Cloning, Expression and Purification of Wheat Acetyl-CoA Carboxylases CT Domain in E. coil

The entire gene of carboxyltransferase（CT） domain of acetyl-CoA carboxylase（ACCase） from Chinese Spring wheat（CSW） plastid was cloned firstly, and the 2.3 kb gene was inserted into PET28a^＋ vector and expressed in E. coil in a soluble state. The （His）6 fusion protein was identified by SDS-PAGE and Western blot. The recombinant protein was purified by affinity chromatography, and the calculated molecular mass（Mr） was 88000. The results of the sequence analysis indicate that the cloned gene（GeneBank accession No. EU124675） was a supplement and revision of the reported ACCase CT partial cDNA from Chinese Spring wheat plastid. The recombinant protein will be significant for us to investigate the recognizing mechanism between ACCase and herbicides, and further to screen new herbicides.

Acetyl-CoA carboxylase inhibitors in non-alcoholic steatohepatitis: Is there a benefit?

De novo lipogenesis(DNL)plays an important role in the pathogenesis of hepatic steatosis and also appears to be implicated in hepatic inflammation and fibrosis.Accordingly,the inhibition of acetyl-CoA carboxylase,which catalyzes the ratelimiting step of DNL,might represent a useful approach in the management of patients with nonalcoholic fatty liver disease(NAFLD).Animal studies and preliminary data in patients with NAFLD consistently showed an improvement in steatosis with the use of these agents.However,effects on fibrosis were variable and an increase in plasma triglyceride levels was observed.Therefore,more longterm studies are needed to clarify the role of these agents in NAFLD and to determine their risk/benefit profile.

Detection of Acetyl-CoA Carboxylase （ACCase） Inhibitor Herbicides Resistance in Sterile Wild Oat （Avena sterilis L.） Using Agar Quick Test

This study was conducted using seeds in Petri dish containing agar medium in order to determine acetyl-CoA carboxylase （ACCase） herbicides resistance （R） in Avena sterilis that was grown in wheat fields at Adana province, Turkey. Seeds were collected from one large suspected field, where clodinafop-propargyl （Aryloxyphenoxypropionate “FOPs”） and pinoxaden （Phenylpyrazoline “DEN”） have been applied for many years. Susceptible （S） population was collected from the road side on the same region. Agar media of concentration 14 g/L was prepared and it was melted in microwave. Then the amount of 20 mL agar media was added into each Petri dish. Five seeds were placed on agar mediums containing discriminating dose of clodinafop and pinoxaden. Petri dishes were placed in growth incubator operating at 10 °C. After 15 d, both radicle and hypocotyl length were measured. The percentage of germinated seed and dose-response curves were determined. At these different concentration levels, there were more than 50% of R and less than 40% of S seed germinated for pinoxaden. However, for clodinafop, more than 60% of R and less than 50% of S seeds were germinated. At higher concentration levels, the populations of resistant and susceptible were not germinated for both herbicides. The resistance value of R population was then compared with that of the S biotype. From the resistance index （RI）, the population was more resistant to pinoxaden （7.43 for radicle and 2.47 for hypocotyl） than the clodinafop-propagyl （1.39 for radicle and 3.77 for hypocotyl）. The method provided a simple, quick and cost effective way to identify ACCase herbicides resistance in most grass weeds.

Photosynthetic Characteristics of Transgenic Rice Plants Overexpressing Maize Phosphoenopyruvate Carboxylase

用转PEPC、PPDK、NADP_ME、PEPC +PPDK 双基因水稻 (OryzasativaL .)及原种为材料 ,以光合酶活性、饱和光合速率及PSⅡ光化学效率 (Fv/Fm)为指标 ,研究了转PEPC基因水稻的光合生理特征。结果如下 :转PEPC基因水稻PEPC活性比原种提高 2 0倍 ;饱和光合速率比原种高 5 5 % ;用高光强或人工光氧化剂甲基紫精 (MV)处理后 ,与原种相比 ,转PEPC基因水稻光化学效率下降较少 ,证明其耐光抑制、耐光氧化能力增强 ,测定其光合日变化看出 :在 1d中不同时间 ,转PEPC基因水稻的光合速率均高于原种 ,且与PEPC活性的日变化有相似的趋势。上述结果为转PEPC基因水稻的生理机制和育种研究提供了依据和途径。

Preliminary Study on Phosphoenolpyruvate Carboxylase(PEPCase) Gene Introduced into Wheat

[ Objective] The aim of this study was to introduce Phosphoenolpyruvate Carboxylase （PEPCase） gene into common wheat Linyou 145. [ Method] With the material of common wheat Linyou 145, Phosphoenolpyruvate Carboxylase （PEPCase） gene was introduced into wheat embryo callus by the agrobacterium-mediated transformation system, and then analyzed through successive selection with selective medium con- taing gygrornycin to detect the gene at the molecular level. [Result] The hyg-resistant plants were obtained, and GUS histochemical staining showed the leaf of resistant plants was stained dark blue. The target bands appeared in PCR analysis. [ Conclusion] Phosphoenolpyruvate Car- boxylase （PEPCase） gene has been primarily introduced into the recipient material.

高效液相色谱法(HPLC)测定保加利亚乳杆菌胞内Acetyl-CoA

建立了一种快速测定保加利亚乳杆菌胞内Acetyl—CoA含量的HPLC分析方法。以C18（HypersilODS25μm）色谱柱为固定相，以缓冲液A（0．2mol／麟酸钠，pH=5）和缓冲液B（800mL0．25m01／I磷酸钠，pH=5和200mL乙腈的混合物）为流动相，梯度洗脱，流速为1mE]min，柱温25℃，采用紫外检测器（254nm）进行检测。结果表明：该方法可以在17rain内实现EAcetyl—CoA与其他物质完全分离和定量，Acetyl—CoA在0．011-0．359μmol／mL范围内的线性关系良好，回归方程线性相关系数R2=0．9995，检出限（以信噪比（S／N）为3计）为0．28mg／L，定量限（r2S／N为10计）为0．32mg／L。该方法用于保加利亚乳杆菌胞内Acetyl—CoA含量的测定，相对标准偏差（n=6）为0．75％，重现性良好，三个水平的加标回收率为94．8％～103．3％，该方法适用于保加利亚乳杆菌胞内Acetyl—CoA快速、高效分离和定量。

Identification and Expression Analysis of the Phosphoenolpyruvate Carboxylase Gene Family in Peanut (Arachis hypogaea L.)

Phosphoenolpyruvate carboxylase （PEPC） is widely distributed in plants and bacteria, and catalyzes the carboxylation of phosphoenolpyruvate to form oxaloacetate and inorganic phosphate. To investigate the molecular mechanisms of the regulation and control of peanut oil, with the degenerated primers and RACE-PCR approach, five PEPC genes were cloned from peanut, and designated as AhPEPC1, AhPEPC2, AhPEPC3, AhPEPC4, and AhPEPC5, respectively. The structure and phylogenetic analysis of PEPC protein indicated that AhPEPC1-4 genes encoded a typical plant-type PEPC-enzyme, and AhPEPC5 a bacterial-type. By real-time quantitative RT-PCR approach the expression pattern of each gene was detected in various tissues of normal and high oil-content peanut varieties. It was found that there was a lower expression level of AhPEPCs genes except for the AhPEPC2 in high-oil peanut than normal-oil peanut line. The results provide some fundamental information for the further investigation of plant PEPC proteins and their role in regulation of oil-content in peanut seeds.

Functional Analysis of the Phosphoenolpyruvate Carboxylase on the Lipid Accumulation of Peanut(Arachis hypogaea L.) Seeds

Phosphoenolpyruvate carboxylase（PEPC;EC 4.1.1.31） catalyses phosphoenolpyruvate（PEP） to yield oxaloacetate,which is involved in protein biosynthesis.Pyruvate kinase（PK;EC 2.7.1.40） catalyzes PEP to yield pyruvate,which is involved in fatty acid synthesis.In this study,five PEPC genes（AhPEPC1,AhPEPC2,AhPEPC3,AhPEPC4,and AhPEPC5） from peanut have been cloned.Using a quantitative real-time RT-PCR approach,the expression pattern of each gene was monitored during the seed development of four peanut varieties（E11,Hebeigaoyou,Naihan 1,and Huayu 26）.It was found that these five genes shared similar expression behaviors over the developmental stages of E11 with high expression levels at 30 and 40 d after pegging（DAP）;whereas these five genes showed irregular expression patterns during the seed development of Hebeigaoyou.In Naihan 1 and Huayu 26,the expression levels of the five genes remained relatively high in the first stage.The PEPC activity was monitored during the seed development of four peanut varieties and seed oil content was also characterized during whole period of seed development.The PEPC activity followed the oil accumulation pattern during the early stages of development but they showed a significantly negative correlation thereafter.These results suggested that PEPC may play an important role in lipid accumulation during the seed development of four peanut varieties tested.

Characterization of ribulose-1,5-bisphosphate carboxylase/oxygenase and transcriptional analysis of its related genes in Saccharina japonica(Laminariales,Phaeophyta)

Saccharina japonica is a common macroalga in sublittoral communities of cold seawater environments, and consequently may have highly efficient ribulose-1, 5-bisphosphate carboxylase/ oxygenase （Rubisco） activity for carbon assimilation. In our study, we cloned the full-length Rubisco gene from S.japonica （SJ-rbc）. It contained an open reading frame for a large subunit gene （SJ-rbcL） of 1 467 bp, a small subunit gene （SJ-rbcS） of 420 bp, and a SJ-rbcL/S intergenie spacer of 269 bp. The deduced peptides of SJ-rbcL and SJ-rbcS were 488 and 139 amino acids with theoretical molecular weights and isoelectric points of 53.97 kDa, 5.81 and 15,84 kDa, 4.71, respectively. After induction with 1 mmol/L isopropyl-β-D- thiogalactopyranoside for 5 h and purification by Ni2＋ affinity chromatography, electrophoresis and western blot detection demonstrated successful expression of the 55 kDa SJ-rbcL protein. Real-time quantitative PCR showed that the mRNA levels of SJ-rbcL in gametophytes increased when transferred into normal growth conditions and exhibited diurnal variations： increased expression during the day but suppressed expression at night. This observation implied that Rubisco played a role in normal gametophytic growth and development. In juvenile sporophytes, mRNA levels of SJ-rbcL, carbonic anhydrase, Calvin-Benson- Bassham cycle-related enzyme, and chloroplast light-harvesting protein were remarkably increased under continuous light irradiance. Similarly, expression of these genes was up-regulated under blue light irradiance at 350 umol/（m2.s）. Our results indicate that long-term white light and short-term blue light irradiance enhances juvenile sporophytic growth by synergistic effects of various photosynthetic elements.

Photosynthetic Characteristics and Heterosis in Transgenic Hybrid Rice with Maize Phosphoenolpyruvate Carboxylase (pepc) Gene

Three F3 hybrids derived from the sterile rice lines Gang 46A, 776A and 2480A and the improved restorer line Shuhui 881 containing maize phosphoenolpyruvate carboxylase （pepc） gene were used to analyze the effect of pepc gene on the heterosis and photosynthetic characteristics, while the F3 obtained by crossing Shuhui 881 with the above three sterile lines served as controls. The dynamics of photosynthetic characteristics in leaves of three F1 with pepc gene and their controls were determined at the initial-tillering, maxium-tillering, elongation, initial-heading, heading, maturity stages, and other different times after flag leaf fully expanded. The PEPCase activities of the three F1 with pepc gene increased significantly as compared with control plants during the whole developmental stages. Moreover, the net photosynthesis rate （Pn） also increased to certain extent. The data showed that PEPCase activity was significantly correlated to Pn with a correlation coefficient of 0.6081. The photosynthetic indexes of the three F1 with pepc gene were obviously superior to respective controls in apparent quantum efficiency, light compensation point and carboxylation efficiency, while the CO2 compensation point was lower than that of corresponding control. The Pn of the three F1 with pepc gene at light saturation point and CO2 saturation point was also higher than that of control plants. in addition, the three F1 with pepc gene had an average increase of 37.10% in grain yields per plant in comparison with control plants. The results indicated that the photosynthetic characteristics of hybrid rice containing pepc gene had been improved to some extent due to the introduction of pepc gene.

Genetic Mutation of Vitamin K-dependent Gamma-glutamyl Carboxylase Domain in Patients with Calcium Oxalate Urolithiasis

To investigate the exon mutation of vitamin K-dependent gamma-glutamyl carboxylase （GGCX or VKDC） in patients with calcium oxalate urolithasis, renal cortex and peripheral blood samples were obtained from severe hydronephrosis patients （with or without calculi）, and renal tumor patients undergoing nephrectomy. GGCX mutations in all 15 exons were examined in 44 patients with calcium oxalate urolithiasis （COU） by polymerase chain reaction （PCR） and denatured high pressure liquid chromatography （DHPLC）, and confirmed by sequencing. Mutation was not found in all COU samples compared to the controls. These data demonstrated that functional GGCX mutations in all 15 exons do not occur in most COU patients. It was suggested that there may be no significant association between the low activity and mutation of GGCX in COU.

Concomitant Increases of the Developing Seed Phosph<i>oenol</i>pyruvate Carboxylase Ac-tivity and the Seed Protein Content of Field-Grown Wheat with Nitrogen Supply

Wheat seed storage protein is of great importance for human food. To increase the contents of storage proteins effectively, nitrogen fertilizer at flowering stages is commonly applied. In our previous study, rice phosphoenolpyruvate carboxylase (PEPCase) activity in developing seeds was observed in response to nitrogen application at a flowering stage and was positively correlated to the response of the protein content in seeds of six cultivars. This observation might indicate that the seeds have a biological system for accepting nitrogen in seeds by using PEPCase. To test whether this physiological event occurs in wheat, we examined the PEPCase activity and protein content in field-grown wheat seeds under different nitrogen supply conditions. With only basal dressing, seeds showed lower PEPCase activity and protein content (both 0.90-fold) compared to seeds without basal fertilizer. With ammonium sulfate application at 8.3 and 25 g/m2 at a flowering stage, seeds showed higher PEPCase activity (1.08- and 1.17-fold, respectively) and protein content (1.15- and 1.42-fold, respectively), depending on the nitrogen level. We investigated the relationship between PEPCase activity and protein content in the seeds among four conditions. The effect of the nitrogen supply on PEPCase activity during grain-filling stages was validated by the results of a hydroponic culture experiment. Together the results demonstrate that our hypothesis seems to apply to field-grown wheat.

Entamoeba histolytica acetyl-CoA synthetase:biomarker of acute amoebic liver abscess

Objective:To characterize the Entamoeba histolytica(E.histolytica)antigen(s)recognized by moribound amoebic liver abscess hamsters.Methods:Crude soluble antigen of E.histolytica was probed with sera of moribund hamsters in 1D-and 2D-Westem blot analyses.The antigenic protein was then sent for tandem mass spectrometry analysis.The corresponding gene was cloned and expressed in Escherichia coli BL21-AI to produce the recombinant E.histolytica ADP-forming acetyl-CoA synthetase(EhACS)protein.A customised ELISA was developed to evaluate the sensitivity and specificity of the recombinant protein.Results:A^75 kDa protein band with a pl value of 5.91-6.5 was found to be antigenic;and not detected by sera of hamsters in the control group.Tandem mass spectrometry analysis revealed the protein to be the 77 kDa E.histolytica ADP-forming acetyl-CoA synthetase(EhACS).The customised ELISA results revealed 100%sensitivity and 100%specificity when tested against infected(n=31)and control group hamsters(n=5)serum samples,respectively.Conclusions:This rinding suggested the significant role of EhACS as a biomarker for moribund hamsters with acute amoebic liver abscess(ALA)infection.It is deemed pertinent that future studies explore the potential roles of EhACS in better understanding the pathogenesis of ALA;and in the development of vaccine and diagnostic tests to control ALA in human populations.

Differential effect of biotin on carboxylase activity and mice skeletal muscle metabolism

In mammalian skeletal muscle there are four carboxylases involved in several biochemical processes like gluconeogenesis, tricarboxylic acid cycle anaplerosis, metabolism of fatty acids and metabolism of various amino acids. It has been shown that biotin deficiency reduces body weight at the expense of muscular mass. When necessary, the liver uses skeletal muscle protein to provide glucose and amino acids to organs in need of such compounds. In this paper we analyzed carboxylase specific activities in hind limb skeletal muscle of 3 weeks old BALB/c male mice, at 0, 1, 4, 7, and 14 days of a specific diet with different biotin concentrations. Biotin was used at 0.0, 1.8 or 98.2 mg per kg of food;and was referred to as biotin deficient, sufficient and supplemented, respectively. Water and food supply and consumption by the three groups of mice were the same. Therefore, the observed effects were directly related to biotin ingestion. The body weight of biotin supplemented mice was the same as the body weight of mice in the biotin sufficient group, while biotin deficiency caused body weight reduction after 7 days of biotin depletion. We found that the total protein concentration in the vastus lateralis muscle is associated with the biotin content in the diet. After 7 days, the muscle total protein content was lower in mice of the biotin deficient group while it was higher in the mice from the biotin supplemented group

重庆地区马唐种群对2种乙酰辅酶A羧化酶抑制剂类除草剂的敏感性

为高效科学防除重庆地区马唐Digitaria sanguinalis(L.)Scop.,采用整株生物测定法,测定了不同马唐种群对2种乙酰辅酶A羧化酶抑制剂类除草剂的ED_(50)和相对抗性倍数(RI)。结果表明,不同马唐种群对噁唑酰草胺、烯草酮均产生了不同程度的抗性(相对敏感种群除外);DJSP02对噁唑酰草胺、烯草酮分别表现为中抗(RI=8.78)和低抗(RI=2.10),其ED_(50)分别为377.31 g/hm^(2)和37.97 g/hm^(2);DJSP03对噁唑酰草胺、烯草酮分别表现为低抗(RI=3.18)和中抗(RI=7.56),其ED_(50)分别为136.84 g/hm^(2)和136.93 g/hm^(2),均已产生了正交互抗性。因此,在对重庆地区抗性马唐种群进行防除时宜与其他类型的除草剂混用或交替使用。

水生植物RuBisCO基因的趋同演化

植物碳固定,作为碳循环中的核心环节,对全球粮食产量与气候变迁具有深远影响。在此过程中,核酮糖-1,5-二磷酸羧化酶(RuBisCO)的作用举足轻重。其催化速率与特异性间存在微妙的平衡:特异性降低,催化速率上升,但副反应中的有毒副产物亦随之增多,从而影响整体效率。然而,水生植物却拥有独特的CO_(2)富集机制,有效减少了副反应的影响,为酶创造了优越的工作环境。本文以高等水生植物入水事件为切入点,深入研究了其核酮糖-1,5-二磷酸羧化酶大亚基蛋白的趋同进化。旨在寻找与入水事件紧密相关的趋同进化位点,并探究CO_(2)富集机制下该酶理化性质的变化,为酶的优化提供新思路。收集了泽泻目、唇形目、虎耳草目、水韭目、山茱萸目和金鱼藻目等6个目共48个物种的酶蛋白质序列信息,并确定了6次关键的入水事件。采用PCOC分析方法,整合了这6次入水事件的数据,深入剖析了水生植物的进化趋势。分析结果显示,水生植物中可能存在一个关键的趋同进化位点,位于蛋白质序列的第328号位置,紧邻酶的活性中心。推测该位点的替换可能增强了反应底物进入活性中心区域的灵活性,使CO_(2)更易进入,从而提升了酶的催化速率。这一发现为后续的酶改进突变实验指明了方向,提供了坚实的理论依据。研究成果不仅有助于优化该酶的催化效率和适应性,也为深入研究植物碳固定机制提供了新的视角。

不同进化类型大豆品种光合碳代谢关键酶活性研究

光合碳代谢是大豆生长发育过程中基础的生理代谢途径,对其自身的产量和品质有重要的影响。为了探索大豆品种演化过程中光合特性的变化规律和碳代谢的演化趋势,本研究以野生、半野生、半栽培和栽培大豆品种为试验材料,对不同进化类型大豆光合特性以及光合碳代谢关键酶活性进行研究。结果表明:净光合速率、气孔导度、蒸腾速率、RuBP羧化酶活性和叶绿素含量在所有类型的大豆品种中均呈现单峰曲线变化,均以栽培型最高,最大值在R2期,栽培型净光合速率、叶绿素含量和RuBP羧化酶活性分别为24.2μmol·m^(-2)·s^(-1)、99.9 mg·g^(-1)和4.5μmol·g^(-1)·min^(-1)。碳代谢相关酶活性也均呈单峰曲线变化,蔗糖合成酶、蔗糖磷酸合成酶、淀粉合成酶、ADPG焦磷酸化酶和UDPG焦磷酸化酶活性最大值均出现在R2期,栽培型大豆品种的蔗糖合成酶和ADPG焦磷酸化酶活性显著高于其它类型品种,分别较野生、半野生、半栽培品种高203.4%、67.8%、49.2%和192.5%、142.4%和131.1%。各类型大豆叶片碳代谢相关酶活性,在V2和R2期与Pn呈显著或者极显著正相关,在R4、R6和R8期与Pn无显著相关性或呈负相关,随着大豆品种的演化,叶片光合碳同化能力改善,其中RuBP羧化酶、蔗糖合成酶和ADPG焦磷酸化酶活性增强幅度较高。研究结果可为大豆品种选育及高光效利用研究提供理论基础。

The Ca^(2+)/CaN/ACC and cAMP/PKA/HK signal pathways are required for PBAN-mediated sex pheromone biosynthesis in Conogethes punctiferalis

Conogethes punctiferalis is a crop and fruit pest that has caused serious economic losses to agricultural production.This pest relies heavily on its sex pheromone to ensure sexual encounters and subsequent mating success.However,the molecular mechanism underlying sex pheromone biosynthesis in this species remains elusive.The present study investigated the detailed mechanism underlying PBAN-regulated sex pheromone biosynthesis in C.punctiferalis by transcriptome sequencing of the C.punctiferalis pheromone glands(PGs)and subsequent functional identification of the target genes.The results showed that female mating started from the first scotophase,and peaked at the second to fifth scotophases in accordance with the release of sex pheromones.PBAN regulated sex pheromone biosynthesis by employing Ca^(2+)and cAMP as secondary messengers,as demonstrated by RNA interference(RNAi),pharmacological inhibitors,and behavioral assays.Further investigation revealed that calcineurin(CaN)and acetyl-CoA carboxylase(ACC)were activated by PBAN/Ca^(2+)signaling,and the RNAimediated knockdown of CaN and ACC transcripts significantly reduced sex pheromone production,ultimately leading to a significantly reduced ability of females to attract males.Importantly,hexokinase(HK)was found to regulate sex pheromone biosynthesis in response to the PBAN/cAMP/PKA signaling pathway,as demonstrated by RNAi,enzyme activity,and pharmacological inhibitor assays.Furthermore,Far2 and Desaturase1 were found to participate in PBAN-regulated sex pheromone biosynthesis.Altogether,our findings revealed that PBAN regulates sex pheromone biosynthesis through the PBANR/Ca^(2+)/CaN/ACC and PBANR/cAMP/PKA/HK pathways in C.punctiferalis,which enriches our comprehension of the details of sex pheromone biosynthesis in moths.

乙酰辅酶A羧化酶2通过调控细胞周期促进肝癌细胞增殖

目的:研究乙酰辅酶A羧化酶2(acetyl-co carboxylase 2,ACC2)对肝癌细胞增殖、迁移能力的影响以及其潜在的作用机制。方法:通过TCGA、GEO、CPTAC数据库对ACC2在肝癌(hepatocellular carcinoma,HCC)患者肝脏组织中的表达和预后价值进行分析。采用CRISPR-Cas9系统构建了ACC2敲低肝癌细胞系,观察肝癌细胞增殖、迁移能力以及细胞周期的变化,Western blot实验检测细胞周期相关蛋白的表达。结果:ACC2在肝癌组织中低表达(P<0.05)且与预后不良相关(P<0.05)。体外细胞功能学实验表明降低ACC2表达显著促进肝癌细胞增殖、迁移能力(P<0.05)。细胞周期流式分析显示敲低ACC2促进肝癌细胞周期G1/S期的转变(P<0.05),细胞周期相关蛋白中CyclinE2和p21的表达降低,而CyclinA2和p-RB的表达升高。结论:ACC2可以促进肝癌细胞增殖和迁移的能力,对细胞增殖的促进作用是通过加速肝癌细胞周期G1向S期的转化实现的。

敲低乙酰辅酶A羧化酶1对食管鳞状细胞癌KYSE450细胞迁移的影响及机制

目的探讨敲低乙酰辅酶A羧化酶1(ACC1)对食管鳞状细胞癌KYSE450细胞迁移能力的影响及机制。方法将对数生长期食管鳞状细胞癌KYSE450细胞随机分为shNC组、shACC1组、shNC+AEB071组及shACC1+AEB071组,shNC组KYSE450细胞转染空白质粒载体,shACC1组KYSE450细胞转染慢病毒质粒载体,shNC+AEB071组KYSE450细胞转染空白质粒载体后加入5μL浓度为2 mmoL·L^(-1)的蛋白激酶C抑制剂AEB071(终浓度为5μmoL·L^(-1)),shACC1+AEB071组KYSE450细胞转染慢病毒质粒载体后加入5μL浓度为2 mmoL·L^(-1)的蛋白激酶C抑制剂AEB071(终浓度为5μmoL·L^(-1))。Transwell法检测各组KYSE450细胞迁移能力;显微镜下观察各组KYSE450细胞形态学变化;Western blot检测4组KYSE450细胞中乙酰辅酶A羧化酶1(ACC1)、组蛋白H3(H3)、乙酰化的组蛋白H3第9赖氨酸(H3K9Ac)及上皮-间质转化(EMT)标志物β-连环蛋白(β-catenin)、波形蛋白(Vimentin)以及锌指转录因子(Snail)等的表达。结果shACC1组KYSE450细胞迁移数显著高于shNC组(P<0.05);shNC+AEB071组与shNC组KYSE450细胞迁移数比较差异无统计学意义(P>0.05);shACC1+AEB071组KYSE450细胞迁移数显著低于shACC1组(P<0.05)。shNC组KYSE450细胞呈椭圆形上皮样细胞形态,shACC1组KYSE450细胞呈类似于梭形的间质样细胞形态,shNC+AEB071组和shACC1+AEB071组KYSE450细胞呈椭圆形上皮样细胞形态。shACC1组KYSE450细胞中ACC1相对表达量显著低于shNC组(P<0.05),β-catenin、Vimentin、Snail的相对表达量及H3K9Ac/H3比值显著高于shNC组(P<0.05);shNC+AEB071组与shNC组KYSE450细胞中ACC1、β-catenin、Vimentin、Snail相对表达量及H3K9Ac/H3比值比较差异无统计学意义(P>0.05);shACC1+AEB071组KYSE450细胞中β-catenin、Vimentin、Snail的相对表达量及H3K9Ac/H3比值显著低于shACC1组(P<0.05);shACC1+AEB071组与shACC1组KYSE450细胞中ACC1相对表达量比较差异无统计学意义(P>0.05)。结论敲低ACC1可促进食管鳞状细胞癌KYSE450细胞的迁移,从而促进肿瘤的进展,其机制可能是通过蛋白激酶C相关信号通路介导的。