ARF6s Identification and Function Analysis Provide Insights Into Flower Development of Punica granatum L.

Yujie Zhao,Yuan Ren,Ming Yan,Yuying Wang,Xueqing Zhao,Zhaohe Yuan

doi:10.3389/fpls.2022.833747

Abstract

Based on the genome and small-RNA sequencing of pomegranate, miRNA167 and three target genes PgARF6 were identified in “Taishanhong” genome. Three PgARF6 genes and their corresponding protein sequences, expression patterns in pomegranate flower development and under exogenous hormones treatments were systematically analyzed in this paper. We found that PgARF6s are nuclear proteins with conserved structures. However, PgARF6s had different protein structures and expression profiles in pomegranate flower development. At the critical stages of pomegranate ovule sterility (8.1–14.0 mm), the expression levels of PgARF6s in bisexual flowers were lower than those in functional male flowers. Interestingly, PgARF6c expression level was significantly higher than PgARF6a and PgARF6b. Under the treatment of exogenous IBA and 6-BA, PgARF6s were down-regulated, and the expression of PgARF6c was significantly inhibited. PgmiR167a and PgmiR167d had the binding site on PgARF6 genes sequences, and PgARF6a has the directly targeted regulatory relationship with PgmiR167a in pomegranate. At the critical stage of ovule development (8.1–12.0 mm), exogenous IBA and 6-BA promoted the content of GA and ZR accumulation, inhibited BR accumulation. There was a strong correlation between the expression of PgARF6a and PgARF6b. Under exogenous hormone treatment, the content of ZR, BR, GA, and ABA were negatively correlated with the expressions of PgARF6 genes. However, JA was positively correlated with PgARF6a and PgARF6c under IBA treatment. Thus, our results provide new evidence for PgARF6 genes involving in ovule sterility in pomegranate flowers.

Highlights

Pomegranate, belonging to Lythraceae (Yuan et al, 2018), is rich in polyphenols such as punicalagin, ellagic acid, and other ellagitannin-based compounds (Johanningsmeier and Harris, 2011)
Pg006237.1, Pg013200.1, and Pg026615.1 were renamed as PgARF6a, PgARF6b, and PgARF6c for convenience in subsequent analysis, based on their potential orthologs proteins in Arabidopsis thaliana (Figure 1)
At the period of ovule primordia formed (5.1–8.0 mm, P2), the expression levels of PgARF6a, PgARF6b, and PgARF6c in bisexual flowers were higher compared with that in functional male flowers. These results showed that PgARF6s expression levels in bisexual flowers were significantly lower than that in functional male flowers at the critical stage of ovule sterility (8.1– 12.0 mm, P3–P4), indicating that PgARF6s was involved in ovule sterility developing into functional male flower

Summary

Introduction

Pomegranate, belonging to Lythraceae (Yuan et al, 2018), is rich in polyphenols such as punicalagin, ellagic acid, and other ellagitannin-based compounds (Johanningsmeier and Harris, 2011). Pomegranate trees produce bisexual flower and functional male flower. Bisexual flower has an apparent tubular shape, and can bear a fruit as time advancement, whereas functional male. ARF6 Function in Pomegranate Flower Development flower is bell-shaped which usually drops and fails to set fruit. Studies have revealed that female sterility in functional male flowers is due to the abnormal ovule development failing to generate egg cells When the vertical diameter of bisexual flowers is 5.1–10.0 mm, inner integument primordia are being form. When bisexual flowers gain their vertical diameter to 13.1–15.0 mm, the integument continues to enlarge, whereas the inner integument of functional male flowers is stagnated and the ovules display wilting It can be concluded that it is an important stage for pomegranate ovule development when the vertical diameter of a flower bud is 5.1–15.0 mm

Methods

Results

Conclusion