Sunday, November 24, 2024
11/24/2024
The human-specific bacterial pathogen Neisseria gonorrhoeae (Ngo) causes the sexually-transmitted infection gonorrhea. Ngo expresses outer membrane transporters to acquire iron and other metals in a TonB-dependent manner. In order to prevent microbial growth and infectivity, humans use high affinity metal-binding proteins to sequester essential metals, a phenomenon known as “nutritional immunity.” Ngo subverts nutritional immunity by using dedicated TonB-dependent transporters (TdTs) that bind to and remove the metals from human metal- binding proteins such as transferrin and lactoferrin, which sequester iron (Fe). Our preliminary data indicates that two Ngo outer membrane transporters, TdfH and TdfJ, enable Ngo to internalize zinc (Zn) from Zn- sequestering, human innate immunity proteins. TdfH affords Ngo the ability to overcome Zn sequestration imposed by the innate immunity protein calprotectin (CP), which is produced in abundance by neutrophils and accumulates to high concentrations at sites of inflammation. Furthermore, in the prior funding period, we determined that TdfJ enables the use of S100A7, also known as psoriasin, as the sole Zn source. Two other transporters, TdfF and TdfG, are not as well defined, but their expression is iron-repressed. We hypothesize that these metal transporters enable Ngo to overcome the growth inhibitory effects of the innate immunity proteins produced during inflammation. These studies are significant because knowing how Ngo acquires essential metals like Zn and Fe from the human host opens new opportunities for developing new therapeutics against ‘superbug’ strains, whether they be targets for a protective vaccine or small molecule inhibitors of crucial transporters. The overarching hypothesis to be tested in the proposed study is that Ngo subverts nutritional immunity imposed by host metal-chelating proteins including S100 proteins, which are produced by neutrophils and epithelial cells, by the deployment of outer membrane transporters, which bind to and relieve these proteins of their sequestered transition metal cargo. To test these hypotheses, the following specific aims are proposed: Aim 1 will interrogate the metal environment sensed by Ngo when interacting with human cervical cells and primary human PMNs, cell types important in human infection. We will identify the infection conditions under which Ngo experiences Fe and Zn limitation using fluorescent reporters and RNA-seq approaches. Aim 2 will define the mechanisms for subversion of host nutritional immunity by Ngo TdTs, employing structural biology and protein-protein interaction techniques. Aim 3 will elucidate the consequences of inhibiting the biological functions of the TdTs in vitro and with host cells. These studies will test monoclonal antibodies, TdT mutagenesis and S100 protein inhibitors for impacts on host cell survival and metal uptake. Overall, these innovative studies will build on exciting findings from the prior funding period and will also move into innovative new areas of investigation, deploying state-of-the-art technologies. The proposed studies will shed new light on fundamental pathogenic mechanisms used by Ngo to overcome nutritional immunity imposed by the human host.
HHS’ Tracking Accountability in Government Grants System (TAGGS) website provides access to detailed descriptions of grants, loans, aggregated direct payments and other types of financial assistance awarded by the HHS Operating Divisions (OPDIVs) and the Office of the Secretary Staff Divisions (STAFFDIVs).
