Background: It is unclear how YAP gene expression is controlled in colorectal cancer (CRC) cells. Results: ␤-Catenin/TCF4 complexes directly regulate YAP gene expression through a novel DNA enhancer element. Conclusion: YAP is a direct Wnt/␤-catenin target gene, and its expression is required for CRC cell growth. Significance: Aberrant Wnt/␤-catenin signaling may account for oncogenic YAP expression in intestinal cells.

Reductive genome evolution has purged many metabolic pathways from obligate intracellular Rickettsia (Alphaproteobacteria; Rickettsiaceae). While some aspects of host-dependent rickettsial metabolism have been characterized, the array of host-acquired metabolites and their cognate transporters remains unknown. This dearth of information has thwarted efforts to obtain an axenic Rickettsia culture, a major impediment to conventional genetic approaches. Using phylogenomics and computational pathway analysis, we reconstructed the Rickettsia metabolic and transport network, identifying 51 host-acquired metabolites (only 21 previously characterized) needed to compensate for degraded biosynthesis pathways. In the absence of glycolysis and the pentose phosphate pathway, cell envelope glycoconjugates are synthesized from three imported host sugars, with a range of additional host-acquired metabolites fueling the tricarboxylic acid cycle. Fatty acid and glycerophospholipid pathways also initiate from host precursors, and import of both isoprenes and terpenoids is required for the synthesis of ubiquinone and the lipid carrier of lipid I and O-antigen. Unlike metabolite-provisioning bacterial symbionts of arthropods, rickettsiae cannot synthesize B vitamins or most other cofactors, accentuating their parasitic nature. Six biosynthesis pathways contain holes (missing enzymes); similar patterns in taxonomically diverse bacteria suggest alternative enzymes that await discovery. A paucity of characterized and predicted transporters emphasizes the knowledge gap concerning how rickettsiae import host metabolites, some of which are large and not known to be transported by bacteria. Collectively, our reconstructed metabolic network offers clues to how rickettsiae hijack host metabolic pathways. This blueprint for growth determinants is an important step toward the design of axenic media to rescue rickettsiae from the eukaryotic cell.

To establish a habitable intracellular niche, various pathogenic bacteria secrete effectors that target intracellular trafficking and modulate phosphoinositide (PI) metabolism. Murine typhus, caused by the obligate intracellular bacterium Rickettsia typhi, remains a severe disease in humans. However, the mechanisms by which R. typhi effector molecules contribute to internalization by induced phagocytosis and subsequent phagosomal escape into the cytosol to facilitate the intracellular growth of the bacteria remain ill-defined. Here, we characterize a new molecule, Risk1, as a phosphatidylinositol 3-kinase (PI3K) secreted effector and the first bacterial secretory kinase with both class I and III PI3K activities. Inactivation of Risk1 PI3K activities reduced the phosphorylation of phosphatidylinositol 4,5-bisphosphate to phosphatidylinositol 3,4,5-trisphosphate within the host, which consequently diminished host colonization by R. typhi. During infection, Risk1 targets the Rab5-EEA1-phosphatidylinositol 3-phosphate [PI(3)P] signaling axis to promote bacterial phagosomal escape. Subsequently, R. typhi undergoes ubiquitination and induces host autophagy; however, maturation to autolysosomes is subverted to support intracellular growth. Intriguingly, only enzymatically active Risk1 binds the Beclin-1 core complex and contributes to R. typhi-induced autophagosome formation. In sum, our data suggest that Risk1, with dual class I and class III PI3K activities, alters host PI metabolism and consequently subverts intracellular trafficking to facilitate intracellular growth of R. typhi. IMPORTANCE Rickettsia species are Gram-negative obligate intracellular bacteria that infect a wide range of eukaryotes and vertebrates. In particular, human body louse-borne Rickettsia prowazekii and flea-borne Rickettsia typhi have historically plagued humankind and continue to reemerge globally. The unavailability of vaccines and limited effectiveness of antibiotics late in infection place lethality rates up to 30%, highlighting the need to elucidate the mechanisms of Rickettsia pathogenicity in greater detail. Here, we characterize a new effector, Risk1, as a secreted phosphatidylinositol 3-kinase (PI3K) with unique dual class I and class III activities. Risk1 is required for host colonization, and its vacuolar phosphatidylinositol 3-phosphate generation modulates endosomal trafficking to arrest autophagosomal maturation. Collectively, Risk1 facilitates R. typhi growth by altering phosphoinositide metabolism and subverting intracellular trafficking.

The Wnt/β-catenin signaling pathway controls intestinal homeostasis and mutations in components of this pathway are prevalent in human colorectal cancers (CRCs). These mutations lead to inappropriate expression of genes controlled by Wnt responsive DNA elements (WREs). T-cell factor/Lymphoid enhancer factor transcription factors bind WREs and recruit the β-catenin transcriptional co-activator to activate target gene expression. Deregulated expression of the c-MYC proto-oncogene (MYC) by aberrant Wnt/β-catenin signaling drives colorectal carcinogenesis. In this review, we discuss the current literature pertaining to the identification and characterization of WREs that control oncogenic MYC expression in CRCs. A common theme has emerged whereby these WREs often map distally to the MYC genomic locus and control MYC gene expression through long-range chromatin loops with the MYC proximal promoter. We propose that by determining which of these WREs is critical for CRC pathogenesis, novel strategies can be developed to treat individuals suffering from this disease.