Mycobacterium tuberculosis(Mtb) infection of macrophages reprograms cellular metabolism to promote lipid retention. While it is clearly known that intracellularMtbutilize host derived fatty acids and cholesterol to fuel the majority of its metabolic demands, the role of macrophage lipid catabolism on the bacteria’s ability to access the intracellular lipid pool remains undefined. We utilized a CRISPR genetic knockdown approach to assess the impact of sequential steps in fatty acid metabolism on the growth of intracellularMtb. Our analyzes demonstrate that knockdown of lipid import, sequestration and metabolism genes collectively impair the intracellular growth ofMtbin macrophages. We further demonstrate that modulating fatty acid homeostasis in macrophages impairsMtbreplication through diverse pathways like enhancing production of pro-inflammatory cytokines, autophagy, restricting the bacteria access to nutrients and increasing oxidative stress. We also show that impaired macrophage lipid droplet biogenesis is restrictive to intracellularMtbreplication, but increased induction of the same by blockade of downstream fatty acid oxidation fails to rescueMtbgrowth. Our work expands our understanding of how host fatty acid homeostasis impactsMtbgrowth in the macrophage.SignificanceMycobacterium tuberculosis(Mtb) primarily infects macrophages in the lungs. In infected macrophages,Mtbuses host lipids as key carbon sources to maintain infection and survive. In this work, we used a CRISPR-Cas9 gene knockout system in murine macrophages to examine the role of host fatty acid metabolism on the intracellular growth ofMtb. Our work shows that macrophages which cannot either import, store or catabolize fatty acids restrictMtbgrowth by both common and divergent anti-microbial mechanisms, including increased glycolysis, increased production of reactive oxygen species, production of pro-inflammatory cytokines, enhanced autophagy and nutrient limitation. Our findings demonstrate that manipulating lipid metabolism in macrophages controlsMtbthrough multiple other mechanisms, beyond limiting the bacteria’s access to nutrients.
