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1.
PeerJ ; 7: e8064, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31824757

RESUMO

Phenylalanine ammonia lyase (PAL) plays an important role in the biosynthesis of secondary metabolites regulating plant growth response. To date, the evolutionary history of the PAL family in Rosaceae plants remains unclear. In this study, we identified 16 PAL homologous genes in five Rosaceae plants (Pyrus bretschneideri, Fragaria vesca, Prunus mume, Prunus persica, and Malus × domestica). We classified these PALs into three categories based on phylogenetic analysis, and all PALs were distributed on 13 chromosomes. We tracked gene duplication events and performed sliding window analysis. These results revealed the evolution of PALs in five Rosaceae plants. We predicted the promoter of the PbPALs by PLANT CARE online software, and found that the promoter region of both PbPAL1 and PbPAL3 have at least one AC element. The results of qRT-PCR analysis found that PbPAL1 and PbPAL2 were highly expressed in the stems and roots, while expression level of PbPAL3 was relatively low in different tissues. The expression of PbPAL1 and PbPAL2 increased firstly and then decreased at different developmental periods of pear fruit. Among them, the expression of PbPAL1 reached the highest level 55 days after flowering. Three PbPALs were induced by abiotic stress to varying degrees. We transfected PbPAL1 and PbPAL2 into Arabidopsis thaliana, which resulted in an increase in lignin content and thickening of the cell walls of intervascular fibres and xylem cells. In summary, this research laid a foundation for better understanding the molecular evolution of PALs in five Rosaceae plants. Furthermore, the present study revealed the role of PbPALs in lignin synthesis, and provided basic data for regulating lignin synthesis and stone cells development in pear plants.

2.
Front Genet ; 10: 632, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31333718

RESUMO

Stone cells are a characteristic trait of pear fruit, but the contents and sizes of stone cells negatively correlate with fruit texture and flavor. Secondary cell wall thickening and lignification have been established as key steps of stone cell development. KNOTTED-LIKE HOMEOBOX (KNOX) proteins play important roles in plant cell growth and development, including cell wall formation and lignification. Although the characteristics and biological functions of KNOX proteins have been investigated in other plants, this gene family has not been functionally characterized in pear. Eighteen PbKNOX genes were identified in the present study, and all of the identified family members contained the KNOX I and/or KNOX II domains. Based on the phylogenetic tree and chromosomal localization, the 18 PbKNOX genes were divided into five subfamilies [SHOOT MERISTEMLESS (STM)-like, BREVIPEDICELLUS (BP)-like, KNOTTED ARABIDOPSIS THALIANA 2/6 (KNAT2/6)-like, KNAT7-like, and KNAT3-5-like] and were distributed among 10 chromosomes. In addition, we identified 9, 11, and 11 KNOX genes in the genomes of grape, mei, and strawberry, respectively, and the greatest number of collinear KNOX gene pairs formed between pears and peaches. Analyses of the spatiotemporal expression patterns showed that the tissue specificity of PbKNOX gene expression was not very significant and that the level of the PbKNOX1 transcript showed an opposite trend to the levels of stone cells and lignin accumulation. Furthermore, PbKNOX1 has high sequence identity and similarity with Arabidopsis BP. Compared with wild-type Arabidopsis, plants overexpressing PbKNOX1 not only showed an approximately 19% decrease in the secondary cell wall thickness of vessel cells but also exhibited an approximately 13% reduction in the lignin content of inflorescence stems. Moreover, the expression of several genes involved in lignin biosynthesis was downregulated in transgenic lines. Based on our results, PbKNOX1/BP participates in cell wall-thickening and lignin biosynthesis and represses the transcription of key structural genes involved in lignin synthesis, providing genetic evidence for the roles of KNOX in cell wall thickening and lignin biosynthesis in pear.

3.
Cells ; 8(6)2019 05 29.
Artigo em Inglês | MEDLINE | ID: mdl-31146469

RESUMO

: The accumulation of lignin in fruit has a significant negative impact on the quality of fruit-producing trees, and in particular the lignin formation stimulates the development of stone cells in pear fruit. Reactive oxygen species (ROS) are essential for lignin polymerization. However, knowledge of the RBOH family, a key enzyme in ROS metabolism, remains unknown in most fruit trees. In this study, a total of 40 RBOHs were identified from five fruit-producing trees (Pyrus bretschneideri, Prunus persica, Citrus sinensis, Vitis vinifera, and Prunus mume), and 10 of these sequences came from Pyrus bretschneideri. Multiple sequence alignments revealed that all 10 PbRBOHs contained the NADPH_Ox domain and the six alpha-helical transmembrane domains (TM-I to TM-VI). Chromosome localization and interspecies phylogenetic tree analysis showed that 10 PbRBOHs irregularly distributed on 8 chromosomes and 3 PbRBOHs (PbRBOHA, PbRBOHB, and PbRBOHD) are closely related to known lignification-related RBOHs. Furthermore, hormone response pattern analysis showed that the transcription of PbRBOHs is regulated by SA, ABA and MeJA. Reverse transcription-quantitative real-time polymerase chain reaction (qRT-PCR) and transcriptome sequencing analysis showed that PbRBOHA, PbRBOHB, and PbRBOHD accumulated high transcript abundance in pear fruit, and the transcriptional trends of PbRBOHA and PbRBOHD was consistent with the change of stone cell content during fruit development. In addition, subcellular localization revealed that PbRBOHA and PbRBOHD are distributed on the plasma membrane. Combining the changes of apoplastic superoxide (O2.-) content and spatio-temporal expression analysis, these results indicate that PbRBOHA and PbRBOHD, which are candidate genes, may play an important role in ROS metabolism during the lignification of pear stone cells. This study not only provided insight into the molecular characteristics of the RBOH family in fruit-producing trees, but also lays the foundation for studying the role of ROS in plant lignification.


Assuntos
Simulação por Computador , Frutas/genética , Genes de Plantas , Lignina/metabolismo , Família Multigênica , NADPH Oxidases/genética , Pyrus/enzimologia , Pyrus/genética , Sequência de Aminoácidos , Cromossomos de Plantas/genética , Sequência Conservada/genética , Éxons/genética , Frutas/efeitos dos fármacos , Duplicação Gênica , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Tamanho do Genoma , Íntrons/genética , NADPH Oxidases/química , NADPH Oxidases/metabolismo , Motivos de Nucleotídeos/genética , Especificidade de Órgãos/efeitos dos fármacos , Especificidade de Órgãos/genética , Filogenia , Reguladores de Crescimento de Planta/farmacologia , Regiões Promotoras Genéticas/genética , Pyrus/efeitos dos fármacos , Sintenia/genética , Árvores/enzimologia , Árvores/genética
4.
Mol Biol Rep ; 46(2): 2153-2175, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30734172

RESUMO

Stone cells are a characteristic trait of pear fruits, and excessive stone cell formation has a significant negative impact on the texture and flavour of the pulp. Lignin is one of the main components of stone cells. Family-1 uridine diphosphate-glycosyltransferases (UGTs) are responsible for the glycosylation modification of monolignols. However, information remains limited regarding the relationship between UGTs and stone cell formation. To address this problem, we identified 139 UGTs from the pear genome, which were distributed in 15 phylogenetic groups (A-M, O, and P). We also performed a collinearity analysis of UGTs among four Rosaceae plants (pear, peach, mei, and strawberry). Phylogenetic analysis suggested that 13 PbUGTs might be related to the glycosylation of monolignols. Analysis of expression patterns demonstrated that most putative monolignol glycosylation-related PbUGTs not only showed high expression levels in flowers and buds but were also induced by exogenous ABA, SA, and MeJA. In addition, the transcript level of Pbr005014.1 (named PbUGT72AJ2) was consistent with the changing trend of lignin content in pear fruit, and the transcript level was also higher in 'Dangshan Su' pear with higher lignin and stone cell contents. Subcellular localization results showed that PbUGT72AJ2 was located mainly in the cytomembrane and cytoplasm. Based on our study, PbUGT72AJ2 is considered to be a monolignol glycosylation-related UGT. Our results provide an important source for the identification of UGTs and a foundation for the future understanding and manipulation of lignin metabolism and stone cell formation in pear fruit.


Assuntos
Glicosiltransferases/genética , Pyrus/genética , Sementes/genética , Frutas/genética , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica de Plantas/genética , Glicosiltransferases/metabolismo , Lignina/genética , Lignina/metabolismo , Família Multigênica , Filogenia , Proteínas de Plantas/genética , Pyrus/metabolismo , Transcriptoma/genética
5.
Gene ; 686: 237-249, 2019 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-30468911

RESUMO

Lignin is the main component of stone cells, which are a key factor in determining pear quality. Therefore, modification of lignin biosynthesis has important implications for regulating stone cell formation. LIMs are involved in plant development, stress response and metabolism. However, there is still a lack of knowledge about the pear LIM family and lignin-related LIMs. To address this problem, we identified 14 LIMs from the pear genome and named them. Phylogenomic and feature domain analysis showed that they were divided into CRP- and DA&DAR-LIM groups and five subclades. LIMs from the genomes of four rosids (Prunus mummer, Prunus persica, Fragaria vesca and Vitis vinifera) were also identified, and microsynteny analysis revealed the most orthologous gene pairs in the cross of pear/grape and pear/mei. The transcript levels of PbLIMs were significantly affected by SA, ABA and MeJA. Spatio-temporal expression analysis showed that PbLIMs of the δLIM2 subfamily were highly expressed in the flowers. Changes in the expression levels of PbWLIM1a and PbWLIM1b during fruit development was consistent with the changes in lignin content. Combining phylogenetic analyses, protein three-dimensional structure determination and sequence alignment analyses, these two genes were suggested as lignin-related PbLIMs. Subcellular localization results showed that PbWLIM1a and PbWLIM1b were located mainly in the chloroplast. This study lays the foundation for revealing the mechanism of LIM-mediated lignin metabolism to regulate stone cell formation.


Assuntos
Proteínas de Cloroplastos , Regulação da Expressão Gênica de Plantas/fisiologia , Lignina , Família Multigênica/fisiologia , Filogenia , Pyrus , Proteínas de Cloroplastos/biossíntese , Proteínas de Cloroplastos/genética , Cloroplastos/genética , Cloroplastos/metabolismo , Lignina/biossíntese , Lignina/genética , Pyrus/genética , Pyrus/metabolismo
6.
Org Biomol Chem ; 16(29): 5350-5358, 2018 07 25.
Artigo em Inglês | MEDLINE | ID: mdl-30004550

RESUMO

An efficient synthesis of (E)-cinnamaldehydes by a metal-free DDQ-mediated oxidative transformation of allylarenes was developed. The protocol provides a practical method to prepare diverse (E)-cinnamaldehydes with broad functional group tolerance in good to excellent yields, including easy access to natural products randainal and geranyloxy sinapyl aldehyde from plant extracts. Finally, the mechanism of a single-electron transfer process was proposed.

7.
Eur J Med Chem ; 139: 674-697, 2017 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-28850922

RESUMO

Recently, more and more concomitant EGFR mutations and ALK rearrangement are observed from the clinic, which still lacks effective single-agent therapy. Starting from ALK inhibitor 14 (TAE684), we have developed a highly potent EGFR/ALK dual kinase inhibitor compound 18 (CHMFL-ALK/EGFR-050), which potently inhibited EGFR L858R, del 19 and T790M mutants as well as EML4-ALK, R1275Q, L1196M, F1174L and C1156Y mutants biochemically. Compound 18 significantly inhibited the proliferation of EGFR mutant and EML4-ALK driven NSCLC cell lines. In the cellular context it strongly affected EGFR and ALK mediated signaling pathways, induced apoptosis and arrested cell cycle at G0/G1 phase. In the in vivo studies, 18 significantly suppressed the tumor growth in H1975 cell inoculated xenograft model (40 mg/kg/d, TGI: 99%) and H3122 cell inoculated xenograft model (40 mg/kg/d, TGI: 78%). Compound 18 might be a potential drug candidate for EGFR- or ALK-individual as well as concomitant EGFR/ALK NSCLC.


Assuntos
Acrilamidas/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Descoberta de Drogas , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Receptores ErbB/antagonistas & inibidores , Neoplasias Pulmonares/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Pirimidinas/farmacologia , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Acrilamidas/síntese química , Acrilamidas/química , Quinase do Linfoma Anaplásico , Animais , Apoptose/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/patologia , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Receptores ErbB/metabolismo , Feminino , Humanos , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Nus , Estrutura Molecular , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/patologia , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Pirimidinas/síntese química , Pirimidinas/química , Receptores Proteína Tirosina Quinases/metabolismo , Relação Estrutura-Atividade
8.
Org Lett ; 15(4): 788-91, 2013 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-23350934

RESUMO

An efficient palladium-catalyzed synthesis of 3-acylindoles using free (N-H) indoles and nitriles has been developed. The acylation reaction proceeded well under the Pd(OAc)(2)/2,2'-bipyridine system and with D-(+)-camphorsulfonic acid as the additive. A possible mechanism involving carbopalladation of nitriles and subsequent hydrolysis of ketimines is proposed.


Assuntos
Indóis/síntese química , Nitrilos/química , Paládio/química , Catálise , Indóis/química , Estrutura Molecular
9.
J Org Chem ; 77(21): 9504-9, 2012 Nov 02.
Artigo em Inglês | MEDLINE | ID: mdl-23025822

RESUMO

A palladium-catalyzed ortho-alkoxylation of anilides with both primary and secondary alcohols via ligand-directed C-H activation has been explored. This alkoxylation promoted by catalytic methanesulfonic acid proceeds well at room temperature in most cases and affords aryl alkyl ethers in moderate to good yields.


Assuntos
Álcoois/química , Anilidas/química , Éteres/química , Paládio/química , Catálise , Ligações de Hidrogênio , Estereoisomerismo
10.
Chem Commun (Camb) ; (46): 7236-8, 2009 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-19921041

RESUMO

A novel tandem protocol involving a Heck reaction process for the synthesis of alpha,beta-unsaturated aldehydes has been developed. In the presence of Pd(OAc), PPh3, NaOAc, TBAB and air, N-allylbenzenamines underwent the reaction with various aryl halides to afford the corresponding alpha,beta-unsaturated aldehydes selectively in moderate to good yields. The protocol can also be used as a valuable route for the deallylation of arylamines.


Assuntos
Aldeídos/síntese química , Aminas/química , Alcenos/química , Catálise , Hidrocarbonetos Aromáticos/química , Métodos , Oxigênio , Paládio
11.
J Org Chem ; 74(22): 8834-7, 2009 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-19848384

RESUMO

Palladium-catalyzed intramolecular hydroarylation of N-arylpropiolamides has been developed for the stereoselective synthesis of 3-(monosubstituted methylene)oxindoles. In the presence of Pd(OAc)(2) and dppf, a variety of N-arylpropiolamides successfully underwent the intramolecular hydroarylation reaction to afford the corresponding 3-(monosubstituted-methylene)oxindoles in moderate to excellent yields. It is noteworthy that the stereoselectivity of the reaction can be controlled by varying the reaction temperature.


Assuntos
Amidas/química , Indóis/síntese química , Compostos Organometálicos/química , Paládio/química , Termodinâmica , Catálise , Indóis/química , Estrutura Molecular , Oxindois , Estereoisomerismo
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