Background Goat milk is increasingly recognized for high digestibility and a distinctive compositional profile.Protein acetylation,an important post-translational modification,regulates biosynthetic and metabolic path...Background Goat milk is increasingly recognized for high digestibility and a distinctive compositional profile.Protein acetylation,an important post-translational modification,regulates biosynthetic and metabolic pathways.This study aimed to identify critical acetylated proteins and specific modification sites involved in milk production and component synthesis in dairy goats,thereby elucidating the molecular mechanisms of lactation.We performed a comparative TMT-based acetylomic and proteomic analysis of mammary tissues from Saanen dairy goats during peak lactation and the dry period using LC–MS/MS.A candidate acetylation site was further investigated in goat mammary epithelial cells(GMECs)through site-directed mutagenesis and lipid metabolic assays,establishing functional links between acetylation and mammary lipid metabolism and providing a foundation for molecular strategies to improve milk quality and yield.Results We established a comprehensive mammary acetylome,identifying 862 significantly acetylated proteins and 2,028 modification sites across the two physiological phases.Differentially acetylated proteins were predominantly localized to the cytoplasm(39.98%).From these,54 key acetylated proteins,including MTOR,BCAT2,QARS1,GOT1,GOT2,BDH1,ACSS1,STAT5B,FABP5,and GPAM were prioritized as candidates involved in milk protein synthesis,milk fat synthesis,lactose synthesis,and other lactation-related processes.Among them,β-hydroxybutyrate dehydrogenase 1(BDH1)acetylation was characterized in detail.Members of the HDAC family were identified as primary regulators mediating BDH1 deacetylation.BDH1 acetylation promoted lipid droplet formation and triglyceride synthesis in GMECs.At the transcriptional level,BDH1 acetylation upregulated LXRα,ACSL1 and SCD1,whereas deacetylation downregulated SCD1,FASN,and ACSL1.Notably,BDH1 acetylation/deacetylation significantly reduced SREBP1 expression,linking this modification to coordinated control of lipogenic gene networks.Conclusions This study established,for the first time,the comprehensive acetylome of mammary gland tissues in dairy goats,revealing a substantial number of differentially acetylated proteins and modification sites.We demonstrate that acetylation of BDH1 regulated by HDACs promotes lipid droplet biogenesis and triglyceride synthesis in GMECs through transcriptional modulation of key lipogenic genes and suppression of SREBP1.These findings provide mechanistic insights into the post-translational regulation of mammary lipid metabolism and offer molecular targets for future genetic and nutritional strategies aimed at enhancing milk quality and yield in dairy goats.展开更多
The current study describes the molecular characterization of a clone which can restore the ability of bdh A mutant strains NGRPA2 and Rm11107 to utilize 3-hydroxybutyrate as a sole carbon source (Hbu+ ). This clone w...The current study describes the molecular characterization of a clone which can restore the ability of bdh A mutant strains NGRPA2 and Rm11107 to utilize 3-hydroxybutyrate as a sole carbon source (Hbu+ ). This clone was screened out by complementation experiment from Bradyrhizobium japonicum US-DAI 10 genomic library, and the presence of bdhA gene in the clone was verified by Bdh assay and Southern blot analysis. Furthermore, the entire sequence of bdhA. gene was sequenced and the sequence was deposited in GenBank database under the accession number AY077581. bdhA gene comprises 789 base pairs and encodes Bdh with 262 amino acid of MW 27.59 kDa. Interposon JlKm was inserted into the bdhA ORF at EcoR I site and the bdhA mutant was constructed in B .japonicum by homologous recombination. Plant assay result did not show obvious effects of mutation of bdhA gene on nodulation and nitrogen-fixation.展开更多
This paper will correct some gaps existing in the proof of a theorem written in an earlier paper by the author on the lower bounds for sums of BDH type published in 1993. Some improvements upon the previous version of...This paper will correct some gaps existing in the proof of a theorem written in an earlier paper by the author on the lower bounds for sums of BDH type published in 1993. Some improvements upon the previous version of that theorem will be obtained.展开更多
目的探讨β-羟丁酸脱氢酶2(BDH2)基因对肝癌细胞增殖能力的抑制作用及其机制。方法合成表达BDH2的慢病毒载体,构建稳定表达BDH2的肝癌细胞株HepG2-BDH2(实验组)和对照细胞株HepG2-Vector(对照组)。RT-PCR检测两组细胞中BDH2 m RNA表达...目的探讨β-羟丁酸脱氢酶2(BDH2)基因对肝癌细胞增殖能力的抑制作用及其机制。方法合成表达BDH2的慢病毒载体,构建稳定表达BDH2的肝癌细胞株HepG2-BDH2(实验组)和对照细胞株HepG2-Vector(对照组)。RT-PCR检测两组细胞中BDH2 m RNA表达水平。采用CCK-8法检测两组肝癌细胞的增殖能力,平板克隆实验观察两组细胞的克隆形成能力。Western blot检测BDH2蛋白和Bcl-2细胞凋亡相关蛋白的表达。实验数据比较采用t检验。结果实验组BDH2 mRNA表达量为(2.20±0.10)×10^(-3),明显高于对照组的(0.20±0.01)×10^(-3)(t=34.95,P<0.05)。实验组细胞培养3、4、5、6、7 d的A_(450)值分别为0.55±0.20、0.73±0.02、1.26±0.12、1.62±0.14、2.19±0.12,明显低于对照组的0.70±0.06、1.13±0.08、1.77±0.15、2.45±0.12、3.02±0.15(t=-5.19,-11.34,-5.96,-10.35,-9.54;P<0.05)。细胞生长曲线显示实验组肝癌细胞的增殖能力明显弱于对照组。平板克隆实验结果显示,实验组细胞克隆形成数目为(184±7)个,明显少于对照组的(429±15)个(t=-25.84,P<0.05)。与对照组相比,实验组的BDH2、cleaved caspase-3蛋白表达明显升高,而Bcl-2蛋白表达明显降低。结论 BDH2基因可抑制肝癌细胞增殖,其机制可能是通过Bcl-2信号通路促进肝癌细胞的凋亡。展开更多
基金supported by the National Key Research and Development Program of China(2022YFF1000102)Xi’an Agricultural Technology Research General Project(24NYGG0025)the National Natural Science Foundation of China(31702098)。
文摘Background Goat milk is increasingly recognized for high digestibility and a distinctive compositional profile.Protein acetylation,an important post-translational modification,regulates biosynthetic and metabolic pathways.This study aimed to identify critical acetylated proteins and specific modification sites involved in milk production and component synthesis in dairy goats,thereby elucidating the molecular mechanisms of lactation.We performed a comparative TMT-based acetylomic and proteomic analysis of mammary tissues from Saanen dairy goats during peak lactation and the dry period using LC–MS/MS.A candidate acetylation site was further investigated in goat mammary epithelial cells(GMECs)through site-directed mutagenesis and lipid metabolic assays,establishing functional links between acetylation and mammary lipid metabolism and providing a foundation for molecular strategies to improve milk quality and yield.Results We established a comprehensive mammary acetylome,identifying 862 significantly acetylated proteins and 2,028 modification sites across the two physiological phases.Differentially acetylated proteins were predominantly localized to the cytoplasm(39.98%).From these,54 key acetylated proteins,including MTOR,BCAT2,QARS1,GOT1,GOT2,BDH1,ACSS1,STAT5B,FABP5,and GPAM were prioritized as candidates involved in milk protein synthesis,milk fat synthesis,lactose synthesis,and other lactation-related processes.Among them,β-hydroxybutyrate dehydrogenase 1(BDH1)acetylation was characterized in detail.Members of the HDAC family were identified as primary regulators mediating BDH1 deacetylation.BDH1 acetylation promoted lipid droplet formation and triglyceride synthesis in GMECs.At the transcriptional level,BDH1 acetylation upregulated LXRα,ACSL1 and SCD1,whereas deacetylation downregulated SCD1,FASN,and ACSL1.Notably,BDH1 acetylation/deacetylation significantly reduced SREBP1 expression,linking this modification to coordinated control of lipogenic gene networks.Conclusions This study established,for the first time,the comprehensive acetylome of mammary gland tissues in dairy goats,revealing a substantial number of differentially acetylated proteins and modification sites.We demonstrate that acetylation of BDH1 regulated by HDACs promotes lipid droplet biogenesis and triglyceride synthesis in GMECs through transcriptional modulation of key lipogenic genes and suppression of SREBP1.These findings provide mechanistic insights into the post-translational regulation of mammary lipid metabolism and offer molecular targets for future genetic and nutritional strategies aimed at enhancing milk quality and yield in dairy goats.
文摘The current study describes the molecular characterization of a clone which can restore the ability of bdh A mutant strains NGRPA2 and Rm11107 to utilize 3-hydroxybutyrate as a sole carbon source (Hbu+ ). This clone was screened out by complementation experiment from Bradyrhizobium japonicum US-DAI 10 genomic library, and the presence of bdhA gene in the clone was verified by Bdh assay and Southern blot analysis. Furthermore, the entire sequence of bdhA. gene was sequenced and the sequence was deposited in GenBank database under the accession number AY077581. bdhA gene comprises 789 base pairs and encodes Bdh with 262 amino acid of MW 27.59 kDa. Interposon JlKm was inserted into the bdhA ORF at EcoR I site and the bdhA mutant was constructed in B .japonicum by homologous recombination. Plant assay result did not show obvious effects of mutation of bdhA gene on nodulation and nitrogen-fixation.
文摘This paper will correct some gaps existing in the proof of a theorem written in an earlier paper by the author on the lower bounds for sums of BDH type published in 1993. Some improvements upon the previous version of that theorem will be obtained.
