scholarly journals Different genome-wide transcriptome responses of Nocardioides simplex VKM Ac-2033D to phytosterol and cortisone 21-acetate

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Victoria Yu Shtratnikova ◽  
Mikhail I. Sсhelkunov ◽  
Victoria V. Fokina ◽  
Eugeny Y. Bragin ◽  
Andrey A. Shutov ◽  
...  
Keyword(s):  
Dehydrogenase Activity ◽  
Transcriptome Profiling ◽  
Comparative Investigation ◽  
Mycolic Acid ◽  
Gene Organization ◽  
Expression Level ◽  
Bacterial Degradation ◽  
Genome Wide ◽  
Steroid Catabolism

Abstract Background Bacterial degradation/transformation of steroids is widely investigated to create biotechnologically relevant strains for industrial application. The strain of Nocardioides simplex VKM Ac-2033D is well known mainly for its superior 3-ketosteroid Δ1-dehydrogenase activity towards various 3-oxosteroids and other important reactions of sterol degradation. However, its biocatalytic capacities and the molecular fundamentals of its activity towards natural sterols and synthetic steroids were not fully understood. In this study, a comparative investigation of the genome-wide transcriptome profiling of the N. simplex VKM Ac-2033D grown on phytosterol, or in the presence of cortisone 21-acetate was performed with RNA-seq. Results Although the gene patterns induced by phytosterol generally resemble the gene sets involved in phytosterol degradation pathways in mycolic acid rich actinobacteria such as Mycolicibacterium, Mycobacterium and Rhodococcus species, the differences in gene organization and previously unreported genes with high expression level were revealed. Transcription of the genes related to KstR- and KstR2-regulons was mainly enhanced in response to phytosterol, and the role in steroid catabolism is predicted for some dozens of the genes in N. simplex. New transcription factors binding motifs and new candidate transcription regulators of steroid catabolism were predicted in N. simplex. Unlike phytosterol, cortisone 21-acetate does not provide induction of the genes with predicted KstR and KstR2 sites. Superior 3-ketosteroid-Δ1-dehydrogenase activity of N. simplex VKM Ac-2033D is due to the kstDs redundancy in the genome, with the highest expression level of the gene KR76_27125 orthologous to kstD2, in response to cortisone 21-acetate. The substrate spectrum of N. simplex 3-ketosteroid-Δ1-dehydrogenase was expanded in this study with progesterone and its 17α-hydroxylated and 11α,17α-dihydroxylated derivatives, that effectively were 1(2)-dehydrogenated in vivo by the whole cells of the N. simplex VKM Ac-2033D. Conclusion The results contribute to the knowledge of biocatalytic features and diversity of steroid modification capabilities of actinobacteria, defining targets for further bioengineering manipulations with the purpose of expansion of their biotechnological applications.

Adventures inRhodococcus — from steroids to explosivesThis article is based on a presentation by Dr. Lindsay Eltis at the 60th Annual Meeting of the Canadian Society of Microbiologists in Hamilton, Ontario, 14  June 2010. Dr. Eltis was the recipient of the 2010 Norgen Biotek Corporation / CSM Award, an annual award sponsored by Norgen Biotek and the Canadian Society of Microbiologists intended to recognize outstanding scientific work in microbiology by a Canadian researcher.

2011 ◽  
Vol 57 (3) ◽  
pp. 155-168 ◽  
Author(s):  
Katherine C. Yam ◽  
Sachi Okamoto ◽  
Joseph N. Roberts ◽  
Lindsay D. Eltis
Keyword(s):  
Polychlorinated Biphenyl ◽  
Molecular Genetic ◽  
Oxidative Degradation ◽  
Model Organism ◽  
Scientific Work ◽  
Mycolic Acid ◽  
Bacterial Degradation ◽  
Canadian Society ◽  
Rhodococcus Jostii Rha1

Rhodococcus is a genus of mycolic-acid-containing actinomycetes that utilize a remarkable variety of organic compounds as growth substrates. This degradation helps maintain the global carbon cycle and has increasing applications ranging from the biodegradation of pollutants to the biocatalytic production of drugs and hormones. We have been using Rhodococcus jostii RHA1 as a model organism to understand the catabolic versatility of Rhodococcus and related bacteria. Our approach is exemplified by the discovery of a cluster of genes specifying the catabolism of cholesterol. This degradation proceeds via β-oxidative degradation of the side chain and O2-dependent cleavage of steroid ring A in a process similar to bacterial degradation of aromatic compounds. The pathway is widespread in Actinobacteria and is critical to the pathogenesis of Mycobacterium tuberculosis , arguably the world’s most successful pathogen. The close similarity of some of these enzymes with biphenyl- and polychlorinated-biphenyl-degrading enzymes that we have characterized is facilitating inhibitor design. Our studies in RHA1 have also provided important insights into a number of novel metalloenzymes and their biosynthesis, such as acetonitrile hydratase (ANHase), a cobalt-containing enzyme with no significant sequence identity with characterized nitrile hydratases. Molecular genetic and biochemical studies have identified AnhE as a dimeric metallochaperone that delivers cobalt to ANHase, enabling its maturation in vivo. Other metalloenzymes we are characterizing include N-acetylmuramic acid hydroxylase, which catalyzes an unusual hydroxylation of the rhodococcal and mycobacterial peptidoglycan, and 2 RHA1 dye-decolorizing peroxidases. Using molecular genetic and biochemical approaches, we have demonstrated that one of these enzymes is involved in the degradation of lignin. Overall, our studies are providing fundamental insights into a range of catabolic processes that have a wide variety of applications.

scholarly journals Downregulated PITX1 Modulated by MiR-19a-3p Promotes Cell Malignancy and Predicts a Poor Prognosis of Gastric Cancer by Affecting Transcriptionally Activated PDCD5

2018 ◽  
Vol 46 (6) ◽  
pp. 2215-2231 ◽  
Author(s):  
Fengchang Qiao ◽  
Pihai Gong ◽  
Yunwei Song ◽  
Xiaohui Shen ◽  
Xianwei Su ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cell Proliferation ◽  
Cell Growth ◽  
Cell Apoptosis ◽  
Expression Level ◽  
Qrt Pcr ◽  
Genome Wide ◽  
Patient Prognosis

Background/Aims: PITX1 has been identified as a potential tumor-suppressor gene in several malignant tumors. The molecular mechanism underlying PITX1, particularly its function as a transcription factor regulating gene expression during tumorigenesis, is still poorly understood. Methods: The expression level and location of PITX1 were determined by quantitative reverse transcription PCR (qRT-PCR) and immunohistochemical staining in gastric cancer (GC). The effect of PITX1 on the GC cell proliferation and tumorigenesis was analyzed in vitro and in vivo. To explore how PITX1 suppresses cell proliferation, we used PITX1-ChIP-sequencing to measure genome-wide binding sites of PITX1 and assessed global function associations based on its putative target genes. ChIP-PCR, electrophoretic mobility shift assay, and promoter reporter assays examined whether PITX1 bound to PDCD5 and regulated its expression. The function of PDCD5 in GC cell apoptosis was further examined in vitro and in vivo. The relationship between the PITX1 protein level and GC patient prognosis was evaluated by the Kaplan-Meier estimator. Meanwhile, the expression level of miR-19a-3p, which is related to PITX1, was also detected by luciferase reporter assay, qRT-PCR, and western blotting. Results: The expression level of PITX1 was decreased in GC tissues and cell lines. Elevated PITX1 expression significantly suppressed the cell proliferation of GC cells and tumorigenesis in vitro and in vivo. PITX1 knockdown blocked its inhibition of GC cell proliferation. PITX1 bound to whole genome-wide sites, with these targets enriched on genes with functions mainly related to cell growth and apoptosis. PITX1 bound to PDCD5, an apoptosis-related gene, during tumorigenesis, and cis-regulated PDCD5 expression. Increased PDCD5 expression in GC cells not only induced GC cell apoptosis, but also suppressed GC cell growth in vitro and in vivo. Moreover, PITX1 expression was regulated by miR-19a-3p. More importantly, a decreased level of PITX1 protein was correlated with poor GC patient prognosis. Conclusion: Decreased expression of PITX1 predicts shorter overall survival in GC patients. As a transcriptional activator, PITX1 regulates apoptosis-related genes, including PDCD5, during gastric carcinogenesis. These data indicate PDCD5 to be a novel and feasible therapeutic target for GC.

Kinase Targets for Mycolic Acid Biosynthesis in Mycobacterium tuberculosis

2019 ◽  
Vol 12 (1) ◽  
pp. 27-49 ◽  
Author(s):  
Shahinda S.R. Alsayed ◽  
Chau C. Beh ◽  
Neil R. Foster ◽  
Alan D. Payne ◽  
Yu Yu ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Enzymatic Activity ◽  
Bacterial Pathogen ◽  
Building Blocks ◽  
Mycolic Acid ◽  
Adaptive Responses ◽  
Mycolic Acids ◽  
Hydroxylated Fatty Acids

Background:Mycolic acids (MAs) are the characteristic, integral building blocks for the mycomembrane belonging to the insidious bacterial pathogen Mycobacterium tuberculosis (M.tb). These C60-C90 long α-alkyl-β-hydroxylated fatty acids provide protection to the tubercle bacilli against the outside threats, thus allowing its survival, virulence and resistance to the current antibacterial agents. In the post-genomic era, progress has been made towards understanding the crucial enzymatic machineries involved in the biosynthesis of MAs in M.tb. However, gaps still remain in the exact role of the phosphorylation and dephosphorylation of regulatory mechanisms within these systems. To date, a total of 11 serine-threonine protein kinases (STPKs) are found in M.tb. Most enzymes implicated in the MAs synthesis were found to be phosphorylated in vitro and/or in vivo. For instance, phosphorylation of KasA, KasB, mtFabH, InhA, MabA, and FadD32 downregulated their enzymatic activity, while phosphorylation of VirS increased its enzymatic activity. These observations suggest that the kinases and phosphatases system could play a role in M.tb adaptive responses and survival mechanisms in the human host. As the mycobacterial STPKs do not share a high sequence homology to the human’s, there have been some early drug discovery efforts towards developing potent and selective inhibitors.Objective:Recent updates to the kinases and phosphatases involved in the regulation of MAs biosynthesis will be presented in this mini-review, including their known small molecule inhibitors.Conclusion:Mycobacterial kinases and phosphatases involved in the MAs regulation may serve as a useful avenue for antitubercular therapy.

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