共查询到20条相似文献,搜索用时 31 毫秒
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Je Hyeong Jung Baskaran Kannan Hugo Dermawan Geoffrey W. Moxley Fredy Altpeter 《Plant molecular biology》2016,92(4-5):505-517
Sugarcane (Saccharum spp. hybrids) is a major feedstock for commercial bioethanol production. The recent integration of conversion technologies that utilize lignocellulosic sugarcane residues as well as sucrose from stem internodes has elevated bioethanol yields. RNAi suppression of lignin biosynthetic enzymes is a successful strategy to improve the saccharification of lignocellulosic biomass. 4-coumarate:coenzyme A ligase (4CL) is a key enzyme in the biosynthesis of phenylpropanoid metabolites, such as lignin and flavonoids. Identifying a major 4CL involved in lignin biosynthesis among multiple isoforms with functional divergence is key to manipulate lignin biosynthesis. In this study, two full length 4CL genes (Sh4CL1 and Sh4CL2) were isolated and characterized in sugarcane. Phylogenetic, expression and RNA interference (RNAi) analysis confirmed that Sh4CL1 is a major lignin biosynthetic gene. An intragenic precision breeding strategy may facilitate the regulatory approval of the genetically improved events and was used for RNAi suppression of Sh4CL1. Both, the RNAi inducing cassette and the expression cassette for the mutated ALS selection marker consisted entirely of DNA sequences from sugarcane or the sexually compatible species Sorghum bicolor. Field grown sugarcane with intragenic RNAi suppression of Sh4CL1 resulted in reduction of the total lignin content by up to 16.5?% along with altered monolignol ratios without reduction in biomass yield. Mature, field grown, intragenic sugarcane events displayed 52–76?% improved saccharification efficiency of lignocellulosic biomass compared to wild type (WT) controls. This demonstrates for the first time that an intragenic approach can add significant value to lignocellulosic feedstocks for biofuel and biochemical production. 相似文献
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Camilla Dornfeld Alexandra J. Weisberg Ritesh K C Natalia Dudareva John G. Jelesko Hiroshi A. Maeda 《The Plant cell》2014,26(7):3101-3114
The aromatic amino acid Phe is required for protein synthesis and serves as the
precursor of abundant phenylpropanoid plant natural products. While Phe is
synthesized from prephenate exclusively via a phenylpyruvate intermediate in model
microbes, the alternative pathway via arogenate is predominant in plant Phe
biosynthesis. However, the molecular and biochemical evolution of the plant arogenate
pathway is currently unknown. Here, we conducted phylogenetically informed
biochemical characterization of prephenate aminotransferases (PPA-ATs) that belong to class-Ib aspartate aminotransferases
(AspAT Ibs) and catalyze the first
committed step of the arogenate pathway in plants. Plant PPA-ATs and succeeding arogenate dehydratases (ADTs) were found to be most closely related to
homologs from Chlorobi/Bacteroidetes bacteria. The Chlorobium
tepidum
PPA-AT and ADT homologs indeed efficiently converted prephenate and arogenate into
arogenate and Phe, respectively. A subset of AspAT
Ib enzymes exhibiting PPA-AT
activity was further identified from both Plantae and prokaryotes and, together with
site-directed mutagenesis, showed that Thr-84 and Lys-169 play key roles in specific
recognition of dicarboxylic keto (prephenate) and amino (aspartate) acid substrates.
The results suggest that, along with ADT, a gene encoding
prephenate-specific PPA-AT was transferred
from a Chlorobi/Bacteroidetes ancestor to a eukaryotic ancestor of Plantae, allowing
efficient Phe and phenylpropanoid production via arogenate in plants today. 相似文献
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Haiyun Pan Rui Zhou Gordon V. Louie Jo?lle K. Mühlemann Erin K. Bomati Marianne E. Bowman Natalia Dudareva Richard A. Dixon Joseph P. Noel Xiaoqiang Wang 《The Plant cell》2014,26(9):3709-3727
The enzymes cinnamoyl-CoA reductase (CCR) and cinnamyl alcohol dehydrogenase (CAD) catalyze the two key reduction reactions in the conversion of cinnamic acid derivatives into monolignol building blocks for lignin polymers in plant cell walls. Here, we describe detailed functional and structural analyses of CCRs from Medicago truncatula and Petunia hybrida and of an atypical CAD (CAD2) from M. truncatula. These enzymes are closely related members of the short-chain dehydrogenase/reductase (SDR) superfamily. Our structural studies support a reaction mechanism involving a canonical SDR catalytic triad in both CCR and CAD2 and an important role for an auxiliary cysteine unique to CCR. Site-directed mutants of CAD2 (Phe226Ala and Tyr136Phe) that enlarge the phenolic binding site result in a 4- to 10-fold increase in activity with sinapaldehyde, which in comparison to the smaller coumaraldehyde and coniferaldehyde substrates is disfavored by wild-type CAD2. This finding demonstrates the potential exploitation of rationally engineered forms of CCR and CAD2 for the targeted modification of monolignol composition in transgenic plants. Thermal denaturation measurements and structural comparisons of various liganded and unliganded forms of CCR and CAD2 highlight substantial conformational flexibility of these SDR enzymes, which plays an important role in the establishment of catalytically productive complexes of the enzymes with their NADPH and phenolic substrates. 相似文献
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Fang Xu Yan Huang Lin Li Patrick Gannon Eric Linster Monika Huber Paul Kapos Willy Bienvenut Bogdan Polevoda Thierry Meinnel Rüdiger Hell Carmela Giglione Yuelin Zhang Markus Wirtz She Chen Xin Li 《The Plant cell》2015,27(5):1547-1562
Nod-like receptors (NLRs) serve as immune receptors in plants and animals. The stability of NLRs is tightly regulated, though its mechanism is not well understood. Here, we show the crucial impact of N-terminal acetylation on the turnover of one plant NLR, Suppressor of NPR1, Constitutive 1 (SNC1), in Arabidopsis thaliana. Genetic and biochemical analyses of SNC1 uncovered its multilayered regulation by different N-terminal acetyltransferase (Nat) complexes. SNC1 exhibits a few distinct N-terminal isoforms generated through alternative initiation and N-terminal acetylation. Its first Met is acetylated by N-terminal acetyltransferase complex A (NatA), while the second Met is acetylated by N-terminal acetyltransferase complex B (NatB). Unexpectedly, the NatA-mediated acetylation serves as a degradation signal, while NatB-mediated acetylation stabilizes the NLR protein, thus revealing antagonistic N-terminal acetylation of a single protein substrate. Moreover, NatA also contributes to the turnover of another NLR, RESISTANCE TO P. syringae pv maculicola 1. The intricate regulation of protein stability by Nats is speculated to provide flexibility for the target protein in maintaining its homeostasis. 相似文献
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Christian P. Craddock Nicolette Adams Fiona M. Bryant Smita Kurup Peter J. Eastmond 《The Plant cell》2015,27(4):1251-1264
Regulation of membrane lipid biosynthesis is critical for cell function. We previously reported that disruption of PHOSPHATIDIC ACID PHOSPHOHYDROLASE1 (PAH1) and PAH2 stimulates net phosphatidylcholine (PC) biosynthesis and proliferation of the endoplasmic reticulum (ER) in Arabidopsis thaliana. Here, we show that this response is caused specifically by a reduction in the catalytic activity of the protein and positively correlates with an accumulation of its substrate, phosphatidic acid (PA). The accumulation of PC in pah1 pah2 is suppressed by disruption of CTP:PHOSPHOCHOLINE CYTIDYLYLTRANSFERASE1 (CCT1), which encodes a key enzyme in the nucleotide pathway for PC biosynthesis. The activity of recombinant CCT1 is stimulated by lipid vesicles containing PA. Truncation of CCT1, to remove the predicted C-terminal amphipathic lipid binding domain, produced a constitutively active enzyme. Overexpression of native CCT1 in Arabidopsis has no significant effect on PC biosynthesis or ER morphology, but overexpression of the truncated constitutively active version largely replicates the pah1 pah2 phenotype. Our data establish that membrane homeostasis is regulated by lipid composition in Arabidopsis and reveal a mechanism through which the abundance of PA, mediated by PAH activity, modulates CCT activity to govern PC content. 相似文献
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Dong-Dong Wang Hua Bai Wei-Qi Chen Hai Lu Xiang-Ning Jiang 《Journal of Plant Biology》2009,52(5):482-491
A cinnamoyl coenzyme A reductase (CCR, EC 1.2.1.44), one of the key enzyme involved in lignin biosynthesis, was cloned from
Populus tomentosa (Chinese white poplar). At the same time, a 4CL1 gene was cloned from P. tomentosa, too. The two genes were subcloned in pQE31 vector and expressed in Escherichia coli M15. Both of them were purified by Ni-NTA. Purified CCR protein was digested by trypsin and analyzed by HPLC-MS; the peptide
segments had 27% similarity with the sequence of the CCR protein. 4CL was thought to be a neighbor enzyme of CCR in lignin
biosynthesis. In this paper, a 4CL1 from P. tomentosa was cloned, and its enzyme reaction products were extracted for the substrates of CCR. Three 4CL1 enzyme reaction products
were monitored by HPLC-MS and then the CCR enzyme reaction was detected by GC-MS. In the CCR reaction, the three corresponding
aldehyde (p-coumaraldehyde, caffealdehyde, and coniferaldehyde) were detected and identified by Frontier3 software. The results showed
that the CCR that we cloned from P. tomentosa had affinities with 4CL1 enzyme reaction products and a ptCCR that was cloned from aspen (Li et al., Plant Cell Physiol 46(7):1073–1082, 2005) only had affinity with feruloyl-CoA. The different results maybe depend on the different study method. The method of exacting
4CL enzyme products as the substrates of CCR in the paper was reliable and can be used in lignin biosynthesis network to detect
the enzymes in the neighborhood that depended on the polarity of the substrates and products. This CCR gene had eight homology
sequence CCR gene when a BLAST was conducted in Populus trichocarpa genome database. The CCR homology genes in Populus suggested that some CCRs may take part in the lignin biosynthesis, too. The gene family would be the hot spot in the future
study. 相似文献
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Uma D. Vempati Xianlin Han Carlos T. Moraes 《The Journal of biological chemistry》2009,284(7):4383-4391
Cytochrome c (cyt c) is a heme-containing protein that
participates in electron transport in the respiratory chain and as a signaling
molecule in the apoptotic cascade. Here we addressed the effect of removing
mammalian cyt c on the integrity of the respiratory complexes in
mammalian cells. Mitochondria from cyt c knockout mouse cells lacked
fully assembled complexes I and IV and had reduced levels of complex III. A
redox-deficient mutant of cyt c was unable to rescue the levels of
complexes I and IV. We found that cyt c is associated with both
complex IV and respiratory supercomplexes, providing a potential mechanism for
the requirement for cyt c in the assembly/stability of complex
IV.The mitochondrial electron transport chain consists of four multisubunit
complexes, namely, NADH-ubiquinone oxidoreductase (complex
I),2
succinate-ubiquinone oxidoreductase (complex II), ubiquinone-cytochrome
c oxidoreductase (complex III), and cytochrome c oxidase
(complex IV, COX). Cytochrome c (cyt c) shuttles electrons
from oxidative phosphorylation complex III to complex IV. Electrons are
transferred from reduced cyt c sequentially to the CuA
site, heme a, heme a3, and CuB
binuclear center in the complex IV before being finally transferred to
molecular oxygen to generate water
(1). Respiratory complexes are
assembled into supercomplexes (also called respirasomes). These contain
complex I bound to dimeric complex III and a variable copy number of complex
IV (2).In Saccharomyces cerevisiae, cyt c is encoded by two
genes: CYC1 and CYC7. Mutagenesis studies in yeast have
shown that cyt c is required for the assembly of COX
(3,
4). In yeast lacking both the
cyt c genes (CYC1 and CYC7), COX assembly was
absent. It was also shown that cyt c is only structurally required
for COX assembly, because a catalytic mutant of cyt c (W65S) was
sufficient to bring about near normal levels of COX. However, because yeast
lacks complex I, they could not analyze the role of cyt c in the
assembly/stability of complex I. Mammals possess two different isoforms of cyt
c encoded on different chromosomes: the somatic (cyt cS)-
and testis (cyt cT)-specific isoforms. In mouse, the cDNAs bear 74%
homology, whereas the proteins possess 86% identity with most dissimilarity in
the C terminus.Cardiolipin (CL) is an anionic phospholipid present almost exclusively in
the mitochondrial membranes and constitutes 25% of its total phospholipids
(5). Work from several
laboratories showed that CL is essential for the membrane anchorage of the
respiratory supercomplexes. CL has two main roles in the mitochondrial
structure and function, namely, stabilization of mitochondrial membranes and
specific interactions with proteins. CL deficiency results in inefficient
energy transformation by oxidative phosphorylation, swelling of mitochondria,
decreased ATP/oxygen ratio, and reduced membrane potential
(6,
7). In accordance, in S.
cerevisiae lacking CL synthase, the supercomplex comprising complexes III
and IV is unstable (8).
Assembly mutants of COX had significantly reduced CL synthase activity,
whereas assembly mutants of respiratory complex III and complex V showed less
inhibition (9). Subsequently,
the proton gradient across the inner mitochondrial membrane was found to be
important for CL formation and that CL synthase was stimulated by alkaline pH
at the matrix side (10). In
this study, we investigated the role of cyt c depletion on CL levels
by examining its content and composition in cyt c null cells.Here we aimed to answer the following questions: What is the role of cyt
c in the assembly and maintenance of the different respiratory
complexes in mammals? Are there changes in the content/composition of lipids
in the cyt c-ablated cells? Analysis of mouse fibroblasts revealed
that cyt c is essential for the assembly/stability of COX, and a
catalytically mutant form of cyt c cannot rescue the COX defect in
the cyt c null cells. CL and triacylglycerols showed significant
differences in the cyt c null cells, both in content and
composition. 相似文献
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Thomas S. Peat Christine B?ttcher Janet Newman Del Lucent Nathan Cowieson Christopher Davies 《The Plant cell》2012,24(11):4525-4538
Auxins are important for plant growth and development, including the control of fruit ripening. Conjugation to amino acids by indole-3-acetic acid (IAA)-amido synthetases is an important part of auxin homeostasis. The structure of the auxin-conjugating Gretchen Hagen3-1 (GH3-1) enzyme from grapevine (Vitis vinifera), in complex with an inhibitor (adenosine-5′-[2-(1H-indol-3-yl)ethyl]phosphate), is presented. Comparison with a previously published benzoate-conjugating enzyme from Arabidopsis thaliana indicates that grapevine GH3-1 has a highly similar domain structure and also undergoes a large conformational change during catalysis. Mutational analyses and structural comparisons with other proteins have identified residues likely to be involved in acyl group, amino acid, and ATP substrate binding. Vv GH3-1 is a monomer in solution and requires magnesium ions solely for the adenlyation reaction. Modeling of IAA and two synthetic auxins, benzothiazole-2-oxyacetic acid (BTOA) and 1-naphthaleneacetic acid (NAA), into the active site indicates that NAA and BTOA are likely to be poor substrates for this enzyme, confirming previous enzyme kinetic studies. This suggests a reason for the increased effectiveness of NAA and BTOA as auxins in planta and provides a tool for designing new and effective auxins. 相似文献
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Chloroplast division is performed by the constriction of envelope membranes at the division site. Although constriction of a ring-like protein complex has been shown to be involved in chloroplast division, it remains unknown how membrane lipids participate in the process. Here, we show that phosphoinositides with unknown function in envelope membranes are involved in the regulation of chloroplast division in Arabidopsis thaliana. PLASTID DIVISION1 (PDV1) and PDV2 proteins interacted specifically with phosphatidylinositol 4-phosphate (PI4P). Inhibition of phosphatidylinositol 4-kinase (PI4K) decreased the level of PI4P in chloroplasts and accelerated chloroplast division. Knockout of PI4Kβ2 expression or downregulation of PI4Kα1 expression resulted in decreased levels of PI4P in chloroplasts and increased chloroplast numbers. PI4Kα1 is the main contributor to PI4P synthesis in chloroplasts, and the effect of PI4K inhibition was largely abolished in the pdv1 mutant. Overexpression of DYNAMIN-RELATED PROTEIN5B (DRP5B), another component of the chloroplast division machinery, which is recruited to chloroplasts by PDV1 and PDV2, enhanced the effect of PI4K inhibition, whereas overexpression of PDV1 and PDV2 had additive effects. The amount of DRP5B that associated with chloroplasts increased upon PI4K inhibition. These findings suggest that PI4P is a regulator of chloroplast division in a PDV1- and DRP5B-dependent manner. 相似文献
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Tae-Houn Kim Hans-Henning Kunz Saikat Bhattacharjee Felix Hauser Jiyoung Park Cawas Engineer Amy Liu Tracy Ha Jane E. Parker Walter Gassmann Julian I. Schroeder 《The Plant cell》2012,24(12):5177-5192
In a chemical genetics screen we identified the small-molecule [5-(3,4-dichlorophenyl)furan-2-yl]-piperidine-1-ylmethanethione (DFPM) that triggers rapid inhibition of early abscisic acid signal transduction via PHYTOALEXIN DEFICIENT4 (PAD4)- and ENHANCED DISEASE SUSCEPTIBILITY1 (EDS1)-dependent immune signaling mechanisms. However, mechanisms upstream of EDS1 and PAD4 in DFPM-mediated signaling remain unknown. Here, we report that DFPM generates an Arabidopsis thaliana accession-specific root growth arrest in Columbia-0 (Col-0) plants. The genetic locus responsible for this natural variant, VICTR (VARIATION IN COMPOUND TRIGGERED ROOT growth response), encodes a TIR-NB-LRR (for Toll-Interleukin1 Receptor–nucleotide binding–Leucine-rich repeat) protein. Analyses of T-DNA insertion victr alleles showed that VICTR is necessary for DFPM-induced root growth arrest and inhibition of abscisic acid–induced stomatal closing. Transgenic expression of the Col-0
VICTR allele in DFPM-insensitive Arabidopsis accessions recapitulated the DFPM-induced root growth arrest. EDS1 and PAD4, both central regulators of basal resistance and effector-triggered immunity, as well as HSP90 chaperones and their cochaperones RAR1 and SGT1B, are required for the DFPM-induced root growth arrest. Salicylic acid and jasmonic acid signaling pathway components are dispensable. We further demonstrate that VICTR associates with EDS1 and PAD4 in a nuclear protein complex. These findings show a previously unexplored association between a TIR-NB-LRR protein and PAD4 and identify functions of plant immune signaling components in the regulation of root meristematic zone-targeted growth arrest. 相似文献
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Alexis Samba Mialoundama Nurul Jadid Julien Brunel Thomas Di Pascoli Dimitri Heintz Mathieu Erhardt Jér?me Mutterer Marc Bergdoll Daniel Ayoub Alain Van Dorsselaer Alain Rahier Paul Nkeng Philippe Geoffroy Michel Miesch Bilal Camara Florence Bouvier 《The Plant cell》2013,25(12):4879-4893
Sterols are vital for cellular functions and eukaryotic development because of their essential role as membrane constituents. Sterol biosynthetic intermediates (SBIs) represent a potential reservoir of signaling molecules in mammals and fungi, but little is known about their functions in plants. SBIs are derived from the sterol C4-demethylation enzyme complex that is tethered to the membrane by Ergosterol biosynthetic protein28 (ERG28). Here, using nonlethal loss-of-function strategies focused on Arabidopsis thaliana ERG28, we found that the previously undetected SBI 4-carboxy-4-methyl-24-methylenecycloartanol (CMMC) inhibits polar auxin transport (PAT), a key mechanism by which the phytohormone auxin regulates several aspects of plant growth, including development and responses to environmental factors. The induced accumulation of CMMC in Arabidopsis erg28 plants was associated with diagnostic hallmarks of altered PAT, including the differentiation of pin-like inflorescence, loss of apical dominance, leaf fusion, and reduced root growth. PAT inhibition by CMMC occurs in a brassinosteroid-independent manner. The data presented show that ERG28 is required for PAT in plants. Furthermore, it is accumulation of an atypical SBI that may act to negatively regulate PAT in plants. Hence, the sterol pathway offers further prospects for mining new target molecules that could regulate plant development. 相似文献