (ovary, style, and stigma) were retained for mixed-pool transcriptome and miRNA sequencing. The transcriptome data (PRJNA754480) were reported in our previous study [16]. Three biological replicates (18 samples in total) were obtained for each test sample.…”

A chromosome-level reference-quality genome ofPunica granatumL

“…The oil obtained from pomegranate seeds is known for its functional and beneficial health properties 5 . Apart from its commercial importance, pomegranate has become an attractive system for studying several valuable biological features, such as color formation in the fruit peel and aril 6 , phenotypic variations between soft-and hard-seeded varieties 7,8 , and the mechanism underlying ovule sterility [9][10][11] . Development of high-quality reference genome would be very useful for understanding the molecular mechanisms underlying these valuable biological features.…”

SlCRCa is a key D‐class gene controlling ovule fate determination in tomato

“…CRABS CLAW (CRC ) is an ancient member of the YABBY family in both gymnosperms and angiosperms (Zhao et al, 2023). Over two decades ago, Eshed et al (1999) found that sepal margins of Arabidopsis with 35S:AtCRC overexpression formed ectopic ovules and stigmas.…”