D-085. The Sap Transporter Is Critical to Survival Strategies by Nontypeable Haemophilus influenzae (NTHi)
Nontypeable Haemophilus influenzae (NTHi) is an opportunistic pathogen of both the upper and lower respiratory tracts. We have previously shown that the sap (sensitivity to antimicrobial peptides) operon gene products mediate NTHi resistance to antimicrobial peptides (APs), in part by AP interaction with the SapA binding protein (SBP) and subsequent up-regulation of sap gene transcription. We further show that the putative Sap transporter ATPase protein, SapD, is required for AP resistance and potassium uptake in NTHi which suggested that NTHI couples a mechanism of AP resistance with rapid potassium uptake to counter potassium loss associated with AP lethality. Further, SBP bound the iron-containing compound heme and was required for iron homeostasis in NTHi. Here, we extended our functional studies to analyze the Sap transporter permease components (SapB and SapC) and determine their role in acquisition of potassium and iron as well as AP resistance. We determined that the SapC-deficient strain required a forty-fold increase in potassium concentration to rescue growth, compared to that of the parental isolate. SapC complementation fully restored NTHi growth and its ability to transport potassium. A SapB-deficient strain was only partially impaired in potassium transport. In contrast, we determined that SapB, but not SapC, was required for growth in heme-starved conditions suggesting a role for this permease component in heme transport. Complementation of the SapB mutation fully restored NTHi growth in a heme-deplete environment. We also determined that SapC, but not SapB, was required for resistance to APs. Collectively, our data support a coupling of potassium uptake with AP resistance, both requiring the SapC permease component, yet independent from SapB-dependent heme transport mechanisms. These data support our hypothesis that Sap proteins differentially associate to support the critical, yet multifunctional, roles of the Sap transporter and thereby link metabolic needs to innate immunity resistance mechanisms. This work was supported by R01 DC03915 from NIDCD/NIH and R21 A1070825 from NIAID/NIH.