The fat is in the lysosome: how Mycobacterium tuberculosis tricks macrophages into storing lipids

In this issue of the JCI, Bedard et al. (16) shine a light on these processes. By using correlative light and electron microscopy (CLEM), Bedard et al. demonstrate that the particular inclusions within swollen lysosomes of human macrophages were of a lipid nature, suggesting that 1-TbAd induced lipid storage in these cells (Figure 1). This finding was confirmed by immunofluorescence microscopy using BODIPY staining. A two-hour pulse of 1-TbAd was sufficient to trigger durable lipid accumulation in macrophages over several days. 1-TbAd–induced lipid accumulation in human macrophages was clearly related to lysosome remodeling, since this phenotype could be recapitulated with chloroquine, a classical lysosomotropic drug, but not with N6-tubercolosinyladenosine (N6-TbAd), a natural isomer of 1-TbAd with no lysosomotropism or antiacid properties (12, 13). To obtain a comprehensive picture of the lipids that accumulated in macrophages treated with 1-TbAd, Bedard et al. (16) performed lipidomic analysis and found that 1-TbAd increased the pools of cholesteryl esters and triacylglycerol, which are the main lipids that accumulate in M. tuberculosis–induced foamy macrophages (24). 1-TbAd also enhanced the amounts of monoalkyl-diacylglycerol, β-glucosylceramide, and lactosylceramide, which are lysosomal hydrolase substrates known to be stored in specific lysosomal storage disorders, including Wolman and Gaucher diseases.

1-TbAd produced by M. tuberculosis induces lysosomal lipid storage.Figure 1

1-TbAd produced by M. tuberculosis induces lysosomal lipid storage. (i) M. tuberculosis (Mtb) produces 1-TbAd, which raises the vacuolar pH and inhibits acidic hydrolases, including lipases, inside the phagolysosome. (ii) Consequently, lipids, including cholesteryl esters and triglycerides, accumulate in the vacuole, mimicking lysosomal storage diseases. (iii) Under conditions of restricted lipid access, M. tuberculosis can use these lipids as a carbon source to promote its intracellular growth.

On the basis of these results, Bedard et al. (16) hypothesized that by raising lysosomal pH, 1-TbAd inhibits the activity of intralysosomal acid hydrolases, resulting in the accumulation of lipids in this compartment. To address this hypothesis, the authors used enzyme-activated fluorogenic probes and showed that 1-TbAd strongly inhibited the acid-dependent glycosidase and protease activity within intact macrophages. Lysosomes play a fundamental role in the autophagic process by fusing with autophagosomes and degrading their contents. Consequently, the autophagic flux can be strongly disrupted as a consequence of any lysosomal dysfunction, as observed in lysosomal storage diseases or when using lysosomotropic drugs such as chloroquine. In agreement, Bedard et al. found that 1-TbAd, but not N6-TbAd, induced the accumulation of autophagosomes in macrophages, most likely by impairing autophagosome-lysosome fusion. Next, Bedard et al. asked whether direct effects of 1-TbAd on lysosomal lipid storage could also be observed in macrophages infected with 1-TbAd–producing M. tuberculosis strains. Electron microscopy (EM) analyses combined with immunogold staining for CD63, a lysosomal marker, confirmed, that infection with wild-type M. tuberculosis, but not with the Rv3378c-KO strain, which does not produce 1-TbAd, induced swelling of macrophage phagolysosomes. In addition, immunofluorescence microscopy revealed a substantial induction of lipid inclusions (indicated by Nile red staining) at day four after infection with wild-type and complemented bacteria, compared with the 1-TbAd–deficient mutant. Lipid inclusions in M. tuberculosis–infected macrophages were clearly localized to phagolysosomal (i.e., LAMP1+) compartments, as revealed by CLEM (16).

Since some lipid inclusions were observed in the phagolysosomes that contained bacteria, Bedard et al. then wondered whether M. tuberculosis could use these lipids, in particular cholesterol, as a carbon source to grow inside these cells. To answer this question, they measured intracellular M. tuberculosis growth in the presence of all-trans-retinoic acid (ATRA), which limits M. tuberculosis growth by restricting access to cholesterol (8), alone or in combination with 1-TbAd. Remarkably, treatment with 1-TbAd, but not N6-TbAd, abrogated ATRA-mediated restriction of M. tuberculosis growth in macrophages in a dose-dependent manner, suggesting that 1-TbAd–induced lipid storage in phagolysosomes provides nutrients to M. tuberculosis to support bacterial growth. Finally, Bedard et al. sought to assess whether the lysosomal failure of macrophages triggered by 1-TbAd could be reversed using pharmacological regulators of lysosomal activity. In particular, they focused on an agonist (C8) of transient receptor potential mucolipin channel 1 (TRPML1), a lysosomal Ca2+ release channel that prevents lysosomal dyshomeostasis (1620). As hypothesized, treatment of 1-TbAd–stimulated macrophages with C8 decreased the size of swollen lysosomes by more than two-fold, dramatically reduced the number of lipid inclusions, and restored glycolipid catabolism (16).

留言 (0)

沒有登入
gif