Apple (Malus × domestica) trees are vulnerable to freezing temperatures. However, there has been only limited success in developing cold-hardy cultivars. This lack of progress is due at least partly ...to lack of understanding of the molecular mechanisms of freezing tolerance in apple.
In this study, we evaluated the potential roles for two R2R3 MYB transcription factors (TFs), MYB88 and the paralogous FLP (MYB124), in cold stress in apple and Arabidopsis. We found that MYB88 and MYB124 positively regulate freezing tolerance and cold-responsive gene expression in both apple and Arabidopsis.
Chromatin-Immunoprecipitation-qPCR and electrophoretic mobility shift assays showed that MdMYB88/MdMYB124 act as direct regulators of the COLD SHOCK DOMAIN PROTEIN 3 (MdCSP3) and CIRCADIAN CLOCK ASSOCIATED 1 (MdCCA1) genes. Dual luciferase reporter assay indicated that MdCCA1 but not MdCSP3 activated the expression of MdCBF3 under cold stress. Moreover, MdMYB88 and MdMYB124 promoted anthocyanin accumulation and H2O2 detoxification in response to cold.
Taken together, our results suggest that MdMYB88 and MdMYB124 positively regulate cold hardiness and cold-responsive gene expression under cold stress by C-REPEAT BINDING FACTOR (CBF)-dependent and CBF-independent pathways.
Minimum miscible pressure (MMP) is an essential design parameter of gas flooding for enhanced oil recovery (EOR) applications. Researchers have developed a number of methods for MMP computations, ...including the analytical methods, the slim-tube simulation method, and the multiple-mixing-cell (MMC) method. Among these methods, the MMC method is widely accepted for its simplicity, robustness, and moderate computational cost An important version of the MMC method is the Jaubert et al. method which has a much lower computational cost than the slim-tube simulation method. However, the original Jaubert et al. method suffers several drawbacks. One notable drawback is that it cannot be applied to the scenario where the oil-gas MMP is lower than the saturation pressure of the crude oil. In this work, we present a modified MMC method that is more versatile and robust than the original version. Our method can handle the scenario where the oil-gas MMP is lower than the saturation pressure of the crude oil. Besides, we propose a modified MMC model that can reduce the computational cost of MMP estimations. This modified model, together with a newly proposed pressure search algorithm, increases the MMP estimation accuracy of the modified method. We demonstrate the good performance of the modified MMC method by testing it in multiple case studies. A good agreement is obtained between the MMPs calculated by the modified method and the tie-line-based ones from the literature.
Aroma affects the sensory quality of fruit and, consequently, consumer satisfaction. Melatonin (MT) is a plant growth regulator used to delay senescence in postharvest fruit during storage; however, ...its effect on aroma of pear fruit remains unclear. In this study, we assessed the effects of 0.1 mmol L
MT on volatiles and associated gene expression in the fruit of pear cultivars 'Korla' (
Rehd) and 'Abbé Fetel' (
L.). MT mainly affected the production of C
aromatic substances in the two varieties. In 'Korla', MT inhibited expression of
, and reduced hydroperoxide lyase (HPL) activity and content of hexanal and (
)-hex-2-enal. In contrast, MT inhibited activity of lipoxygenase (LOX), reduced expression of
and
, promoted
gene expression, increased alcohol acyltransferase (AAT) activity, and increased propyl acetate, and hexyl acetate content in 'Abbé Fetel' that similarly led to the reduction in content of hexanal and (
)-hex-2-enal. Content of esters in 'Abbé Fetel' pear increased with increasing postharvest storage period. Although mechanisms differed between the two varieties, effects on aroma volatiles mediated by MT were driven by expression of genes encoding LOX, HPL, and AAT enzymes.
As a class of natural antioxidants in plants, fruit flavonol metabolites are beneficial to human health. However, the regulatory networks for flavonol biosynthesis in most fruits are largely unknown. ...Previously, we reported a spontaneous pear bud sport 'Red Zaosu' (Pyrus bretschneideri Rehd.) with a high flavonoid content in its fruit. The identification of the flavonol biosynthetic regulatory network in this mutant pear fruit is crucial for elucidating the flavonol biosynthetic mechanism in fruit.
Here, we demonstrated the PbMYB12b positively regulated flavonols biosynthesis in 'Red Zaosu' fruit. Initially, we investigated the accumulation patterns of four major quercetin glycosides and two major isorhamnetin glycosides in the fruit of 'Red Zaosu' and its wild-type 'Zaosu'. A PRODUCTION OF FLAVONOL GLYCOSIDES (PFG)-type MYB transcription factor PbMYB12b was also screened for because of its correlation with flavonol accumulation in pear fruit. The biofunction of PbMYB12b was verified by transient overexpression and RNAi assays in pear fruit and young leaves. Overexpression of PbMYB12b enhanced the biosynthesis of quercetin glycosides and isorhamnetin glycosides by positively regulating a general flavonoids biosynthesis gene PbCHSb and a flavonol biosynthesis gene PbFLS. This finding was also supported by dual-luciferase transient expression assay and transient β-glucuronidase (GUS) reporter assay.
Our study indicated that PbMYB12b positively regulated flavonol biosynthesis, including four major quercetin glycosides and two major isorhamnetin glycosides, by promoting the expression of PbCHSb and PbFLS in pear fruit.
Parthenocarpy, the production of seedless fruit without fertilization, has a variety of valuable qualities, especially for self-incompatible species, such as pear. To explore whether melatonin (MT) ...induces parthenocarpy, we used 'Starkrimson' pear as a material for morphological observations. According to our results, exogenous MT promoted the expansion and division of the mesocarp cells in a manner similar to hand pollination. However, the seeds of exogenous MT-treated fruit were undeveloped and aborted later in the fruit-setting stage. To further investigate how MT induced parthenocarpy, we studied changes of related hormones in the ovaries and found that MT significantly increased the contents of the gibberellins (GAs) GA
and GA
. Thus, paclobutrazol (PAC), a GA-biosynthesis inhibitor, was used to study the relationship between GAs and MT. In addition, spraying MT after treatment with PAC did not increase GA content nor lead to parthenocarpy. Through a transcriptome analysis, we discovered that MT can cause significant upregulation of
and downregulation of
. However, no significant difference was observed in
compared with the control after PAC and MT applications. Thus, MT induces parthenocarpy by promoting GA biosynthesis along with cell division and mesocarp expansion in pear.
Subgroup 4 R2R3 MYBs play vital roles in the regulation of anthocyanin biosynthesis. However, there is limited knowledge regarding the functions of MYB repressors in pear (
). Here, PbMYB120 was ...identified as a potential regulator of anthocyanin biosynthesis. A phylogenetic analysis revealed that PbMYB120 was clustered into the FaMYB1-like clade of the subgroup 4 R2R3 MYBs.
was expressed higher in red peels than in green peels in five pear cultivars.
expression was positively correlated with anthocyanin accumulation. However, the transient overexpression of
led to the inhibition of anthocyanin accumulation and
expression. Promoter binding and activation assays indicated that PbMYB120 binds to the promoter of
and represses the promoter's activity. Thus, the inhibition of anthocyanin accumulation by PbMYB120 may be correlated with the repression of
. Furthermore, during anthocyanin induction, the expression levels of anthocyanin activators and
were upregulated. This study demonstrated that
was highly expressed in pear tissues having higher anthocyanin accumulations but acted as a repressor in the regulation of anthocyanin accumulation. PbMYB120 may work coordinately with anthocyanin activators and serve as a balancer of anthocyanin accumulation.
The N-terminal of Myc-like basic helix-loop-helix transcription factors (bHLH TFs) contains an interaction domain, namely the MYB-interacting region (MIR), which interacts with the R2R3-MYB proteins ...to regulate genes involved in the anthocyanin biosynthetic pathway. However, the functions of MIR-domain bHLHs in this pathway are not fully understood. In this study, PbbHLH2 containing the MIR-domain was identified and its function investigated. The overexpression of
in "Zaosu" pear peel increased the anthocyanin content and the expression levels of late biosynthetic genes. Bimolecular fluorescence complementation showed that PbbHLH2 interacted with R2R3-MYB TFs PbMYB9, 10, and 10b in onion epidermal cells and confirmed that MIR-domain plays important roles in the interaction between the MIR-domain bHLH and R2R3-MYB TFs. Moreover, PbbHLH2 bound and activated the dihydroflavonol reductase promoter in yeast one-hybrid (Y1H) and dual-luciferase assays. Taken together these results suggested that the MIR domain of PbbHLH2 regulated anthocyanin biosynthesis in pear fruit peel.
Seedless fruit is a feature appreciated by consumers. The ovule abortion process is highly orchestrated and controlled by numerous environmental and endogenous signals. However, the mechanisms ...underlying ovule abortion in pear remain obscure. Here, we found that gibberellins (GAs) have diverse functions during ovules development between seedless pear ‘1913’ and seeded pear, and that GA4+7 activates a potential programmed cell death process in ‘1913’ ovules. After hormone analyses, strong correlations were determined among jasmonic acid (JA), ethylene and salicylic acid (SA) in seedless and seeded cultivars, and GA4+7 treatments altered the hormone accumulation levels in ovules, resulting in significant correlations between GA and both JA and ethylene. Additionally, SA contributed to ovule abortion in ‘1913’. Exogenously supplying JA, SA or the ethylene precursor 1-aminocyclopropane-1-carboxylic acid promoted ‘Bartlett’ seed death. The regulatory mechanism in which ethylene controls ovule death has been demonstrated; therefore, JA’s role in regulating ‘1913’ ovule abortion was investigated. A further study identified that the JA signaling receptor MYC2 bound the SENESCENCE-ASSOCIATED 39 promoter and triggered its expression to regulate ovule abortion. Thus, we established ovule abortion-related relationships between GA and the hormones JA, ethylene and SA, and we determined their synergistic functions in regulating ovule death.
Anthocyanin biosynthesis exhibits a rhythmic oscillation pattern in some plants. To investigate the correlation between the oscillatory regulatory network and anthocyanin biosynthesis in pear, the ...anthocyanin accumulation and the expression patterns of anthocyanin late biosynthetic genes (
s) were investigated in fruit skin of 'Red Zaosu' (Pyrus bretschneideri Rehd.). The anthocyanin accumulated mainly during the night over three continuous days in the fruit skin, and the
s' expression patterns in 'Red Zaosu' fruit skin were oscillatory. However, the expression levels of typical anthocyanin-related transcription factors did not follow this pattern. Here, we found that the expression patterns of four
s (
s), members of a class of atypical anthocyanin-regulated
s, were consistent with those of
s in 'Red Zaosu' fruit skin over three continuous days. Additionally, transient expression assays indicated that the four
s promoted anthocyanin biosynthesis by regulating the expression of the anthocyanin biosynthetic genes encoding dihydroflavonol-4-reductase (DFR) and anthocyanidin synthase (ANS) in red pear fruit skin, which was verified using a dual-luciferase reporter assay. Moreover, a yeast one-hybrid assay indicated that
,
and
directly bound to
and
promoters. Thus,
s promote anthocyanin accumulation at night by up-regulating the expression levels of
and
in 'Red Zaosu' fruit skin.
Decrease in anthocyanin content results in the loss of red color in leaves, petals and receptacles during development. The content of anthocyanin was affected by the biosynthesis and degradation of ...anthocyanin. Compared with the known detailed mechanism of anthocyanin biosynthesis, the degradation mechanism is not fully investigated. It is vital to study the degradation mechanism of anthocyanin in pear for promoting the accumulation of anthocyanin and inhibiting the red fading in pear.
Here, we reported that laccase encoded by PbLAC4-like was associated with anthocyanin degradation in pear. The expression pattern of PbLAC4-like was negatively correlated with the content of anthocyanin during the color fading process of pear leaves, petals and receptacles. Phylogenetic analysis and sequence alignment revealed that PbLAC4-like played a vital role in anthocyanin degradation. Thus, the degradation of anthocyanin induced by PbLAC4-like was further verified by transient assays and prokaryotic expression. More than 80% of anthocyanin compounds were degraded by transiently over-expressed PbLAC4-like in pear fruitlet peel. The activity of crude enzyme to degrade anthocyanin in leaves at different stages was basically consistent with the expression of PbLAC4-like. The anthocyanin degradation ability of prokaryotic induced PbLAC4-like protein was also verified by enzyme activity assay. Besides, we also identified PbMYB26 as a positive regulator of PbLAC4-like. Yeast one-hybrid and dual luciferase assay results showed that PbMYB26 activated PbLAC4-like expression by directly binding to the PbLAC4-like promoter.
Taken together, the PbLAC4-like activated by PbMYB26, was involved in the degradation of anthocyanin, resulting in the redness fading in different pear tissues.
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