Kaijie Qi

Map-based cloning of the pear gene MYB114 identifies an interaction with other transcription factors to coordinately regulate fruit anthocyanin biosynthesis

Red fruits are popular and widely accepted by consumers because of an enhanced appearance and enriched anthocyanins. The molecular mechanism of anthocyanin regulation in red-skinned pear (Pyrus) has been studied, and the genes encoding the biosynthetic steps and several transcription factors (TFs) have been characterized. In this study, a candidate R2R3 MYB TF, PyMYB114, was identified by linkage to the quantitative trait loci (QTL) for red skin color on linkage group 5 in a population of Chinese pear (Pyrus bretschneideri). The function of PyMYB114 was verified by transient transformation in tobacco (Nicotinana tabacum) leaves and strawberry (Fragaria) and pear fruits, resulting in the biosynthesis of anthocyanin. Suppression of PyMYB114 could inhibit anthocyanin biosynthesis in red-skinned pears. The ERF/AP2 TF PyERF3 was found to interact with PyMYB114 and its partner PybHLH3 to co-regulate anthocyanin biosynthesis, as shown by a dual luciferase reporter system and a yeast two-hybrid assay. In addition, the transcript abundance of PyMYB114 and PyMYB10 were correlated, and co-transformation of these two genes into tobacco and strawberry led to enhanced anthocyanin biosynthesis. This interaction network provides insight into the coloration of fruits and the interaction of different TFs to regulate anthocyanin biosynthesis.

Genome-wide identification and comparative analysis of the cation proton antiporters family in pear and four other Rosaceae species

The monovalent cation proton antiporters (CPAs) play essential roles in plant nutrition, development, and signal transduction by regulating ion and pH homeostasis of the cell. The CPAs of plants include the Na+/H+ exchanger, K+ efflux antiporter, and cation/H+ exchanger families. However, currently, little is known about the CPA genes in Rosaceae species. In this study, 220 CPA genes were identified from five Rosaceae species (Pyrus bretschneideri, Malus domestica, Prunus persica, Fragaria vesca, and Prunus mume), and 53 of which came from P. bretschneideri. Phylogenetic, structure, collinearity, and gene expression analyses were conducted on the entire CPA genes of pear. Gene expression data showed that 35 and 37 CPA genes were expressed in pear fruit and pollen tubes, respectively. The transcript analysis of some CPA genes under abiotic stress conditions revealed that CPAs may play an important role in pollen tubes growth. The results presented here will be useful in improving understanding of the complexity of the CPA gene family and will promote functional characterization in future studies.

Transcriptome analysis unravels an ethylene response factor involved in regulating fruit ripening in pear

Ethylene response factor (ERF) has been widely studied in regulating fruit ripening in tomato, apple, banana and kiwifruit, but little is known in pear. In this study 1-methylcyclopropene (1-MCP) treatment, an inhibitor of ethylene perception, was conducted at approximately 30 days before harvest to delay fruit ripening in a climacteric white pear cultivar Yali. Transcriptome libraries were constructed and sequenced in pre-ripening, ripening, and 1-MCP treated fruits. Data analysis showed that 73 candidate genes related to fruit ripening were induced by 1-MCP, among which two were positively related, namely 1-aminocyclopropane-1-carboxyla oxidase and an ERF gene (designated as ACO54 and ERF24). Transient transformations in pear fruit revealed that over-expression of ACO54 enhance transcription level of ERF24 and most ripening-related genes. Meanwhile, over-expression of ERF24 raises expression level of ACO54 and partially ripening-related genes. Moreover, dual-luciferase and yeast-one-hybrid assays unravel an interaction between ERF24 and the ACO54 promoter. Therefore, the ERF24 could directly regulate ACO54 expression by binding to its promoter. These results suggested that the first identified ERF24 is involved in regulating fruit ripening in Chinese white pear.

Diversification and independent domestication of Asian and European pears

BackgroundPear (Pyrus) is a globally grown fruit, with thousands of cultivars in five domesticated species and dozens of wild species. However, little is known about the evolutionary history of these pear species and what has contributed to the distinct phenotypic traits between Asian pears and European pears.ResultsWe report the genome resequencing of 113 pear accessions from worldwide collections, representing both cultivated and wild pear species. Based on 18,302,883 identified SNPs, we conduct phylogenetics, population structure, gene flow, and selective sweep analyses. Furthermore, we propose a model for the divergence, dissemination, and independent domestication of Asian and European pears in which pear, after originating in southwest China and then being disseminated throughout central Asia, has eventually spread to western Asia, and then on to Europe. We find evidence for rapid evolution and balancing selection for S-RNase genes that have contributed to the maintenance of self-incompatibility, thus promoting outcrossing and accounting for pear genome diversity across the Eurasian continent. In addition, separate selective sweep signatures between Asian pears and European pears, combined with co-localized QTLs and differentially expressed genes, underline distinct phenotypic fruit traits, including flesh texture, sugar, acidity, aroma, and stone cells.ConclusionsThis study provides further clarification of the evolutionary history of pear along with independent domestication of Asian and European pears. Furthermore, it provides substantive and valuable genomic resources that will significantly advance pear improvement and molecular breeding efforts.

A HD-ZIP II HOMEBOX transcription factor, PpHB.G7, mediates ethylene biosynthesis during fruit ripening in peach

Homeobox transcription factors belong to a superfamily that has been widely studied in plant growth and development, but little is known regarding their role in fruit development and ripening. Using a genome-wide expression analysis of homeobox (HB) genes and quantitative real-time PCR, a HD-ZIP II member, PpHB.G7, which presented higher levels of expression in ripening fruits than in developing fruits in all of the tested cultivars, was isolated from peach. Transient transformations showed that PpHB.G7 affects ethylene production and the expression of ethylene biosynthesis genes (PpACS1 and PpACO1). Both dual-luciferase and yeast one-hybrid assays confirmed that PpHB.G7 interacts with the promoters of PpACS1 and PpACO1. Thus, PpHB.G7 mediates ethylene biosynthesis by stimulating PpACS1 and PpACO1 activities. Furthermore, we also found that the other eight HB genes were differentially expressed in the developing fruits, with seven of these genes belonging to the HD-ZIP family. These results suggest that the HB genes in the HD-ZIP family play important roles in fruit development and ripening.

Genome-wide identification, evolution, and expression analysis of the KT/HAK/KUP family in pear

The K+ transporter/high-affinity K+/K+ uptake (KT/HAK/KUP) family, as one of the largest K+ transporter families in higher plants, plays an essential role in plant growth, mineral element absorption, salt stress tolerance, and other physiological processes. However, little is known about this family in pear (Pyrus). Here, we identified 20 K+ transporter genes in pear (P. bretschneideri) using genome-wide analysis. Their gene structure, chromosomal distribution, conserved motifs, phylogenetics, duplication events, and expression patterns were also examined. The results of phylogenetic analysis showed that PbrKT/HAK/KUP genes were clustered into three major groups (Groups I-III). Among the 20 PbrKT/HAK/KUP genes, 18 were mapped to nine chromosomes and two to scaffolds. Four WGD/segmental gene pairs were identified, indicating that WGD/segmental duplication may have contributed to the expansion of the KT/HAK/KUP family in pear. Among the four pairs of WGD/segmentally duplicated genes, both members of three pairs had been subjected to purifying selection, whereas the fourth pair had been subjected to positive selection. Furthermore, phenotypic experiments showed that the growth of pear seedlings was affected by potassium deficiency treatment. Expression patterns of 20 PbrKT/HAK/KUP genes in roots were further assayed with qRT-PCR. PbrHAK1 and PbrHAK12/16 were significantly expressed in response to K+ deficiency, suggesting that these genes are crucial for K+ uptake in pear, especially under the condition of K+ starvation. Our results provide a foundation for further study on the function of KT/HAK/KUP genes in pear.

Chemical composition and crystal morphology of epicuticular wax in mature fruits of 35 pear (Pyrus spp.) cultivars

An evaluation of fruit wax components will provide us with valuable information for pear breeding and enhancing fruit quality. Here, we dissected the epicuticular wax concentration, composition and structure of mature fruits from 35 pear cultivars belonging to five different species and hybrid interspecies. A total of 146 epicuticular wax compounds were detected, and the wax composition and concentration varied dramatically among species, with the highest level of 1.53 mg/cm2 in Pyrus communis and the lowest level of 0.62 mg/cm2 in Pyrus pyrifolia. Field emission scanning electron microscopy (FESEM) analysis showed amorphous structures of the epicuticular wax crystals of different pear cultivars. Cluster analysis revealed that the Pyrus bretschneideri cultivars were grouped much closer to Pyrus pyrifolia and Pyrus ussuriensis, and the Pyrus sinkiangensis cultivars were clustered into a distant group. Based on the principal component analysis (PCA), the cultivars could be divided into three groups and five groups according to seven main classes of epicuticular wax compounds and 146 wax compounds, respectively.

PbCOL8 is a clock-regulated flowering time repressor in pear

The floral transition is controlled by diverse endogenous and exogenous cues. In many species, COL (CONSTANS-like) genes integrate light and circadian clock signals to regulate flowering time. However, little is known about COLs in perennial woody plants. Here, we identified 15 PbCOLs in pear (Pyrus bretschneideri). PbCOLs were classified into three groups by phylogenetic tree analysis using protein sequences. Multiple sequence alignment analysis revealed conserved B-box and CCT (CO, CO-like, and TOC1) domains in all PbCOL members. This result suggested that PbCOLs might possess conserved functions as other species. Six PbCOLs were found to be regulated by both circadian clock and photoperiod. Here, we showed that PbCOL8, a member of group 2, suppressed the flowering signal integrators FT and SOC1 and could repress flowering time. These findings will contribute to elucidation of the mechanism of floral initiation in pear.

TGTT and AACA: two transcriptionally active LTR retrotransposon subfamilies with a specific LTR structure and horizontal transfer in four Rosaceae species

BackgroundLong terminal repeat retrotransposons (LTR-RTs) are major components of plant genomes. Common LTR-RTs contain the palindromic dinucleotide 5′-‘TG’–‘CA’-3′ motif at the ends. Thus, further analyses of non-canonical LTR-RTs with non-palindromic motifs will enhance our understanding of their structures and evolutionary history.ResultsHere, we report two new LTR-RT subfamilies (TGTT and AACA) with atypical dinucleotide ends of 5′-‘TG’–‘TT’-3′, and 5′-‘AA’–‘CA’-3′ in pear, apple, peach and mei. In total, 91 intact LTR-RTs were identified and classified into four TGTT and four AACA families. A structural annotation analysis showed that the four TGTT families, together with AACA1 and AACA2, belong to the Copia-like superfamily, whereas AACA3 and AACA4 appeared to be TRIM elements. The average amplification time frames for the eight families ranged from 0.05 to 2.32 million years. Phylogenetics coupled with sequence analyses revealed that the TGTT1 elements of peach were horizontally transferred from apple. In addition, 32 elements from two TGTT and three AACA families had detectable transcriptional activation, and a qRT-PCR analysis indicated that their expression levels varied dramatically in different species, organs and stress treatments.ConclusionsTwo novel LTR-RT subfamilies that terminated with non-palindromic dinucleotides at the ends of their LTRs were identified in four Rosaceae species, and a deep analysis showed their recent activity, horizontal transfer and varied transcriptional levels in different species, organs and stress treatments. This work enhances our understanding of the structural variation and evolutionary history of LTR-RTs in plants and also provides a valuable resource for future investigations of LTR-RTs having specific structures in other species.

Evolution, expression analysis, and functional verification of Catharanthus roseus RLK1-like kinase (CrRLK1L) family proteins in pear (Pyrus bretchneideri)

The Catharanthus roseus RLK1-like kinase (CrRLK1L) family is involved in multiple processes during plant growth. However, little is known about CrRLK1L in the wood of the pear fruit tree Pyrus bretchneideri. In this study, 26 CrRLK1L gene members were identified in pear and were grouped into six subfamilies according to phylogenetic analyses. Evolutionary analysis indicated that recent whole genome duplication (WGD) and dispersed gene duplications may contribute to the expansion of the CrRLK1L gene family in pear. Moreover, tissue-specific expression analyses suggested that CrRLK1Ls are involved in the development of various pear tissues. Subsequent qRT-PCR analyses indicated that CrRLK1Ls might play important roles in pollen tube growth. Finally, experiments with antisense oligonucleotides (ASO) demonstrated that PbrCrRLK1L26 have functions in pollen tube elongation and that PbrCrRLK1L3 regulates pollen tube rupture. These results will be useful for elaborating the biological roles of CrRLK1Ls in pear growth and development.