Clp ATPases differentially affect natural competence development in Streptococcus mutans

Abstract

In naturally competent bacteria, DNA transformation through horizontal gene transfer is an evolutionary mechanism to receive extracellular DNA. Bacteria need to maintain a state of competence to accept foreign DNA, and this is an energy-driven phenomenon that is tightly controlled. In Streptococcus, competence development is a complex process that is not fully understood. In this study, we used Streptococcus mutans, an oral bacterium, to determine how cell density affects competence development. We found that in S. mutans the transformation efficiency is maximum when the transforming DNA was added at low cell density and incubated for 2.5 h before selecting for transformants. We also found that S. mutans cells remain competent until the mid-logarithmic phase, after which the competence decreases drastically. Surprisingly, we observed that individual components of Clp proteolytic complexes differentially regulate competence. If the transformation is carried out at the early growth phase, both ClpP protease and ClpX ATPase are needed for competence. In contrast, we found that both ClpC and ClpE negatively affect competence. We also found that if the transformation is carried out at the mid-logarithmic growth phase ClpX is still required for competence, but ClpP negatively affects competence. While the exact reason for this differential effect of ClpP and ClpX on transformation is currently unknown, we found that both ClpC and ClpE have a negative effect on transformation, which was not reported before.

Diverse nature of ClpX degradation motifs in Streptococcus mutans

“…While the amount of the proteolytic component ClpP is inducible by various cellular stresses, the amount of the ATPase component, ClpX, is relatively stable throughout the growth ( 12 ). Consistent with ClpX/P’s key role in regulated proteolysis, inactivation, or functional loss of either ClpX or ClpP leads to pleiotropic effects, including growth defects, aberrant biofilm formation, reduced bacteriocin production, improper natural competence development, and others physiological effects ( 31 , 47 , 48 ). While the importance of ClpX/P in cellular physiology is unraveling, the identity of the substrates targeted by this proteolytic complex is largely unknown in S. mutans and other streptococci.…”

Synthesis of a Novel Lantibiotic Using Mutacin II Biosynthesis Apparatus

“…When necessary, 5 μg/mL erythromycin was included in the THY medium. All of the streptococcal strains were transformed by means of natural transformation according to standard protocols, with the addition of competence-stimulating peptides ( 52 , 53 ).…”

Mass spectrometry and split luciferase complementation assays reveal the MecA protein interactome of Streptococcus mutans

“…For this reason, such proteins exhibit extremely diverse protein interactomes, the vast majority of which remain poorly characterized in most bacteria. In the oral pathobiont Streptococcus mutans , these functions are largely performed by the Clp protease system, which consists of a ClpP protease subunit paired with any of several associated Clp ATPases: ClpC, ClpE, and ClpX ( 1 – 5 ). Of these three Clp ATPases, ClpC is unique in its requirement for an additional adaptor protein called MecA to stimulate proteolysis by ClpCP complexes ( 6 , 7 ).…”