Microarray analysis for differential gene expression in M1-SDHHbtail
Ontology highlight
ABSTRACT:
ORGANISM(S): Streptococcus pyogenes
PROVIDER: GSE15231 | GEO | 2011/06/15
SECONDARY ACCESSION(S): PRJNA116425
REPOSITORIES: GEO
Similar Datasets
Project description:A total of 163 genes were found to be differentially expressed; 38 genes of them were up regulated and 125 genes were down regulated. Most notably the functional categories that were affected include Virulence genes (18 genes 11% of the total significantly differentiated genes), carbohydrate metabolism and cell envelop realted genes ( 31, 19.1% of the total), and aminoacid transport genes (21genes, 12.9%). Among the virulence-related genes, the most notable one were those belong to the production of capsule, the M protein, exotoxin, NAD glycohydrolase and Streptolysin S. Some of the capsule related genes were down regulated by more than 64 fold. These results indicate that CdhA (group A Streptococcal Cell division controlling and Chain-forming cell wall hydrolase) plays an important role in the regulation of virulence.
2010-10-01 | GSE15546 | GEO
Project description:A total of 215 gene were found to be differentially expressed; 90 of them were up regulated and 125 genes were down regulated. Most notably the functional categories that were affected include Virulence genes, carbohydrate genes and lipid synthesis genes. Some of the virulence genes were down regulated by more than 16-32 fold. These results indicate that SDH plays an important role in the regulation of virulence. S.pyogenes strain SF370 (wild-type) was procured from ATCC (ATCC 700294). M1-SDHHbtail (SDH cytoplasmically retained) mutant strain PGB-1 was obtained using pFW5 vector as described before ( Boel G. Jin, H. and Pancholi, V. (2005). Inhibition of cell surface export of groupA streptococcal anchorless surface dehydrogenase (SDH) affects bacterial adherence and antiphagocytic properties. Infect. Immun. 73:6237-6248). Both strains (M1-Wild-type and PGB1(M1-SDHHbtail) were grown in Todd-Hewitt broth to their late log phase. Bacteria were harvested by centrifugation and washed twice with sterile PBS. Total RNA was isolated from these washed streptococci using Qiagen RNeasy Mini kit. For synthesis and labeling of cDNA, 20 ug of total RNA, 1.5 ug of random hexamer, 0.5mM dNTPs (except that 0.2mM of dTTP was replaced by the same amount of amino-allyl dUTP to incorporate dUTP into first-strand cDNA) were used in each reverse transcriptase reaction (Superscript II Reverse Transcriptase, Invitrogen). Purified cDNA preparations from the wild-type and PGB1 mutant strains were labeled with either Alexafluor-555 or Alexafluor-647 depending on the experimental design ( i.e. dye swap experiment). Differentially labeled probes were then combined and purified. Using three independently isolated RNA preparations (biological replicates), a total of 8 experiments (incorporating dye swaps) were performed. Accordingly eight hybridization measurements for this mutant were performed. Thus Exp-1 and -2 (GSM380310, GSM380320) are the technical replicates of the biological sample-1, Exp-3,-4 and -5 (GSM380321, GSM380322, GSM380331) are technical replicates of biological sample-2 and Exp-6,-7, and -8 (GSM380332,GSM380333, GSM380334) are technical replicates of the biological sample-3. Signals of the bound reagents on the microarray spots in terms of relative fluorescence values were measured and quantified by a laser scanner (GenePix 4100 ) at 10um/pixel resolution. The resulting images were processed using Gene Pix Pro software (version 4.0, Axon instruments). The raw data were obtained in the form of GenPix *.gpr output files. The web application CARMAweb (Comprehensive R based microarray analysis web service) was used for the normalization and analysis of microarray data. All raw data (in the form of *. GPR files) were uploaded to the web application in the data directory. Using appropriate navigation tree, background correction from the foreground signal was applied and within microarray normalization was achieved using the Lowess method. Genes flagged as bad spots by the scanning software were excluded from the analysis and all flagged spots were given a weight of zero. The normalized data were then subjected to fold-change analysis and t-statistics using Bioconductor multtest package. CARMAweb allows to set Log2 cut-off values for both the M (regulation) and the A (average expression) values. Differentially expressed genes (Log2 values of Mutant647-red vs. wild-type555-green or Log2 values Mutant-555(green) vs Wild-type 647-red) were defined based on cut-off value >1 Log2< i.e. all genes that show a two or more-fold up- or down-regulation. Bioconductors Multtest package provided suitable method to adjust P values according to multiple hypothesis testing problems.
2011-06-15 | E-GEOD-15231 | biostudies-arrayexpress
Project description:A total of 192 genes were found to be differentially expressed; 38 genes of them were up regulated and 154 genes were down regulated. Most notably the functional categories that were affected include carbohydrate metabolism and cell envelop realted genes ( 56, 29.1% of the total),Virulence genes (12 genes 6.25% of the total significantly differentiated genes), and aminoacid transport genes (7 genes, 3.6 %). Among the virulence-related genes, the most notable ones were those belong to the production of capsule, the M protein, exotoxin, NAD glycohydrolase and Streptolysin S. Some of the capsule related genes were down regulated by more than 64 fold. Additionally 11 differentially expressed lipid metabolism related genes and four aminoacid transport and metabolism-related genes were upregulated. These results indicate that the C-terminal CHAP domain of CdhA (group A Streptococcal Cell division controlling and Chain-forming cell wall hydrolase) plays an important role in the regulation of virulence. Since these mutant lack cell wall hydrolase activity but are not defective in cell division or growth, we believe that CdhA isa multifunctional protein with N-terminal region controlling cell division and c-terminal region responsible for regulating bacterial virulence.
2010-10-01 | GSE15598 | GEO
Project description:A total of 163 genes were found to be differentially expressed; 38 genes of them were up regulated and 125 genes were down regulated. Most notably the functional categories that were affected include Virulence genes (18 genes 11% of the total significantly differentiated genes), carbohydrate metabolism and cell envelop realted genes ( 31, 19.1% of the total), and aminoacid transport genes (21genes, 12.9%). Among the virulence-related genes, the most notable one were those belong to the production of capsule, the M protein, exotoxin, NAD glycohydrolase and Streptolysin S. Some of the capsule related genes were down regulated by more than 64 fold. These results indicate that CdhA (group A Streptococcal Cell division controlling and Chain-forming cell wall hydrolase) plays an important role in the regulation of virulence. S.pyogenes strain SF370 (wild-type) was procured from ATCC (ATCC 700294). M1-CdhA(-) mutant strain was obtained using pFW5 vector containing spectinomycin resistance marker (aad9). The mutant lacking CdhA is highly defective in cell division and growth, lacks antiphagocytic property and attenuated for virulence. Both strains (M1-Wild-type and M1-CdhA(-) were grown in Todd-Hewitt broth to their late log phase. Bacteria were harvested by centrifugation and washed twice with sterile PBS. Total RNA was isolated from these washed streptococci using Qiagen RNeasy Mini kit. For synthesis and labeling of cDNA, 20 ug of total RNA, 1.5 ug of random hexamer, 0.5mM dNTPs (except that 0.2mM of dTTP was replaced by the same amount of amino-allyl dUTP to incorporate dUTP into first-strand cDNA) were used in each reverse transcriptase reaction (Superscript II Reverse Transcriptase, Invitrogen). Purified cDNA preparations from the wild-type and M1CdhA(-) mutant strains were labeled with either Alexafluor-555 or Alexafluor-647 depending on the experimental design ( i.e. dye swap experiment). Differentially labeled probes were then combined and purified. Using three independently isolated RNA preparations (biological replicates), a total of 11 experiments (incorporating dye swaps) were performed. Accordingly eleven hybridization measurements for this mutant were performed. Thus Exp-1 to -4 (GSM388703, GSM388717, GSM388718 and GSM388719) are the technical replicates of the biological sample-1, Exp-5 to 8(GSM388732, GSM388733, GSM388734, and GSM388735) are technical replicates of biological sample-2 and Exp-9,-10, and -11 (GSM388736,GSM388737, and GSM388738) are technical replicates of the biological sample-3. Signals of the bound reagents on the microarray spots in terms of relative fluorescence values were measured and quantified by a laser scanner (GenePix 4100 ) at 10um/pixel resolution. The resulting images were processed using Gene Pix Pro software (version 4.0, Axon instruments). The raw data were obtained in the form of GenPix *.gpr output files. The web application CARMAweb (Comprehensive R based microarray analysis web service) was used for the normalization and analysis of microarray data. All raw data (in the form of *. GPR files) were uploaded to the web application in the data directory. Using appropriate navigation tree, background correction from the foreground signal was applied and within microarray normalization was achieved using the Lowess method. Genes flagged as bad spots by the scanning software were excluded from the analysis and all flagged spots were given a weight of zero. The normalized data were then subjected to fold-change analysis and t-statistics using Bioconductor multtest package. CARMAweb allows to set Log2 cut-off values for both the M (regulation) and the A (average expression) values. Differentially expressed genes (Log2 values of Mutant647-red vs. wild-type555-green or Log2 values Mutant-555(green) vs Wild-type 647-red) were defined based on cut-off value >1 Log2< i.e. all genes that show a two or more-fold up- or down-regulation. Bioconductors Multtest package provided suitable method to adjust P values according to multiple hypothesis testing problems.
2010-10-01 | E-GEOD-15546 | biostudies-arrayexpress
Project description:A total of 192 genes were found to be differentially expressed; 38 genes of them were up regulated and 154 genes were down regulated. Most notably the functional categories that were affected include carbohydrate metabolism and cell envelop realted genes ( 56, 29.1% of the total),Virulence genes (12 genes 6.25% of the total significantly differentiated genes), and aminoacid transport genes (7 genes, 3.6 %). Among the virulence-related genes, the most notable ones were those belong to the production of capsule, the M protein, exotoxin, NAD glycohydrolase and Streptolysin S. Some of the capsule related genes were down regulated by more than 64 fold. Additionally 11 differentially expressed lipid metabolism related genes and four aminoacid transport and metabolism-related genes were upregulated. These results indicate that the C-terminal CHAP domain of CdhA (group A Streptococcal Cell division controlling and Chain-forming cell wall hydrolase) plays an important role in the regulation of virulence. Since these mutant lack cell wall hydrolase activity but are not defective in cell division or growth, we believe that CdhA isa multifunctional protein with N-terminal region controlling cell division and c-terminal region responsible for regulating bacterial virulence. S. pyogenes strain SF370 (wild-type) was procured from ATCC (ATCC 700294). M1-CdhA55(-) mutant strain was obtained using pFW5 vector containing spectinomycin resistance marker (aad9). The mutant expressing truncated CdhA that lack C-terminal 55 residues constituting catalytic site of the CHAP domain is not defective in cell division and growth, but lacks antiphagocytic property and attenuated for virulence. Both strains (M1-Wild-type and M1-CdhA55(-) were grown in Todd-Hewitt broth to their late log phase. Bacteria were harvested by centrifugation and washed twice with sterile PBS. Total RNA was isolated from these washed streptococci using Qiagen RNeasy Mini kit. For synthesis and labeling of cDNA, 20 ug of total RNA, 1.5 ug of random hexamer, 0.5mM dNTPs (except that 0.2mM of dTTP was replaced by the same amount of amino-allyl dUTP to incorporate dUTP into first-strand cDNA) were used in each reverse transcriptase reaction (Superscript II Reverse Transcriptase, Invitrogen). Purified cDNA preparations from the wild-type and M1-CdhA55(-) mutant strains were labeled with either Alexafluor-555 or Alexafluor-647 depending on the experimental design ( i.e. dye swap experiment). Differentially labeled probes were then combined and purified. Using three independently isolated RNA preparations (biological replicates), a total of 7 experiments (incorporating dye swaps) were performed. Accordingly 7 hybridization measurements for this mutant were performed. Thus Exp-1 and 2 (GSM389993, GSM389994) are the technical replicates of the biological sample-1, Exp-3 and 4 (GSM389995, GSM390054) are technical replicates of biological sample-2 and Exp-5,-6, and -7 (GSM390055,GSM390056, and GSM390057) are technical replicates of the biological sample-3. Signals of the bound reagents on the microarray spots in terms of relative fluorescence values were measured and quantified by a laser scanner (GenePix 4100 ) at 10um/pixel resolution. The resulting images were processed using Gene Pix Pro software (version 4.0, Axon instruments). The raw data were obtained in the form of GenPix *.gpr output files. The web application CARMAweb (Comprehensive R based microarray analysis web service) was used for the normalization and analysis of microarray data. All raw data (in the form of *. GPR files) were uploaded to the web application in the data directory. Using appropriate navigation tree, background correction from the foreground signal was applied and within microarray normalization was achieved using the Lowess method. Genes flagged as bad spots by the scanning software were excluded from the analysis and all flagged spots were given a weight of zero. The normalized data were then subjected to fold-change analysis and t-statistics using Bioconductor multtest package. CARMAweb allows to set Log2 cut-off values for both the M (regulation) and the A (average expression) values. Differentially expressed genes (Log2 values of Mutant647-red vs. wild-type555-green or Log2 values Mutant-555(green) vs Wild-type 647-red) were defined based on cut-off value >1 Log2< i.e. all genes that show a two or more-fold up- or down-regulation. BioConductors Multtest package provided suitable method to adjust P values according to multiple hypothesis testing problems.
2010-10-01 | E-GEOD-15598 | biostudies-arrayexpress
Project description:Francisellosis outbreaks have been reported in the literature as occurring seasonally when water temperature is below 24°C. The aim of this study was to quantify the median lethal doses (LD50) of FNO in experimental challenges at 28°C and 22°C, and to investigate the impact of temperature changes in whole genome expression using microarray technology. The LD50 for Nile tilapia at 28°C was ~105.7, whereas at 22°C, the LD50 was ~102.2, showing that the decrease in temperature enhanced pathogen virulence. Out of 1,917 genes screened, a total of 31 and 19 genes were down- and up-regulated at 22°C, respectively. These genes were grouped by orthology into functional categories of: amino acid, inorganic ion, and carbohydrate transport and metabolism; transcription; and posttranslational modification, protein turnover, and chaperones. Expression of genes related to metabolism, oxidative stress, and thermal shock were regulated by temperature changes, reflecting an ability of FNO to adapt to the environment. Expression of virulence genes usually required for the Francisella genus was not changed between tested temperatures, including that of genes located on the Francisella Pathogenicity Island.
2019-12-31 | GSE125115 | GEO
Project description:This analysis revealed ~17.% of all ORFs (292 out of a total 1697 ORFs) differed by at least 1.5 fold (Log2 +0.6) between the mutant and wild-type strains. Of these, ~45% of the ORFs (130 out of 292 ORFs) revealed at least 2-fold (Log2 +1) differences. The complete spectrum of differentially expressed genes in the muatnt strain revealed almost equal number of genes up (143, 49%) or down-regulated (149, 51%). However, at a higher stringency (at least 2-fold), this distribution revealed slightly more leaning towards down-regulated genes (74 out of 130, 57%) indicating the lack of SP-STK results into down regulation of several genes. Amongst the genes belonging to different functional categories, the most notable genes that were affected by the absence of SP-STK were those correspond to Carbohydrate transport and metabolism (57 out of 292, 20% at low stringency and 32 out of 130 ,~25% at high stringency), phage (28, ~10% at low stringency and 24, ~18% at high stringency), and virulence (5-6% irrespective of stringency level). Almost 20-25% of differentially expressed genes revealed at two stringencies were found to be either with unknown or undefined functions. Keywords: knockout mutant
2013-02-28 | GSE10914 | GEO
Project description:In order to characterize the changes in global gene expression in the distal colon of constipated SD rats in response to the laxative effects induced by aqueous extract of Liriope platyphylla (AEtLP) including isoflavone, saponin, oligosaccharide, succinic acid and hydroxyproline, total RNA extracted from the distal colon of AEtLP-treated constipation rats was hybridized to oligonucleotide microarrays.Overall, 581 genes were up-regulated and 216 genes were down-regulated by constipation induced by loperamide, while 67 genes were up-regulated and 421 genes were down-regulated by AEtLP treatment in constipated rats compared to controls. Among the transcripts up-regulated by constipation, 89 were significantly down-regulated and 20 were recovered to normal levels by AEtLP treatment. The major genes in the down-regulated categories included Slc9a5, klk10, Fgf15 and Alpi, while the major genes in the recovered categories were Cyp2b2, Ace, G6pc and Setbp1. However, nine of these genes that were down-regulated by constipation were significantly up-regulated and four were recovered to normal levels by AEtLP treatment. The major genes in the up-regulated categories included Serpina3n, Lcn2 and Slc5a81, while the major genes in the recovered categories were Tmem45a, Rerg and Rgc32.
2014-10-04 | GSE62041 | GEO
Project description:This analysis revealed ~17.% of all ORFs (292 out of a total 1697 ORFs) differed by at least 1.5 fold (Log2 +0.6) between the mutant and wild-type strains. Of these, ~45% of the ORFs (130 out of 292 ORFs) revealed at least 2-fold (Log2 +1) differences. The complete spectrum of differentially expressed genes in the muatnt strain revealed almost equal number of genes up (143, 49%) or down-regulated (149, 51%). However, at a higher stringency (at least 2-fold), this distribution revealed slightly more leaning towards down-regulated genes (74 out of 130, 57%) indicating the lack of SP-STK results into down regulation of several genes. Amongst the genes belonging to different functional categories, the most notable genes that were affected by the absence of SP-STK were those correspond to Carbohydrate transport and metabolism (57 out of 292, 20% at low stringency and 32 out of 130 ,~25% at high stringency), phage (28, ~10% at low stringency and 24, ~18% at high stringency), and virulence (5-6% irrespective of stringency level). Almost 20-25% of differentially expressed genes revealed at two stringencies were found to be either with unknown or undefined functions. Keywords: knockout mutant This analysis is based on a total of three independent prearations of total RNA from the wild-type S. pyogenes M1SF370 and the isogenic knockout mutant lacking SP-STK (SPy1625). First two preparations included dyeswap experiment and hence repeated twice (exp1-4). The third preparation done only one (Exp#5). Thus, the present analysis is based on five separate experiments
2013-02-28 | E-GEOD-10914 | biostudies-arrayexpress
Project description:The transcriptional regulator Mga of Streptococcus pyogenes (the group A streptococcus, GAS) is known to directly activate several virulence genes important for colonization and immune evasion during exponential growth. Transcriptome analysis comparing two mga-1 serotypes (M1 SF370, M6 JRS4) and one mga-2 serotype (M4 GA40634) against their isogenic mga-inactivated strains uncovered a broader Mga regulon profile containing both activated and repressed genes with predicted functions primarily related to the uptake and metabolism of sugars. Although the divergent M1 and M4 Mga profiles were similar in size and content, the M6 JRS4 strain was clearly distinct, even from other M6 strains. Real-time RT-PCR and northern blot analysis validated our microarray results and confirmed that established core Mga regulon genes directly activated by Mga (emm, scpA, sof, fba) exhibited the highest activation levels across all strains tested. A novel ORF (Spy2036) encoding a cytosolic hypothetical protein was highly activated in all three serotypes and was called gene regulated by Mga or grm. Mga was shown to bind directly to Pgrm, which overlaps the Mga-regulated Psof in OF+ strains, suggesting that grm is part of the core Mga regulon and is able to activate two divergently transcribed genes from a single site in a class II background. Both class and serotype specific Mga-regulated genes, such as speB, were apparent. In fact, Mga activated speB as long as it was expressed in the wild type strain, although direct binding of Mga to the PspeB promoter could not be demonstrated. Thus, Mga is able to both directly and indirectly regulate genes shown to be important for virulence and the metabolic homeostasis of GAS. Keywords: Wild-type vs Mga-
2006-08-31 | GSE4596 | GEO