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. 2021 Mar 18;10:e66160. doi: 10.7554/eLife.66160

Figure 1. PomX consists of two domains that are both required for function.

(A) Similarity and identity analysis of PomX, PomY, and PomZ homologs. The three Myxococcales suborders are indicated. An open box indicates that a homolog is not present. (B) Similarity and identity of PomX domains in different PomX homologs. Similarity and identity were calculated based on the domains of M. xanthus PomX shown in C. (C) PomX truncations used in this study. Numbers on top indicate the start and stop positions of the truncations relative to full-length PomXWT. (D) Cell length distribution of cells of indicated genotypes. Cells below stippled line are minicells. Numbers indicate mean cell length±STDEV. In the boxplots, boxes include the 25th and the 75th percentile, whiskers data points between the 10% and 90% percentile, outliers are shown as black dots. Black and red lines indicate the median and mean, respectively. Number of analyzed cells is indicated. In the complementation strains, pomX alleles were expressed from plasmids integrated in a single copy at the attB site. (E) Fluorescence microscopy of cells of indicated genotypes. Phase-contrast and fluorescence images of representative cells were overlayed. Numbers indicate fraction of cells with fluorescent clusters. Demographs show fluorescence signals of analyzed cells sorted according to length and with off-center signals to the right. Numbers in upper right indicate number of cells used to create demographs. Scale bar, 5 µm. (F) Fluorescence microscopy of cells of indicated genotypes. Images of representative cells and demographs were created as in (E). Scale bar, 5 µm. For experiments in D, E and F similar results were obtained in two independent experiments.

Figure 1—source data 1. Source Data for Figure 1D.
elife-66160-fig1-data1.xlsx (101.5KB, xlsx)

Figure 1.

Figure 1—figure supplement 1. PomX variants accumulate in M.xanthus.

Figure 1—figure supplement 1.

(A) Schematic of pomXYZ locus (upper panel) and the construct used for ectopic expression of mCh-pomX and its variants from the attB site (lower panel). The brown region upstream of pomZ was used as a promoter for the expression of mCh-pomX variants. All coordinates are relative to the first nucleotide in pomZ start codon (+1). (B) Western blot analysis of mCh-PomX (71.0 kDa), mCh-PomXN (50.2 kDa), and mCh-PomXC (48.7 kDa) accumulation in indicated strains. Protein from the same number of cells was loaded per lane. Molecular mass markers are indicated on the left and analyzed proteins on the right including calculated MW. The same blots were sequentially analyzed with α-PomX (top panel), α-mCh (middle panel), and α-PilC (lower panel). PilC was used as a loading control. Note PomXWT (43.9 kDa) does not migrate at the expected size in SDS-PAGE but as a protein of a molecular weight of ~72 kDa. Similarly, mCh-PomXWT, mCh-PomXN, and mCh-PomXC migrate at ~110 kDa, ~60 kDa, and ~62 kDa, respectively. Note that the three bands labeled * in the right and left α-mCh western blot of (B) are unspecific bands that sometimes appear in the western blots with α-mCh antibodies.
Figure 1—figure supplement 1—source data 1. Source data for Figure 1—figure supplement 1B.