TABLE 3.
Co-immunoprecipitation of FLAG-tagged divisome proteins from cross-linked Spn cellsa.
| Bait Used | Prey | Mean ratiob | Detected prey in complex?c | Strains usedd |
| FLAG-FtsA | FtsZ-Myc | 18.1 ± 10.8 | Yes | IU9713/IU11476 |
| EzrA-HA | 223 ± 49 | Yes | ||
| DivIVA-FLAG3 | FtsZ-Myc | 1.5 ± 0.5 | Marginal | IU9713/IU11414 |
| EzrA-HA | 1.8 ± 0.5 | Marginal | ||
| MapZ-FLAG3 | FtsZ-Myc | 1.6 ± 0.5 | Marginal | IU9713/IU11430 |
| EzrA-HA | 6.4 ± 0.5 | Yes | ||
| FtsK-FLAG2 | FtsZ-Myc | 1.5 ± 0.5 | Marginal | IU9713/IU11664 |
| EzrA-HA | 4.3 ± 0.7 | Yes | ||
| ZapA-FLAG | EzrA-HA | 8.2 ± 1.5 | Yes | IU11939/IU11840 |
| FtsZ | 14.2 ± 3.4 | Yes | ||
| FtsA | 2.2 ± 0.1 | Yes | ||
| aPBP1a-FLAG3 | EzrA-HA | 3.6 ± 1.9 | Yes | IU6810/IU12069 |
| FtsZ | 1.3 ± 0.2 | No | ||
| FtsA | 1.2 ± 0.2 | No | ||
| MreC | 4.6 ± 2.5 | Yes | ||
| SepF-FLAG | EzrA-HA | 15.9 ± 6.2 | Yes | IU6810/IU12076 |
| FtsZ | 1.1 ± 0.2 | No | ||
| FtsA | 13.3 ± 2 | Yes | ||
| StkP-FLAG2 | EzrA-HA | 13.9 ± 2.3 | Yes | IU6810/IU12077 |
| FtsZ | 1.2 ± 0.1 | No | ||
| FtsA | 1.7 ± 0.3 | Marginal | ||
| bPBP2x-FLAG3 | EzrA-HA | 1.8 ± 0.6 | Marginal | IU6810/IU11880 |
| FtsZ | 1.1 ± 0.0 | No | ||
| FtsA | 1.7 ± 0.2 | Marginal | ||
| FtsZ-FLAG | ZapA-HA | 5.9 ± 3.4 | Yes | IU10267/IU11322 |
| FtsA | 5.7 ± 1.0 | Yes | ||
| FtsZ | 173 ± 79 | Yes |
aCo-IP experiments were performed as described in section “Materials and Methods.”
bThe mean ratio is determined by dividing Western blot ROI signals of epitope tagged prey in strain expressing bait+ by the ROI signal of epitope tagged prey in strain expressing bait-FLAG3 ([ROI prey-HA]bait+/[ROI prey-HA]bait–FLAG3). ROI values are determined as described in Materials and Methods. ±, SEM from two independent biological replicates.
cA positive association was determined based on the mean ratio value greater than 2.0. A mean value between 1.5 and 2.0 was labeled “marginally detected” association because prey proteins were indeed detected in the bait complex, although not “high” relative to prey protein in non-bait complex.
dComplete genotypes are listed in Supplementary Table 1. Strain numbers are listed in the following order: the strain listed first expresses the epitope-tagged prey and untagged/no bait FtsA (i.e., IU9713; ftsZ-Myc ezrA-HA ftsA+), while the second strain listed contains the epitope tagged prey and a FLAG tagged bait protein (i.e., IU11476 ftsZ-Myc ezrA-HA FLAG-ftsA). All strains used here showed wild-type cell morphology and growth characteristics, with the exception of IU11430 that showed some slight cell size variability with rounder cells.