Comment[ArrayExpressAccession]	E-GEOD-60565
MAGE-TAB Version	1.1						
Public Release Date	2014-08-21
Investigation Title	Comparison of Expression Profiles for Agrobacterium tumefaciens visR mutant to wild-type strain.						
Comment[Submitted Name]	Comparison of Expression Profiles for Agrobacterium tumefaciens visR mutant to wild-type strain.
Experiment Description	Many bacteria colonize surfaces and transition to a sessile mode of growth. The plant pathogen Agrobacterium tumefaciens produces a unipolar polysaccharide (UPP) adhesin at single cell poles that contact surfaces. Here we report that elevated levels of the intracellular signal cyclic diguanosine monophosphate (c-di-GMP) lead to surface-contact-independent UPP production and a red colony phenotype due to production of UPP and the exopolysaccharide cellulose, when A. tumefaciens is incubated with the polysaccharide stain Congo Red. Transposon mutations with elevated Congo Red staining identified presumptive UPP-negative regulators, mutants for which were hyperadherent, producing UPP irrespective of surface contact. Multiple independent mutations were obtained in visN and visR, activators of flagellar motility in A. tumefaciens, now found to inhibit UPP and cellulose production. Expression analysis in a visR mutant and isolation of suppressor mutations, identified three diguanylate cyclases inhibited by VisR. Null mutations for two of these genes decrease attachment and UPP production, but do not alter cellular c-di-GMP levels. However, analysis of catalytic site mutants revealed their GGDEF motifs are required to increase UPP production and surface attachment. Mutations in a specific presumptive c-di-GMP phosphodiesterase also elevate UPP production and attachment, consistent with c-di-GMP activation of surface-dependent adhesin deployment. Three biological replicates, independent RNA preparations, one dye swap.						
Term Source Name	ArrayExpress	EFO
Term Source File	http://www.ebi.ac.uk/arrayexpress/	http://www.ebi.ac.uk/efo/efo.owl					
Person Last Name	Heindl	Xu	Kim	Koestler	Choi	Waters	Fuqua
Person First Name	Jason	Jing	Jinwoo	Benjamin	Jeong-Hyeon	Christopher	Clay
Person Mid Initials	Eugene			J		M	
Person Email	jheindl@indiana.edu						
Person Affiliation	Indiana University						
Person Phone	812-856-5186						
Person Address	Biology, Indiana University, 1001 E. 3rd St., Jordan Hall 142, Bloomington, IN, USA						
Person Roles	submitter						
Protocol Name	P-GSE60565-1	P-GSE60565-2	P-GSE60565-5	P-GSE60565-6	P-GSE60565-3	P-GSE60565-4	P-GSE60565-7
Protocol Description	Limma Software (version 3.6.9) from Bioconductor (version 2.7) was used for background subtraction, Loess within arrays normalization  and Quantile between arrays normalization. ID_REF =  VALUE = normalized M values, log2 (visR/WT)	Limma Software (version 3.6.9) from Bioconductor (version 2.7) was used for background subtraction, Loess within arrays normalization  and Quantile between arrays normalization. ID_REF =  VALUE = normalized M values, log2 (visR/WT) INV_VALUE = normalized M values, log2 (WT/visR)	Labeled cDNA with Invitrogen SuperScript Indirect Labeling Kit, cDNA was purified on Qiagen QIAQuick columns. cDNA was labeled with AlexaFluor 555 and 647 dyes using Invitrogen SuperScript cDNA Labeling Kit	Hybridization reactions were performed using Agilent in situ Hybridization Kit Plus, boiled 5 min 95 C, applied to Agilent custom arrays for A. tumefaciens C58, and hybridized overnight at 65 C. Hybridized arrays were washed with Agilent Wash Solutions 1 and 2, rinsed with acetonitrile, and incubated in Agilent Stabilization and Drying Solution immediately prior to scanning the arrays	Grown w/shaking in ATGN minimal medium, 28 °C, mid-log phase cultures	11 mL of cultures at OD600~0.6. Cells lysed with lysozyme and RNA extracted using Qiagen RNease Kit.	Scanned on an GenePix 4200A 01 scanner. Images were quantified using GenePix Pro (version 6.0.1.25.)
Protocol Type	normalization data transformation protocol	normalization data transformation protocol	labelling protocol	hybridization protocol	growth protocol	nucleic acid extraction protocol	array scanning protocol
Experimental Factor Name	GENOTYPE						
Experimental Factor Type	genotype						
Publication Title	Genetic analysis of Agrobacterium tumefaciens unipolar polysaccharide production reveals complex integrated control of the motile-to-sessile switch.						
Publication Author List	Xu J, Kim J, Koestler BJ, Choi JH, Waters CM, Fuqua C						
PubMed ID	23829710						
Publication DOI	10.1111/mmi.12321						
Comment[SecondaryAccession]	GSE60565						
Comment[GEOReleaseDate]	2014-08-21						
Comment[ArrayExpressSubmissionDate]	2014-08-20						
Comment[GEOLastUpdateDate]	2014-08-21						
Comment[AEExperimentType]	transcription profiling by array						
Comment[AdditionalFile:Data1]	GSE60565_VisR_WT_Microarray_Data_with_gene_names_and_pvalues.txt						
SDRF File	E-GEOD-60565.sdrf.txt