c‐di‐AMP reports DNA integrity during sporulation in Bacillus subtilis

Abstract

The bacterium Bacillus subtilis produces the DNA integrity scanning protein (DisA), a checkpoint protein that delays sporulation in response to DNA damage. DisA scans the chromosome and pauses at sites of DNA lesions. Structural analysis showed that DisA synthesizes the small molecule cyclic diadenosine monophosphate (c-di-AMP). Here, we demonstrate that the intracellular concentration of c-di-AMP rises markedly at the onset of sporulation in a DisA-dependent manner. Furthermore, exposing sporulating cells to DNA-damaging agents leads to a global decrease in the level of this molecule. This drop was associated with stalled DisA complexes that halt c-di-AMP production and with increased levels of the c-di-AMP-degrading enzyme YybT. Reduced c-di-AMP levels cause a delay in sporulation that can be reversed by external supplementation of the molecule. Thus, c-di-AMP acts as a secondary messenger, coupling DNA integrity with progression of sporulation.

Cyclic Di-AMP Homeostasis in Bacillus subtilis

“…DisA scans the DNA and stops c-di-AMP production if it encounters problems with DNA integrity such as Holliday junctions. The reduced DisA-mediated c-di-AMP synthesis results in a delay of sporulation (8,38). This specific function is rather unlikely to be involved in the essential role of c-di-AMP.…”

“…The genome of B. subtilis encodes three proteins with a DAC domain that may be potential diadenylate cyclases, DisA, CdaA, and CdaS (5, 13). The diadenylate cyclase activity of DisA has been demonstrated experimentally (7,8). disA is the promoter-distal gene of the hexacistronic ctsR-mcsA-mcsB-clpC-radA-disA operon.…”

“…If the protein arrives at branched DNA molecules present in Holliday junctions, then the catalytic activity is inhibited, and this reduction in c-di-AMP concentration results in the delay of sporulation (7,8). DisA contains a catalytic domain, called the diadenylate cyclase (DAC) domain (previously referred to as domain of unknown function, DUF147) (7).…”

Diadenylate cyclase evaluation of ssDacA (SSU98_1483) in Streptococcus suis serotype 2

“…Cyclic diadenosine monophosphate (c-di-AMP) was recently discovered and recognized as a signaling molecule utilized by many bacteria, including bacterial pathogens during infection. In Bacillus subtilis, bacterial c-di-AMP levels are reduced in response to DNA damage, resulting in delayed sporulation (Oppenheimer-Shaanan et al, 2011). High c-di-AMP levels result in smaller bacterial size in Staphylococcus aureus (Corrigan et al, 2011), c-di-AMP secreted by Listeria monocytogenes triggers a cytosolic pathway for innate immunity (Woodward et al, 2010), and c-di-AMP homeostasis is required for the virulence of Streptococcus pyogenes and Streptococcus pneumonia (Bai et al, 2013;Cho and Kang, 2013;Witte et al, 2013).…”

Systematic identification of conserved bacterial c-di-AMP receptor proteins

“…c-di-AMP is synthesized by DisA_N domain-containing diadenylate cyclases DacA, DisA, and YojJ and degraded by the phosphodiesterase enzyme GdpP (4,5,11,(15)(16)(17)(18). A variety of different phenotypes have been linked to altered c-di-AMP levels; an increase in c-di-AMP levels correlates with increased acid resistance (16,19) and altered antibiotic resistance, including an increase in methicillin resistance in S. aureus (11,18,20).…”