Due to self-incompatibility (SI) prevents self-fertilization, natural or artificial cross-pollination has been conducted in many orchards to stabilize fruit yield. However, it is still puzzled which routes of self S-RNase arresting pollen tube growth. Herein, 17 COBRA genes were isolated from pear genome. Of these genes, the pollen-specifically expressed PbCOB.A.1 and PbCOB.A.2 positively mediates pollen tube growth. The promoters of PbCOB.A.1 and/or PbCOB.A.2 were bound and activated by PbABF.E.2 (an ABRE-binding factor) and PbC2H2.K16.2 (a C2H2-type zinc finger protein). Notably, the expressions of PbCOB.A.1, PbCOB.A.2, and PbC2H2.K16.2 were repressed by self S-RNase, suggesting that self S-RNase reduces the expression of PbCOB.A.1 and PbCOB.A.2 by decreasing the expression of their upstream factors, such as PbC2H2.K16.2, to arrest pollen tube growth. PbCOB.A.1 or PbCOB.A.2 accelerates the growth of pollen tubes treated by self S-RNase, but can hardly affect level of reactive oxygen species and deploymerization of actin cytoskeleton in pollen tubes and cannot physically interact with any reported proteins involved in SI. These results indicate that PbCOB.A.1 and PbCOB.A.2 may not relieve S-RNase toxicity in incompatible pollen tube. The information provides a new route to elucidate the arresting pollen tube growth during SI reaction.

Pear fruit senescence under high- and low-temperature conditions has been reported to be mediated by microRNAs. Long non-coding RNAs (lncRNAs), which can function as competing endogenous RNAs that interact with microRNAs, may also be involved in temperature-affected fruit senescence. Based on the transcriptome and microRNA sequencings, in this study, 3330 lncRNAs were isolated from Pyrus pyrifolia fruit. Of these lncRNAs, 2060 and 537 were responsive to high- and low-temperature conditions, respectively. Of these differentially expressed lncRNAs, 82 and 24 correlated to the mRNAs involved in fruit senescence under high- and low-temperature conditions, respectively. Moreover, three lncRNAs were predicted to be competing endogenous RNAs (ceRNAs) that interact with the microRNAs involved in fruit senescence, while one and two ceRNAs were involved in fruit senescence under high- and low-temperature conditions, respectively. A dual-luciferase assay showed that the interaction of an lncRNA with a microRNA disrupts the action of the microRNA on the expression of its target mRNA(s). Furthermore, four alternative splicing-derived lncRNAs interacted with miR172i homologies (Novel_88 and Novel_69) to relieve the repressed expression of their target and produce an miR172i precursor. Correlation analysis of microRNA expression suggested that Novel_69 is likely involved in the cleavage of the pre-miR172i hairpin to generate mature miR172i. Taken together, lncRNAs are involved in pear fruit senescence under high- or low-temperature conditions through ceRNAs and the production of microRNA.

The Chinese pear (Pyrus spp.) exhibits typical gametophytic self-incompatibility (GSI), which inhibits self-crossing and promotes out-crossing, similar to other fruit species in the Rosaceae family. Thus, S-compatible cultivars are required in pear orchards to ensure successful pollination and stable yields. In this study, 84 native Chinese pear accessions were genotyped by allele-specific PCR using one pair allele consensus primers and 29 pairs of S-allele-specific primers that were designed in this study. After cloning and sequencing the PCR products, the S-genotypes of all 84 pear accessions, including wild and cultivated accessions, were determined. The reported 34 S-alleles and a novel S-allele were isolated from these pear accessions. These S-alleles were expressed specifically in the style. Sequence analysis identified that six pear cultivars originated in China shared the same S-RNases with P. communis (Pc). These findings supported the hypothesis that oriental and occidental Pyrus spp. may share the same pool of alleles at the S-locus. A novel S-RNase was isolated in 'Putiandouli', 'Daguoshanli', 'Yunhongli 1′, and 'Dianli' and deposited as S₆₇-RNase under accession number MT773568. Furthermore, the deduced amino acid sequences exhibited high similarity (99.56%) to S₃₂-RNase in Malus. The high similarity between S-RNase in Pyrus and Malus indicated that the existence of S-RNase could have predated speciation between Pyrus and Malus. Furthermore, S-allele information was rearranged in Asian and European pears to provide information for selecting the best pollinator for widely cultivated pear cultivars in China. This information is useful for pear production, cross-breeding, and understanding the mechanism of the self-incompatibility reaction.

With-No-Lysine kinases (WNK) have been reported to be associated with plant growth regulation in Arabidopsis, soybean, and rice, but little is known of their roles in fruits. In this study, a total of 114 WNK genes were identified from 8 fruit trees species, and these WNK genes belonged to 2 classes (Ⅰ and Ⅱ) that respectively contain 4 (A, B, C, D) and 2 groups (E, F). The WNK genes had variable exon-intron structures and were randomly distributed among most chromosomes of each genus. The expression levels of six of the 18 WNK genes in peach were almost undetectable in fruits, suggesting that they may not be associated with fruit development and ripening. Of the other 12 WNK genes, PpWNK.B1 exhibited stronger stability of expression levels than the best reference genes reported previously in fruits from two peach cultivars. These results suggested that PpWNK.B1 could be a reliable reference gene for gene expression studies of peach fruit. Moreover, qRT-PCR assays yielded evidence that when using single reference gene as internal control, mistakes were more possible, indicating that the use of three reference genes is necessary for gene expression studies. Based on the gene expression profiles validated by qRT-PCR, PpWNK.A1 is predicted to likely be involved in fruit ripening, while PpWNK.A2 and PpWNK.E3.1 are likely associated with early fruit development. The present study represented the WNK genes in fruits, and it will be a valuable resource in continuing investigation of gene regulation network during fruit development and ripening.