FIG 8 .

Model for the integration of environmental calcium signaling and PAPS production in the regulation of brp expression. BrpT and c-di-GMP bias the sulfate assimilation pathway toward the accumulation of PAPS in a feed-forward regulatory network (framed in green). Points of activation and repression are indicated by solid blue arrowheads and flat lines, respectively. Environmental calcium stimulates sulfate uptake. The sulfate is converted to adenosine 5′-phosphosulfate (APS) by the action of CysD and CysN and converted to 3′-phosphoadenosine 5′-phosphosulfate (PAPS) by CysC. The accumulation of PAPS triggers an increase in the intracellular c-di-GMP level, which increases brpT expression via the regulator BrpR and also represses expression of the cysJIH operon that drains the PAPS pool by processing it to PAP and sulfite and, eventually, to cysteine. BrpT activates expression of the brp exopolysaccharide, cab matrix genes, and cysDNTC operons (blue, purple, and green block arrows, respectively), which generates more PAPS, c-di-GMP, and BrpT, thus establishing a feed-forward signaling network that biases the sulfur metabolism pathway toward accumulating PAPS to increase brp expression and biofilm formation. PAPS also has a separate and distinct impact on rugose colony development. The operon structure of cysDNTC and cysJIH (red block arrow) allows for their simultaneous regulation by c-di-GMP and, for the regulation of cysDNTC, by BrpT. Environmental calcium may also influence cellular c-di-GMP levels independent of sulfate assimilation (dashed blue line). Other environmental cues (e.g., temperature) can directly affect intracellular c-di-GMP concentrations that may then feed into the PAPS–c-di-GMP loop.