Zinc is an essential nutrient for microbial

Zinc is an essential nutrient for microbial growth, but can be toxic in excess (Blencowe and Morby, 2003; Coleman, 1998). The delicate balance between zinc sufficiency and toxicity is maintained by bacterial metal homeostasis systems (Klaus, 2005; Moore and Helmann, 2005). The GAS metalloregulator, adhesin competence repressor (AdcR), regulates zinc homeostasis by monitoring the intracellular zinc concentration and modulating GAS adaptive response to zinc limitation (Sanson et al., 2015). During zinc sufficiency, the zinc-bound AdcR represses the expression of target genes, whereas the apo-AdcR relieves the repression during zinc deficiency. When purchase AKT inhibitor VIII encounter zinc limitation, GAS upregulates genes encoding zinc acquisition systems (adcA, adcAII/lmb, adcBC, phtD, and phtY), and zinc sparing responses (rpsN.2 and adh) (Sanson et al., 2015). The primary GAS zinc uptake system, AdcABC, is composed of a cell surface-exposed zinc-binding protein (AdcA), an inner membrane permease (AdcB), and a cytosolic ATPase (AdcC) that provides the energy for zinc import by ATP hydrolysis. GAS also employs additional factors such as AdcAII/Lmb, PhtD, and PhtY to overcome zinc limitation (Sanson et al., 2015). Both AdcA and AdcAII share homologous N-terminal ZnuA-like extracellular zinc-binding domains. However, AdcA has an additional C-terminal ZinT-like domain and the role of the ZinT-like domain of AdcA in GAS homeostasis is yet to be elucidated. Although AdcAII/Lmb and Pht proteins contribute to streptococcal adaptive responses to zinc limitation (Bayle et al., 2011; Moulin et al., 2016; Tedde et al., 2016), the exact mechanisms by which they facilitate zinc acquisition remain unknown. Nevertheless, the significance of the AdcR signaling pathway to GAS pathogenesis is underscored by the observation that inactivation of adcR caused dysregulation of zinc homeostasis and significantly attenuated GAS virulence (Sanson et al., 2015). The host recruits calprotectin (CP), an S100A8/A9 heterodimer, at microbial colonization surfaces and inhibits bacterial proliferation by sequestration of zinc, and manganese (Corbin et al., 2008; Damo et al., 2013; Diaz-Ochoa et al., 2016; Liu et al., 2012; Lusitani et al., 2003). The antibacterial activity of CP against bacterial and fungal pathogens has been demonstrated (Corbin et al., 2008; Damo et al., 2013; Diaz-Ochoa et al., 2016; Liu et al., 2012; Sohnle, et al., 1991; Urban, et al., 2009), but its role against most of the streptococcal pathogens is unknown. Emerging evidence indicates that GAS encounters host-mediated zinc immune mechanisms (Brenot et al., 2007; Ong et al., 2014). However, molecular details underlying host defense mechanisms, bacterial countermeasures, and their role in GAS pathogenesis are lacking. Using a multidisciplinary approach, we discovered that CP is a major host defense factor against GAS infection in different niches and mediates growth inhibition primarily by zinc sequestration. Conversely, GAS employs a high-affinity zinc uptake system and a refined sensory system to overcome CP-mediated growth inhibition. To realize the translational potential of our findings, we assessed and validated the extracellular component of the zinc importer, AdcA, as a potential GAS vaccine candidate.
Materials and Methods
Results
Discussion Molecular dialog between host and invading pathogens shapes the course of infection and often determines the disease outcome. We identified a previously unknown GAS-host interaction that occurs during infection and demonstrated that zinc is at the forefront of the host-GAS battle. Furthermore, we showed that the molecular arsenals of the pathogen involved in this host-pathogen conflict could be effectively targeted for disease prevention. Herein, we report that GAS-infected lesions are enriched with CP in two different host compartments and CP retards GAS growth by scavenging zinc from the colonization interface. Our results are consistent with the observations made in other bacterial pathogens including S. aureus, A. baumannii, S. typhimurium, and H. pylori (Gaddy et al., 2014; Hood et al., 2012; Corbin et al., 2008, Damo et al., 2013; Liu et al., 2012). In addition to zinc, CP also inhibits bacterial growth by sequestering manganese and iron (Damo et al., 2013; Diaz-Ochoa et al., 2016; Nakashige et al., 2015). Given that GAS adaptive responses to manganese limitation are not fully understood, the role of CP-mediated manganese sequestration on GAS pathogenesis is yet to be elucidated. Similarly, CP binds iron and causes bacterial growth inhibition in vitro (Nakashige et al., 2015), however, the in vivo significance of CP-mediated iron limitation as a host defense remains unclear (Garcia et al., 2017).