Phototaxis and impaired motility in adenylyl cyclase and cyclase receptor protein mutants of Synechocystis sp. strain PCC 6803

Abstract

We have carefully characterized and reexamined the motility and phototactic responses of Synechocystis sp. adenylyl cyclase (Cya1) and catabolite activator protein (SYCRP1) mutants to different light regimens, glucose, 3-(3,4-dichlorophenyl)-1,1-dimethylurea, and cyclic AMP. We find that contrary to earlier reports, cya1 and sycrp1 mutants are motile and phototactic but are impaired in one particular phase of phototaxis in comparison with wild-type Synechocystis sp.

FIG. 1.

Phases of phototaxis in Synechocystis sp. cells. Cells were spotted on motility plates, placed in unidirectional white light, and photographed. After overnight growth, cells are either single or in small groups; the faint green edge visible at the front of the spot is caused by cells that have moved and accumulated at the front edge (A and D); after 2 days (B and E), many more cells are in groups and have migrated to the front of the spot, typical of phase 2; after 4 days (C), fingerlike projections of motile cells are seen, typical of phase 3. Lower panels: single cells (D) and cell groups (E) from spots in panels A and B are shown at higher magnification. Motility plates contained 0.4% agarose in BG-11 and 10 mM glucose. The arrow shows the direction of the light. Spots are ∼3 to 4 mm in diameter. Bar, 10 μm.

FIG. 2.

Effect of cAMP on phototaxis of Synechocystis sp. WT and cya1 and sycrp1 mutant cells. Upper panels: WT (left), cya1 mutant (middle), and sycrp1 mutant (right) cells were spotted on 0.4% agarose BG-11 motility (no glucose added) plates in the absence (A) or presence (B) of 0.1 mM cAMP and photographed after 4 days. Lower panels: the front edge of the spot containing cya1 mutant cells is shown after day 2 (C) and day 4 (D). Note that cells are evenly distributed at day 2, but after day 4, cells have clearly accumulated at the front. In the presence of 0.1 mM cAMP (B), cya1 mutant cells shows the typical fingerlike projections seen in WT cells; sycrp1 mutant cells do not respond to cAMP. The arrow shows the direction of the light. Spots are ∼3 to 4 mm in diameter. Bar, 10 μm.

FIG. 3.

Glucose effect on motility of Synechocystis sp. WT and cya1 and sycrp1 mutant cells. WT (left), cya1 mutant (middle), and sycrp1 mutant (right) cells were spotted on 0.4% agarose motility plates containing no glucose (A) or 0.1 mM (B), 0.2 mM (C), 1 mM (D), 5 mM (E), or 10 mM (F) glucose. Plates were placed in directional white light and photographed after 4 days. Note that cya1 mutant cells show slight formation of fingerlike projections in 10 mM glucose, but sycrp1 mutant cells do not.

FIG. 4.

Motility in the presence of glucose and DCMU. Synechocystis sp. WT cells were spotted on 0.4% agarose BG-11 plates (A) or plates containing 10 μM DCMU (B), 10 mM glucose (C), or 10 mM glucose and 10 μM DCMU (D). Spots were photographed after 4 days. After 4 days the cells growing in DCMU have not divided and cannot be easily seen at this magnification. Spot diameter is ∼4 mm.

FIG. 5.

Motility of Synechocystis sp. WT and cya1 and sycrp1 mutant cells in different light qualities. WT (left), cya1 mutant (middle), and sycrp1 mutant (right) cells were spotted on 0.4% agarose BG-11-10 mM glucose motility plates in the absence (top panels) or the presence (bottom panels) of 0.1 mM cAMP. Plates were placed in different light sources (white, red, green, or blue) and photographed after 4 days. Spot diameter is ∼4 mm.

FIG. 6.

Representative examples of movement tracks of WT cells and cya1 mutants in white light. WT cells (left) and cya1 mutant cells (middle) were spotted on 0.4% agarose BG-11-10 mM glucose motility plates under directional white light for 24 h. We identified single cells (since groups of cells are not possible to track), tracked cell movements for 30 min by time-lapse video microscopy, and quantified them with Metamorph tracking software. A composite made up of randomly selected individual movement tracks was copied and combined into a single figure. The direction of the white light source is shown by an arrow. Bar, 100 μm. The panel on the right shows the path length for a particular track from start to finish and the displacement which is calculated from the start to the finish points.