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Mycobacterium tuberculosis, the causative agent of tuberculosis disease (TB), is one of the top infectious killers in the world. The sole licensed vaccine, M. bovis BCG, does not reliably confer protective immunity in adults. Although anti-mycobacterial antibiotics are available and routinely prescribed, the rise of antibiotic resistance and the shortcomings of the BCG vaccine highlight the importance of new host-directed therapies and vaccine strategies against TB. A more complete understanding of the immune response to M. tuberculosis is necessary to combat and prevent TB. Two components of innate immunity have been shown to both promote and hinder host control of M. tuberculosis: neutrophils and STING-dependent immune activation.Neutrophil influx and infection occurs during the innate phase of infection. A lack of relevant in vitro models has historically hampered neutrophil studies, given the short half-life of ex vivo neutrophils. We overcame this problem by utilizing a model for neutrophil differentiation and responses to M. tuberculosis infection based on ER-Hoxb8 conditional immortalization of Cas9-expressing neutrophil progenitors. CRISPR/Cas9- mediated gene editing facilitated unbiased genome-wide pooled screens for genes that mediate survival during differentiation and infection. Innate immune activation is also necessary for improved TB protein subunit vaccine design. We aimed to better characterize STING activation in the context of a mucosally administered cyclic-di- nucleotide adjuvanted protein subunit TB vaccine. A previous study had correlated IL-17 producing CD4 T cells (Th17s) with mucosal vaccine protection, instead of the IFN-γ- producing CD4 T cell (Th1) response seen with subcutaneous BCG or protein subunit vaccination. We discovered that both IL-17 and IFN-γ contributed to immunity. The transcription factors STAT6 and IRF3 were not essential for mucosal vaccine-induced immunity downstream of STING, although type I IFN signaling contributed to vaccine efficacy. Dependent on vaccine antigen, our mucosal vaccine could promote durable Th17 responses and antibacterial resistance after the efficacy of BCG had waned. These results provide a clearer picture of several aspects of innate and adaptive immunity.

The p53-regulated long noncoding RNA lincRNA-p21 has been proposed to act in trans via several mechanisms ranging from repressing genes in the p53 transcriptional network to regulating mRNA translation and protein stability. To further examine lincRNA-p21 function, we generated a conditional knockout mouse model. We find that lincRNA-p21 predominantly functions in cis to activate expression of its neighboring gene, p21. Mechanistically, we show that lincRNA-p21 acts in concert with hnRNP-K as a coactivator for p53-dependent p21 transcription. Additional phenotypes of lincRNA-p21 deficiency could be attributed to diminished p21 levels, including deregulated expression and altered chromatin state of some Polycomb target genes, a defective G1/S checkpoint, increased proliferation rates, and enhanced reprogramming efficiency. These findings indicate that lincRNA-p21 affects global gene expression and influences the p53 tumor suppressor pathway by acting in cis as a locus-restricted coactivator for p53-mediated p21 expression., National Institutes of Health (U.S.), Howard Hughes Medical Institute, Ludwig Center for Molecular Oncology, Damon Runyon Cancer Research Foundation

The two unrelated miRNAs miR-143 and miR-145, coexpressed from the miR-143/145 cluster, have been proposed to act as tumor suppressors in human cancer, and therapeutic benefits of delivering miR-143 and miR-145 to tumors have been reported. In contrast, we found that tumor-specific deletion of miR-143/145 in an autochthonous mouse model of lung adenocarcinoma did not affect tumor development. This was consistent with the lack of endogenous miR-143/145 expression in normal and transformed lung epithelium. Surprisingly, miR-143/145 in the tumor microenvironment dramatically promoted tumor growth by stimulating the proliferation of endothelial cells. Loss of miR-143/145 in vivo led to derepression of the miR-145 target CAMK1D, an inhibitory kinase, which when overexpressed prevents mitotic entry of endothelial cells. As a consequence, tumors in miR-143/145-deficient animals exhibited diminished neoangiogenesis, increased apoptosis, and their expansion was limited by the tumor’s ability to co-opt the alveolar vasculature. These findings demonstrate that stromal miR-143/145 promotes tumorigenesis and caution against the use of these miRNAs as agents in cancer therapeutics.SIGNIFICANCE: This study shows that miR-143/145 expressed from the tumor microenvironment stimulates neoangiogenesis and supports tumor expansion in the lung, demonstrating a surprising role for the putative tumor suppressor miRNA cluster in promoting tumorigenesis. We propose inhibition of miR-143/145 as a therapeutic avenue to modulate tumor neoangiogenesis., National Institutes of Health (U.S.) (Grant P01-CA42063-26), National Cancer Institute (U.S.) (Grant P30-CA14051)