TABLE 1 .
Verification that selected genes were σ22 dependent by using a lacZ fusion analysis of each promoter in algT (σ22-deficient) and mucA (anti-sigma-factor-deficient) backgrounds
| Gene | Gene product descriptiona | Stress induction (15/60 min)b | P-lacZ WT:algT:mucA activityc |
|---|---|---|---|
| PA0059 | OsmC, redox protein, osmotically induced | 9.9/8.9 | 156:15:450 |
| PA0062 | Hypothetical, predicted type II lipoprotein | 11.4/10.5 | ND |
| PA0460 | Hypothetical, predicted periplasmic | 18.9/7.7 | 1,569:116:5,161 |
| PA0567d | Proteolipid homolog, membrane integrity | 6.0/6.2 | 220:142:976 |
| PA0854 | FumC2, fumarate hydrase | 4.7/10.7 | ND |
| PA0919 | Hypothetical, predicted secretion usher | 3.0/<2 | Operon |
| PA0920d | Aminoacyl-phosphatidylglycerol synthase | 3.0/<2 | 3,264:2,825:5,239 |
| PA1243 | Predicted transcriptional regulator, PAS domain | 2.0/<2 | 87:30:307 |
| PA1323 | Hypothetical, DUF883 family | 19.8/15.9 | 269:13:1,278 |
| PA1324 | Predicted to bind/transport polysaccharides | 19.1/14.3 | Operon |
| PA2167 | Hypothetical, unclassified | <2/20.7 | 72:21:470 |
| PA2176 | Hypothetical, unclassified | <2/7.0 | 76:19:296 |
| PA2177d | Predicted sensor response hybrid, PAS domain | 2.2/<2 | 99:101:244 |
| PA2717 | Cpo, chloroperoxidase | <2/14.4 | ND |
| PA3040d | Hypothetical, DUF883 family | 7.1/6.2 | 1,352:1,073:4,148 |
| PA3459d | Predicted glutamine amidotransferase | 4.4/<2 | 402:415:839 |
| PA3691 | Hypothetical, predicted lipoprotein | 15.7/17.8 | 608:13:2,216 |
| PA3795d | Predicted oxidoreductase | 5.2/8.8 | 707:605:1,593 |
| PA3819 | SlyB homolog, outer membrane protein | 10.3/8.2 | 1,221:568:3,351 |
| PA4311 | Predicted glycosyltransferase | 4.0/6.9 | ND |
| PA4394 | Predicted mechanosensitive channel, McsS | 3.0/4.0 | 130:15:797 |
| PA4717d | Predicted periplasmic metalloprotease | 5.3/6.3 | 1,404:1,101:3,215 |
| PA5107 | Lipocalin Blc, outer membrane protein | 8.1/4.4 | Operon |
| PA5108 | Hypothetical, predicted lipoprotein | 7.5/3.0 | 714:387:3,732 |
| PA5178 | Hypothetical, LysM domain | 9.6/7.1 | 452:284:2,532 |
| PA5212 | Hypothetical, unclassified | 15.9/7.8 | 1,555:44:5,758 |
| PA5424d | Hypothetical, predicted inner membrane protein | 8.3/7.4 | 807:660:3,904 |
PAO1 gene names and descriptions were obtained from the Pseudomonas Genome Database (33) and are listed numerically. Sequence-defined transposon insertions in each gene were purchased from the University of Washington Genome Center. All were transduced into the PAO1/PDO1 reference isolate and verified by PCR analysis.
Stress induction shows data previously described (19) for the fold increase of the genes’ transcriptional activity expressed from the PAO1 chromosome when treated with d-cycloserine (400 µg/ml) for 15 or 60 min as determined by microarray analysis.
PAO1 (wild type [WT]), PDO-LS586 (algT), and PDO351 (mucA) strains containing each lacZ fusion plasmid were grown in L broth at 37°C with aeration, collected during logarithmic growth, and assayed for β-galactosidase activity. Data show each gene’s promoter activity (Miller units) when fused to lacZ (P-lacZ) when expressed in the wild type or in an algT or mucA mutant. “Operon” indicates that the gene is in an operon of a gene already tested. ND, not determined.
The mutant did not show the predicted σ22 stimulon phenotype until tested under cell wall stress conditions (see Table 2).