Tuberculosis (TB) is a major public health concern, affecting millions of people worldwide. Current Mycobacterium tuberculosis (Mtb) treatment strategies have many limitations including long treatment duration, drug toxicity, emergence of drug-resistant strains, and inadequate efficacy. One new strategy to eradicate Mtb is the use of host directed therapy; however, we must first gain a better understanding of how the host responds to Mtb infection. Understanding that IFNg is critical for Mtb control, we used IFNg to dissect macrophage responses. Here, we used a CRISPR Cas9 screen to broadly understand genes necessary for IFNg-dependent MHCII expression. MHCII drives T-cell activation needed for pathogen clearance. Additionally, we took advantage of a new alveolar macrophage model, known as FLAMs, that was optimized by our lab, to better understand AM IFNg-responses. Our findings reveal that IFNg robustly activates both macrophage types; however, the profile of activated IFNg-stimulated genes varies significantly. Notably, FLAMs show limited activation of costimulatory markers essential for T cell activation upon IFNg stimulation alone. However, with the inhibition of GSK3a/b, a well-conserved multifunctional kinase, FLAMs express a high amount of co-stimulatory molecules, particularly CD40. We also discovered that TNF and IFNg contribute to the increase in costimulatory molecules during GSK3a/b inhibition and IFNg stimulation. Together, these data suggest that AMs' capacity to respond to IFNg is restricted in a GSK3a/b dependent manner and that IFNg responses differ across distinct macrophage populations.
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