Shutian Tao

Evaluation of the volatile profile of 33 Pyrus ussuriensis cultivars by HS-SPME with GC–MS

Evaluation of the volatile compounds in fruit provides useful information for plant breeding for improved fruit aroma. In this study, headspace solid-phase microextraction (HS-SPME) combined with gas chromatography-mass spectrometry (GC-MS) was used to assess the volatile profile of 33 cultivars of the Chinese pear Pyrus ussuriensis. In all, 108 volatile compounds were identified and there were significant differences in the composition and concentration of volatiles among cultivars. On the basis of principal components analysis (PCA), the cultivars could be divided into four groups: Group 1 contained Reli, Jinxiang, Hongbalixiang, Baibalixiang and Fuwuxiang, cultivars with a high concentration of esters and a low concentration of hydrocarbons. Group 2 contained Qiuxiang, Fuanjianba, Longxiang, Guanhongxiao, Shanli24 and Wuxiangli, cultivars with high concentrations of hydrocarbons and low concentrations of esters. Group 3 contained Shatangli and Manyuanxiang, cultivars with high concentrations of aldehydes. Group 4 contained the other 25 cultivars.

Reciprocal regulation of Ca2+‐activated outward K+ channels of Pyrus pyrifolia pollen by heme and carbon monoxide

• The regulation of plant potassium (K+) channels has been extensively studied in various systems. However, the mechanism of their regulation in the pollen tube is unclear. • In this study, the effects of heme and carbon monoxide (CO) on the outward K+ (K+(out)) channel in pear (Pyrus pyrifolia) pollen tube protoplasts were characterized using a patch-clamp technique. • Heme (1 μM) decreased the probability of K+(out) channel opening without affecting the unitary conductance, but this inhibition disappeared when heme was co-applied with 10 μM intracellular free Ca²+. Conversely, exposure to heme in the presence of NADPH increased channel activity. However, with tin protoporphyrin IX treatment, which inhibits hemeoxygenase activity, the inhibition of the K+(out) channel by heme occurred even in the presence of NADPH. CO, a product of heme catabolism by hemeoxygenase, activates the K+(out) channel in pollen tube protoplasts in a dose-dependent manner. The current induced by CO was inhibited by the K+ channel inhibitor tetraethylammonium. • These data indicate a role of heme and CO in reciprocal regulation of the K+(out) channel in pear pollen tubes.

Evolution of the aroma volatiles of pear fruits supplemented with fatty acid metabolic precursors.

To examine the biochemical metabolism of aroma volatiles derived from fatty acids, pear fruits were incubated in vitro with metabolic precursors of these compounds. Aroma volatiles, especially esters, were significantly increased, both qualitatively and quantitatively, in pear fruits fed on fatty acid metabolic precursors. Cultivars having different flavor characteristics had distinctly different aroma volatile metabolisms. More esters were formed in fruity-flavored “Nanguoli” fruits than in green-flavored “Dangshansuli” fruits fed on the same quantities of linoleic acid and linolenic acid. Hexanal and hexanol were more efficient metabolic intermediates for volatile synthesis than linoleic acid and linolenic acid. Hexyl esters were the predominant esters produced by pear fruits fed on hexanol, and their contents in “Dangshansuli” fruits were higher than in “Nanguoli” fruits. Hexyl esters and hexanoate esters were the primary esters produced in pear fruits fed on hexanal, however the content of hexyl ester in “Dangshansuli” was approximately three times that in “Nanguoli”. The higher contents of hexyl esters in “Dangshansuli” may have resulted from a higher level of hexanol derived from hexanal. In conclusion, the synthesis of aroma volatiles was largely dependent on the metabolic precursors presented.

Heteroallelic diploid pollen led to self-compatibility in tetraploid cultivar ‘Sha 01’ (Pyrus sinkiangensis Yü)

Pear cultivar ‘Sha 01’ is a sport from ‘Kuerlexiangli’. Field pollination data revealed that ‘Sha 01’ displayed self-compatibility (SC), whereas ‘Kuerlexiangli’ showed self-incompatibility upon self-pollination. Reciprocal pollinations between the two varieties showed that 76% of ‘Kuerlexiangli’ flowers pollinated with ‘Sha 01’ pollen set fruit, but only 7% of ‘Sha 01’ flowers set fruit when pollinated with ‘Kuerlexiangli’ pollen. The pollen performance was monitored with fluorescence microscopy, and it was observed that ‘Sha 01’ accepted self-pollen but rejected ‘Kuerlexiangli’ pollen, whereas ‘Kuerlexiangli’ rejected self-pollen but accepted ‘Sha 01’ pollen. Taken together, ‘Sha 01’ showed SC as a pollen-part mutant. Molecular S-genotyping of ‘Sha 01’, its selfed progeny, and wild-type ‘Kuerlexiangli’ showed that all contained S22-RNase and S28-RNase alleles, but showed no nucleotide sequence difference or changes in transcription. After flow cytometry and chromosome number analyses, ‘Sha 01’ was found to be a tetraploid (2n = 68) and ‘Kuerlexiangli’ a diploid (2n = 34). Thus, the S-genotype for ‘Sha 01’ was S22S22S28S28 and for ‘Kuerlexiangli’ was S22S28. Ploidy level of ‘Sha 01’ selfed progeny were also determined from leaves using flow cytometry; all seedlings were tetraploids. The S-genotyping for the progeny was identified by S-RNase gene relative optical density for both alleles. Closer inspection of 52 progeny showed that they could be classified into three types S22S22S28S28: S22S22S22S28: S22S28S28S28 with the distribution 28:10:14 ≈ 4:1:1; no individual homozygous for either allele was found. Therefore, it could be concluded that only heteroallelic diploid pollen S22S28 could achieve ‘Sha 01’ fertilization through a so-called competitive interaction.

Identification of differentially expressed genes in a spontaneous mutant of ‘Nanguoli’ pear (Pyrus ussuriensis Maxim) with large fruit

Summary The selection of mutants is one of the most important steps in horticultural breeding. Fruit size is an important breeding objective in pear (Pyrus spp.) because of its economic importance. The pear cultivar ‘Da Nanguoli’ (Pyrus ussuriensis Maxim) produces larger fruit than ‘Nanguoli’, from which ‘Da Nanguoli’ was a bud mutation. The molecular basis for this bud mutation remains unknown. In the present study, ploidy analysis showed that ‘Da Nanguoli’ did not exhibit any alteration in the overall ploidy level. Seventy-six transcript-derived fragments (TDFs) of genes that were differentially expressed between the two cultivars were sequenced using cDNA-AFLP technology. BLAST-X analysis showed that 26 (34.2%) of the TDFs had high sequence homology with known proteins in the non-redundant National Center for Biotechnology Information (NCBI) protein sequence database (i.e., E-values < –3.00). In total, 17 TDFs were chosen and their patterns of expression revealed using cDNA-AFLP and confirmed using quantitative real-time polymerase chain reaction (qRT-PCR). In addition, 98 TDFs, representing candidate genes, were studied in more detail to determine their functions in order to dissect the complex molecular mechanisms involved in the fruit-size mutation.

Genome-wide Annotation and Comparative Analysis of Long Terminal Repeat Retrotransposons between Pear Species of P. bretschneideri and P. Communis

Recent sequencing of the Oriental pear (P. bretschneideri Rehd.) genome and the availability of the draft genome sequence of Occidental pear (P. communis L.), has provided a good opportunity to characterize the abundance, distribution, timing, and evolution of long terminal repeat retrotransposons (LTR-RTs) in these two important fruit plants. Here, a total of 7247 LTR-RTs, which can be classified into 148 families, have been identified in the assembled Oriental pear genome. Unlike in other plant genomes, approximately 90% of these elements were found to be randomly distributed along the pear chromosomes. Further analysis revealed that the amplification timeframe of elements varies dramatically in different families, super-families and lineages, and the Copia-like elements have highest activity in the recent 0.5 million years (Mys). The data also showed that two genomes evolved with similar evolutionary rates after their split from the common ancestor ~0.77–1.66 million years ago (Mya). Overall, the data provided here will be a valuable resource for further investigating the impact of transposable elements on gene structure, expression, and epigenetic modification in the pear genomes.

Mitochondrial dysfunction mediated by cytoplasmic acidification results in pollen tube growth cessation in Pyrus pyrifolia

The length of pollen tubes grown in synthetic media is normally shorter than those grown in vivo. However, the mechanism(s) underlying the cessation of pollen tube growth under culture conditions remain(s) largely unknown. Here, we report a previously unknown correlation between vacuolar function and the cells ability to sustain mitochondrial functions in pear pollen tubes. The pear pollen tubes in vitro grew slowly after 15 hours post-cultured (HPC) and nearly ceased growth at 18 HPC. There was increased malondialdehyde content and membrane ion leakage at 15 HPC compared with 12 HPC. Furthermore, cytoplasmic acidification mainly mediated by decreased vacuolar H(+)-ATPase [V-ATPase, Enzyme Commission (EC) 3.6.1.3] activity was observed in pollen tubes after 15 HPC, and this further resulted in mitochondrial dysfunction, including mitochondrial structure disruption, mitochondrial membrane potential collapse and decreases in both oxygen consumption and ATP production. Our findings suggest that vacuoles and mitochondria intimately linked in regulating pollen tube elongation.

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.

The unique evolutionary pattern of the Hydroxyproline-rich glycoproteins superfamily in Chinese white pear (Pyrus bretschneideri)

BackgroundThe hydroxyproline-rich glycoprotein (HRGP) superfamily, comprising three families (arabinogalactan-proteins, AGPs; extensins, EXTs; proline-rich proteins, PRPs), is a class of proline-rich proteins that exhibit high diversity and are involved in many aspects of plant biology.ResultsIn this study, 838 HRGPs were identified from Chinese white pear (Pyrus bretschneideri) by searching for biased amino acid composition and conserved motifs. 405 HRGPs were derived from whole genome duplication (WGD) events which is suggested to be the major force of driving HRGPs expansion and the recent WGD event shared by apple and pear generated most duplicated HRGPs in pear. This duplication event drived the structural variation of the HRGPs encoding hydroxyproline (Hyp)-rich motifs. The rate of HRGPs evolution mainly impacted the Hyp-rich motifs even in chimeric HRGPs. During the evolution of 53 PRPs that are also typified by 7-deoxyloganetin glucosyltransferase-like genes, the duplication from PRP to non-PRP was indirectly modified by positive selection. These results suggested that the rate of HRGP evolution mainly influenced the Hyp-rich motifs even in chimeric HRGPs. The expression divergence of HRGPs was higher than that of other commonly duplicated genes. In pear pistil, 601 HRGPs exhibited expression, while in pear pollen, 285 HRGPs were expressed. The qPCR results revealed that Pbr036330.1 and Pbr010506.1 showed different expression profile in self-incompatibility of pear pistil.ConclusionsThe researches indicated that WGD events was the main duplication type during the evolution of HRGPs, and the highly variable Hyp-motifs might be accountable for the expansion, evolution and expression divergence of HRGPs and that this divergence may be responsible for the gain of new functions in plants.

The mining and evolutionary investigation of AP2/ERF genes in pear (Pyrus)

BackgroundIn plants, ERF genes participate in a variety of regulatory pathways, such as plant growth and biotic and/or abiotic stress responses. Although the genome of Chinese white pear (‘Dangshansuli’) has been released, knowledge regarding the ERF family in pear, such as gene functions, evolutionary history and expression patterns, remains limited.ResultsIn our study, a total of 155 members of ERF families were identified in pear (Pyrus bretschneideri). The Ka and Ks values suggested that whole-genome duplication (WGD) and dispersed duplication have effectively contributed to the expansion of the pear ERF family. Gene structure and phylogeny analysis divided the PbrERF family into 12 groups, and their gene functions were predicted by comparative analysis. qRT-PCR was carried out to verify the relative expression levels of 7 genes in group III using wild and cultivated pear fruits at three key developmental stages. Wild samples had higher expression of these genes than cultivated samples, especially at the enlarged fruit stage. The transcriptome data of pear seedlings subjected to dehydration treatment further revealed that 4 of the 7 genes responded to drought conditions.ConclusionThe AP2/ERF gene family is greatly expanded in pear. Comparative analysis revealed the probability of ERF genes performing functional roles in multiple pathways. Expression analysis at different stages of pear fruit development in wild and cultivated samples indicated that genes in group III might be involved in abiotic and/or biotic stresses. Further transcriptome data on seedlings subjected to drought treatment verified the potential role of ERF genes in stress response. These results will provide a valuable reference for understanding the function and evolution of the ERF family in higher plants.