Candida albicans Adhesins Als1 and Hwp1 Modulate Interactions with Streptococcus mutans.

Candida albicans and Streptococcus mutans are known to synergistically interact with each other in the oral cavity. For example, glucosyltransferase B (GtfB), secreted by S. mutans, can bind to the C. albicans cell surface, promoting dual-species biofilm formation. However, the fungal factors mediating interactions with S. mutans are unknown. The C. albicans adhesins Als1, Als3, and Hwp1 are key players in C. albicans single-species biofilm formation, but their roles, if any, in interacting with S. mutans have not been assessed. Here, we investigated the roles of the C. albicans cell wall adhesins Als1, Als3, and Hwp1 on forming dual-species biofilms with S. mutans. We assessed the abilities of the C. albicans wild-type als1Δ/Δ, als3Δ/Δ, als1Δ/Δ/als3Δ/Δ, and hwp1Δ/Δ strains to form dual-species biofilms with S. mutans by measuring optical density, metabolic activity, cell enumeration, biomass, thickness, and architecture of the biofilms. We observed that the C. albicans wild-type strain formed enhanced dual-species biofilms in the presence of S. mutans in these different biofilm assays, confirming that C. albicans and S. mutans synergistically interact in the context of biofilms. Our results reveal that C. albicans Als1 and Hwp1 are major players in interacting with S. mutans, since dual-species biofilm formation was not enhanced when the als1Δ/Δ or hwp1Δ/Δ strains were cultured with S. mutans in dual-species biofilms. Als3, however, does not seem to play a clear role in interacting with S. mutans in dual-species biofilm formation. Overall, our data suggest that the C. albicans adhesins Als1 and Hwp1 function to modulate interactions with S. mutans and could be potential targets for future therapeutics.

Divide and conquer: the Pseudomonas aeruginosa two-component hybrid SagS enables biofilm formation and recalcitrance of biofilm cells to antimicrobial agents via distinct regulatory circuits.

The opportunistic pathogen Pseudomonas aeruginosa forms antimicrobial resistant biofilms through sequential steps requiring several two-component regulatory systems. The sensor-regulator hybrid SagS plays a central role in biofilm development by enabling the switch from the planktonic to the biofilm mode of growth, and by facilitating the transition of biofilm cells to a highly tolerant state. However, the mechanism by which SagS accomplishes both functions is unknown. SagS harbours a periplasmic sensory HmsP, and phosphorelay HisKA and Rec domains. SagS domain was used as constructs and site-directed mutagenesis to elucidate how SagS performs its dual functions. It was demonstrated that HisKA-Rec and the phospho-signalling between SagS and BfiS contribute to the switch to the biofilm mode of growth, but not to the tolerant state. Instead, expression of SagS domain constructs harbouring HmsP rendered ΔsagS biofilm cells as recalcitrant to antimicrobial agents as wild-type biofilms, likely by restoring BrlR production and cellular c-di-GMP levels to wild-type levels. Restoration of biofilm tolerance by HmsP was independent of biofilm biomass accumulation, RsmA, RsmYZ, HptB and BfiSR-downstream targets. Our findings thus suggest that SagS likely makes use of a "divide-and-conquer" mechanism to regulate its dual switch function, by activating two distinct regulatory networks via its individual domains.

Doxorubicin and 5-fluorouracil induced accumulation and transcriptional activity of p53 are independent of the phosphorylation at serine 15 in MCF-7 breast cancer cells.

The chemotherapeutic agents doxorubicin (dox) or 5-fluorouracil (5FU) are used to treat cancer cells as they cause irreparable DNA damage, inducing these aberrant cells to undergo cell death. The mediator of this process is presumed to be in part the tumor suppressor p53 which regulates genes involved in DNA repair and cell death. When MCF-7 breast cancer cells are treated with these drugs, we observed that the level of p53 and the p53 negative regulator, Mdm2, increased, as seen by others. But contrary to some reports, we observed minimal phosphorylation of p53 at serine 15 in MCF-7 cells after drug treatment. Interestingly, we determined that there was differential regulation of the kinases ATM and Chk2 with the drug treatments, likely the cause for the lack of phosphorylation of p53. We found a dramatic drop in p53 DNA binding affinity for p21 and other gene response elements (RE) after drug treatment. To determine if the p53 that accumulated in the drug treated cells was functionally active, we monitored changes in the protein products of two p53-regulated genes following drug treatment with and without the addition of a p53-specific siRNA. In response to 5FU, both p21 and Mdm2 proteins increased and that increase was alleviated if a p53-specific siRNA was added. This effect was not seen with the addition of dox. Thus, the phosphorylation at serine 15 is not necessary for the functional activation of this transcription factor. We propose a new model for the regulation of p53, Mdm2, and MdmX after drug treatment.