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Gastric cancer is the third most lethal cancer worldwide, and evaluation of the genomic status of gastric cancer cells has not translated into effective prognostic or therapeutic strategies.We therefore hypothesize that outcomes may depend on the tumor microenvironment (TME), in particular, cancerassociated fibroblasts (CAF). However, very little is known about the role of CAFs in gastric cancer. To address this, we mapped the transcriptional landscape of human gastric cancer stroma by microdissection and RNA sequencing of CAFs from patients with gastric cancer. A stromal gene signature was associated with poor disease outcome, and the transcription factor heat shock factor 1 (HSF1) regulated the signature. HSF1 upregulated inhibin subunit beta A and thrombospondin 2, which were secreted in CAF-derived extracellular vesicles to the TME to promote cancer. Together, our work provides the first transcriptional map of human gastric cancer stroma and highlights HSF1 and its transcriptional targets as potential diagnostic and therapeutic targets in the genomically stable tumor microenvironment. Significance: This study shows how HSF1 regulates a stromal transcriptional program associated with aggressive gastric cancer and identifies multiple proteins within this program as candidates for therapeutic intervention. better understand the molecular basis of this disease, and to identify 48 biomarkers that may predict outcome and guide therapy.49 Gastric cancer is a heterogeneous disease. Traditionally, anatomical 50 location (true gastric vs. gastro-esophageal) and histologic character-51 istics (diffuse vs. intestinal; tubular vs. papillary) have been used to 52 classify gastric cancer subtypes (2). Recent advances in molecular 53 understanding have enabled classification of gastric cancer into dif-54 ferent subtypes based on chromosomal instability, microsatellite 55 instability, genomic stability, presence of Epstein-Barr virus, and 56 epithelial-mesenchymal transition (EMT), which were associated with 57 different survival outcomes (3-6). Mutations in CDH1 and KRAS, and 58 overexpression of HER2, EGFR, FGFR2, VEGF, were shown to 59 contribute to disease progression and correlate with poor out-60 come (7, 8). Despite serving as valuable guides in deciphering the 61 complexity of gastric cancer, there has been little success in applying 62 these molecular classifiers to treatment stratification and development 63 of targeted therapies (3). Prognosis in the clinic is still mostly evaluated 64 on the basis of TNM staging (tumor size, lymph node involvement, and 65 metastasis), and the standard of care for localized gastric cancer is 66 surgical intervention combined with chemotherapy (7). 67 Increasing evidence over the past decade highlighted the indispens-68 able contribution of the tumor microenvironment (TME) to disease 69 progression and treatment resistance (9). The TME is comprised of 70 various cell types, including endothelial cells, fibroblasts, macro-71 phages, and lymphocytes, as well as extracellular matrix co...

In the colon, long-term exposure to chronic inflammation drives colitis-associated colon cancer (CAC) in patients with inflammatory bowel disease. While the causal and clinical links are well established, molecular understanding of how chronic inflammation leads to the development of colon cancer is lacking. Here we deconstruct the evolving microenvironment of CAC by measuring proteomic changes and extracellular matrix (ECM) organization over time in a mouse model of CAC. We detect early changes in ECM structure and composition, and report a crucial role for the transcriptional regulator heat shock factor 1 (HSF1) in orchestrating these events. Loss of HSF1 abrogates ECM assembly by colon fibroblasts in cell-culture, prevents inflammation-induced ECM remodeling in mice and inhibits progression to CAC. Establishing relevance to human disease, we find high activation of stromal HSF1 in CAC patients, and detect the HSF1-dependent proteomic ECM signature in human colorectal cancer. Thus, HSF1-dependent ECM remodeling plays a crucial role in mediating inflammation-driven colon cancer.

Enteropathogenic E . coli (EPEC) is an extracellular diarrheagenic human pathogen which infects the apical plasma membrane of the small intestinal enterocytes. EPEC utilizes a type III secretion system to translocate bacterial effector proteins into its epithelial hosts. This activity, which subverts numerous signaling and membrane trafficking pathways in the infected cells, is thought to contribute to pathogen virulence. The molecular and cellular mechanisms underlying these events are not well understood. We investigated the mode by which EPEC effectors hijack endosomes to modulate endocytosis, recycling and transcytosis in epithelial host cells. To this end, we developed a flow cytometry-based assay and imaging techniques to track endosomal dynamics and membrane cargo trafficking in the infected cells. We show that type-III secreted components prompt the recruitment of clathrin (clathrin and AP2), early (Rab5a and EEA1) and recycling (Rab4a, Rab11a, Rab11b, FIP2, Myo5b) endocytic machineries to peripheral plasma membrane infection sites. Protein cargoes, e.g. transferrin receptors, β1 integrins and aquaporins, which exploit the endocytic pathways mediated by these machineries, were also found to be recruited to these sites. Moreover, the endosomes and cargo recruitment to infection sites correlated with an increase in cargo endocytic turnover (i.e. endocytosis and recycling) and transcytosis to the infected plasma membrane. The hijacking of endosomes and associated endocytic activities depended on the translocated EspF and Map effectors in non-polarized epithelial cells, and mostly on EspF in polarized epithelial cells. These data suggest a model whereby EPEC effectors hijack endosomal recycling mechanisms to mislocalize and concentrate host plasma membrane proteins in endosomes and in the apically infected plasma membrane. We hypothesize that these activities contribute to bacterial colonization and virulence.

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