Macrophage Second-line Responses to Group A Streptoccocus Infection - PROJECT SUMMARY
The resurgence of severe invasive Group A Streptococcus (GAS) infections in the past decade
is a major public health concern in the United States, and gaps in understanding the resilience
of GAS to the immune response has impeded the development of a much-needed vaccine. The
long-term goal is to understand GAS survival and persistence in macrophages to provide a cell-
targeted basis for improved treatment for infection. Toward the long-term goal, the objective in
this application is to examine macrophage second-line responses to GAS following the failure of
lysosomes to eliminate bacteria. The central hypothesis is that GAS-induced lysosomal
permeabilization enables bacterial and lysosomal proteins to enter the cytosol, which continues
to alter macrophage cellular processes, including limiting the oxidative burst response and
inducing histone release and non-lytic exocytosis of GAS. Guided by preliminary data produced
by the applicant’s laboratory, this hypothesis will be explored in three specific aims. The work in
Aim 1 will determine whether GAS NADase in the cytosol impairs the oxidative burst response.
Measurements of reactive oxygen species, cellular energy pools, and NADPH oxidase complex
assembly will determine whether this bacterial enzyme negatively impacts the macrophage
oxidative burst response. Aim 2 will explore whether GAS-induced damage of the nuclear
envelope causes DNA-free histone release and the extent to which these histones have
antimicrobial activity against GAS in the macrophage cytosol. Preliminary data demonstrate that
macrophages expel GAS in exosomes and the work in Aim 3 will characterize these GAS-
containing exosomes. Time-lapse microscopic observations will be complemented by electron
microscopy and proteomics analyses of isolated exosomes. The outcome of this work will inform
approaches aimed at restoring or enhancing host cell function, which both avoids the problems
of broad-spectrum antibiotic treatment and enhances the natural ability of macrophages to
eradicate GAS. The proposed research is innovative because understanding these second-line
responses, which are intermediate processes between lysosomal failure and disease pathology,
will provide new druggable target proteins and pathways. The contribution will be a significant
first step toward developing pharmacological strategies that prevent, rather than treat severe
invasive disease and refractory infections.
