Limited functional conservation of a global regulator among related bacterial genera: Lrp in , and -7

Ng standard errors. At least four points from each of two independent experiments were used to generate each plotted value. Conditions were MOPS-glucose minimal medium, supplemented as described in Methods, in logarithmic (MinLog) or stationary phase (MinSta), or MOPS glucose defined rich medium in those growth phases (RchLog, RchSta). . Direct comparison of mRNA levels.. A baseline amount of total RNA (from a mid-log phase culture in MOPS glucose plus nicotinate) was mixed with varying amounts of test RNA (all from log-phase cultures) from glucose (open circle) or rich (closed circle) cultures. The mixes were used as template for simultaneous amplification with three primer pairs. If the test cDNA preparation has the same proportion of cDNA as the reference pool, the detected amount of cDNA should rise with a slope of 1.0 (actual . detected, based on the varied amounts of test cDNA added); this is shown as a dotted line in each panel.<p><b>Copyright information:</b></p><p>Taken from "Limited functional conservation of a global regulator among related bacterial genera: Lrp in , and "</p><p>http://www.biomedcentral.com/1471-2180/8/60</p><p>BMC Microbiology 2008;8():60-60.</p><p>Published online 11 Apr 2008</p><p>PMCID:PMC2374795.</p><p></p

Wild-type strains of , and were grown in MOPS glucose plus nicotinate or MOPS glucose defined-rich media

The data are from two independent experiments (open and closed symbols). Growth curves (, MOPS glucose medium; , MOPS rich medium) and Lrp protein levels (, glucose; , rich) are shown. Equal amounts of total protein were loaded in each lane, and a standard curve of purified Lrp was included on each gel for quantitation. shows a comparative western blot. Cell pellets were boiled and equal amounts of total protein from (Ec) and (Pm) were resolved side-by-side via SDS polyacrylamide gel electrophoresis. The subsequent blot was probed with polyclonal antiserum raised against Lrp.<p><b>Copyright information:</b></p><p>Taken from "Limited functional conservation of a global regulator among related bacterial genera: Lrp in , and "</p><p>http://www.biomedcentral.com/1471-2180/8/60</p><p>BMC Microbiology 2008;8():60-60.</p><p>Published online 11 Apr 2008</p><p>PMCID:PMC2374795.</p><p></p

The Venn diagram shows subsets of genes that were differentially regulated in response to Lrp orthologs from the indicated species (but not to the vector control)

Gene expression was assessed by two-color microarray analysis as described in Methods. The pie chart represents the relative distribution of genes significantly responsive to the Lrp that are also significantly responsive to the other Lrp orthologs. Details of the statistical analysis of these data are in Methods, and the gene-specific results are available [see Additional file ].<p><b>Copyright information:</b></p><p>Taken from "Limited functional conservation of a global regulator among related bacterial genera: Lrp in , and "</p><p>http://www.biomedcentral.com/1471-2180/8/60</p><p>BMC Microbiology 2008;8():60-60.</p><p>Published online 11 Apr 2008</p><p>PMCID:PMC2374795.</p><p></p

Strains, all carrying 10 and Δ, were transformed with plasmids carrying various alleles (or vector control)

Transformants were grown in unsupplemented MOPS glucose medium. . Western blot analysis of Lrp accumulation (Eco, Lrp; Pmi, Lrp; Vch, Lrp; pCC1, vector control) using polyclonal antiserum raised against Lrp. The arrow indicates the direction of electrophoresis. . P(B), P(C) and P(D) activity were measured via ONPG hydrolysis, and plotted . culture density to ensure that the cultures were in balanced growth. The Lrp orthologs used are from (triangles) and (squares), as well as the positive control (circles) and the vector control (diamonds). . Isoleucine, Leucine and Valine was added to the medium ("+Leu") for experiments depicted in the lower panels: (E), (F) and (G). The correlation coefficients for the least-squares fits to the data were all at least 0.97.<p><b>Copyright information:</b></p><p>Taken from "Limited functional conservation of a global regulator among related bacterial genera: Lrp in , and "</p><p>http://www.biomedcentral.com/1471-2180/8/60</p><p>BMC Microbiology 2008;8():60-60.</p><p>Published online 11 Apr 2008</p><p>PMCID:PMC2374795.</p><p></p

Growth rates were determined from a fit to the exponential portion of the growth curve, extending in all but one case (, glucose minimal medium) through at least four mass doublings

Open symbols refer to growth in MOPS glucose plus required supplements (nicotinate, panthothenate and thiamine; see Methods), while closed symbols represent growth in MOPS glucose defined-rich medium. . growth rates. The values shown are the specific growth rate constants, , calculated as ln2/(doubling time, in h). For comparison, values of 0.5, 1, and 2 correspond respectively to doubling times of 83, 42, and 21 min. The rich medium results are clustered and therefore not labeled; for the minimal medium, the abbreviations used are Eco (), Pmi (), and Vch (). The diagonal line shows where points should fall if mutation has no effect on the growth rate in these media. . Complementation of the low growth rate in the glucose minimal medium described in (A). The dashed lines indicate growth data for the mutant (open circles; 193 min doubling time) and the mutant bearing the vector control (gray circles; 191 min). Remaining lines show the WT (closed circles; 66 min); and the mutant bearing plasmids with the genes from (triangles; 69 min), (squares; 81 min), or (diamonds; 81 min).<p><b>Copyright information:</b></p><p>Taken from "Limited functional conservation of a global regulator among related bacterial genera: Lrp in , and "</p><p>http://www.biomedcentral.com/1471-2180/8/60</p><p>BMC Microbiology 2008;8():60-60.</p><p>Published online 11 Apr 2008</p><p>PMCID:PMC2374795.</p><p></p

In every panel, the x-axis shows the gene expression ratio for 12 Δrelative to that in the same strain complemented by (pEcoLrp)

The y-axes indicate the equivalent ratio, where the complementation is by vector alone (pCC1) or the alleles from (pVchLrp) or (pPmiLrp). Full complementation relative to that by pEcoLrp would yield a slope of 1.0 for the linear fit. . This column shows responses of the gene set yielding statistically-significant increases in expression associated with mutation in , as reflected by an expression ratio significantly above 1.0 on the x-axis. This set includes genes that are repressed (directly or indirectly) by Lrp. . This column includes the set of genes showing significant decreases in expression associated with mutation in , indicating direct or indirect activation by Lrp. . This column shows the set of 57 genes recognized in RegulonDB [71, 72] as being directly controlled by Lrp, whether the control is positive or negative. This set includes genes that are controlled by Lrp, but not under the growth conditions used by us, so the cluster of genes showing little or no effect of Lrp is not surprising [66, 69]. The relative transcript abundances were estimated from at least three independent biological replicas using a linear model similar to one introduced before [141, 142]. Significantly expressed genes were identified at a fixed false discovery rate of 5% at the 90percentile [138]. Details of the statistical analysis of these data are in the text, and a list of the 57 RegulonDB Lrp targets is in the Methods section.<p><b>Copyright information:</b></p><p>Taken from "Limited functional conservation of a global regulator among related bacterial genera: Lrp in , and "</p><p>http://www.biomedcentral.com/1471-2180/8/60</p><p>BMC Microbiology 2008;8():60-60.</p><p>Published online 11 Apr 2008</p><p>PMCID:PMC2374795.</p><p></p