Mycobacterium tuberculosis (Mtb) survives within macrophages by evading delivery to the lysosome and promoting the accumulation of lipid bodies, which serve as a bacterial source of nutrients. Here we show that by inducing miR-33 and its passenger strand miR-33*, Mtb inhibits integrated pathways involved in autophagy, lysosomal function and fatty acid oxidation to support bacterial replication. Silencing of miR-33 and miR-33* by genetic or pharmacological means promotes autophagy flux through derepression of key autophagy effectors such as ATG5, ATG12, LC3B and LAMP1 and AMPK-dependent activation of the transcription factors FOXO3 and TFEB, enhancing lipid catabolism and Mtb xenophagy. These data define a mammalian miRNA circuit utilized by Mtb to coordinately inhibit autophagy and reprogram host lipid metabolism to enable intracellular survival and persistence in the host.

BackgroundA powerful approach to understanding complex processes such as aging is to use model organisms amenable to genetic manipulation, and to seek relevant phenotypes to measure. Caenorhabditis elegans is particularly suited to studies of aging, since numerous single-gene mutations have been identified that affect its lifespan; it possesses an innate immune system employing evolutionarily conserved signaling pathways affecting longevity. As worms age, bacteria accumulate in the intestinal tract. However, quantitative relationships between worm genotype, lifespan, and intestinal lumen bacterial load have not been examined. We hypothesized that gut immunity is less efficient in older animals, leading to enhanced bacterial accumulation, reducing longevity. To address this question, we evaluated the ability of worms to control bacterial accumulation as a functional marker of intestinal immunity.ResultsWe show that as adult worms age, several C. elegans genotypes show diminished capacity to control intestinal bacterial accumulation. We provide evidence that intestinal bacterial load, regulated by gut immunity, is an important causative factor of lifespan determination; the effects are specified by bacterial strain, worm genotype, and biologic age, all acting in concert.ConclusionsIn total, these studies focus attention on the worm intestine as a locus that influences longevity in the presence of an accumulating bacterial population. Further studies defining the interplay between bacterial species and host immunity in C. elegans may provide insights into the general mechanisms of aging and age-related diseases.

Store-operated Ca2+ entry (SOCE) through Ca2+ release-activated Ca2+ (CRAC) channels is essential for immunity to infection. CRAC channels are formed by ORAI1 proteins in the plasma membrane and activated by stromal interaction molecules 1 (STIM1) and STIM2 in the endoplasmic reticulum (ER). Mutations in ORAI1 and STIM1 genes that abolish SOCE cause severe immunodeficiency with recurrent infections due to impaired T cell function. SOCE has also been observed in cells of the innate immune system such as macrophages and dendritic cells (DC) and may provide Ca2+ signals required for their function. The specific role of SOCE in macrophage and DC function, and its contribution to innate immunity, however, is not well defined. We found that non-selective inhibition of Ca2+ signaling strongly impairs many effector functions of bone marrow-derived macrophages (BMDMs) and dendritic cells (BMDCs) including phagocytosis, inflammasome activation, and priming of T cells. Surprisingly however, macrophages and DCs from mice with conditional deletion of Stim1 and Stim2 genes – and therefore complete inhibition of SOCE – showed no major functional defects. Their differentiation, FcR-dependent and independent phagocytosis, phagolysosome fusion, cytokine production, NLRP3 inflammasome activation and their ability to present antigens to activate T cells was preserved. Our findings demonstrate that STIM1, STIM2 and SOCE are dispensable for many critical effector functions of macrophages and DCs, which has important implications for CRAC channel inhibition as a therapeutic strategy to suppress pathogenic T cells while not interfering with myeloid cell functions required for innate immunity.

The microbial communities that reside within the intestinal tract in vertebrates are complex and dynamic. In this report, we establish the utility of Caenorhabditis elegans as a model system for identifying the factors that contribute to bacterial persistence and for host control of gut luminal populations. We found that for N2 worms grown on mixed lawns of bacteria, Salmonella enterica serovar Typhimurium substantially outcompeted Escherichia coli, even when E. coli was initially present at 100-foldhigher concentrations. To address whether innate immunity affects the competition, the daf-2 and daf-16 mutants were studied; their total gut bacterial levels reflect overall capacity for colonization, but Salmonella outcompeted E. coli to an extent similar to wild-type worms. To address the role of virulence properties, Salmonella ⌬spi-1 ⌬spi-2 was used to compete with E. coli. The net differential was significantly less than that for wild-type Salmonella; thus, spi-1 spi-2 encodes C. elegans colonization factors. An E. coli strain with repeated in vivo passage had an enhanced ability to compete against an in vitro-passed E. coli strain and against Salmonella. Our data provide evidence of active competition for colonization niches in the C. elegans gut, as determined by bacterial factors and subject to in vivo selection.