Regulation of the LATS2/YAP/TAZ pathway during Helicobacter pylori induced gastric carcinogenesis
Molina Silvia 1,2, Staedel Cathy 3, Giraud Julie 1,2, Tiffon Camille 1,2, Fernandez Solène 1,2, Sifré Elodie 1,2, Bœuf Hélène4, Lehours Philippe 1,2, Dubus Pierre 1,2,
Mégraud Francis 1,2, Varon Christine 1,2
1
University of Bordeaux, Bordeaux, France
2
INSERM U1053 BaRITOn, Bordeaux, France
3
INSERM U1212 ARNA, Bordeaux, France
4
INSERM U1026 BIOTIS, Bordeaux, France
Introduction: Cancer stem cells (CSC) have been identified in gastric carcinoma, in which they
control tumor initiation, growth and dissemination. We reported that Helicobacter pylori infection leads through an epithelial-mesenchymal transition to the emergence of CD44+ cells with CSC properties. The YAP/TAZ co-transcription factors of the Hippo pathway control cancer initiation and progression in many cancers, but their regulation in the context of H. pylori mediated gastric carcinogenesis has not been described.
Materials and Methods: This work aimed to study the role of the Hippo/YAP/TAZ pathway during H. pylori mediated EMT and emergence of CSC-like cells. Coculture experiments of MKN74,
MKN45 and AGS gastric cell lines with the 7.13 cagA+ and cagA mutant H. pylori strains were performed. We evaluated YAP/TEAD transcriptional activity and YAP/TAZ/TEAD target genes expression (RT-qPCR). The consequences of LATS2 and YAP/TAZ inhibition by siRNA and by a YAP/TEAD inhibitor were evaluated on H. pylori-induced EMT and CSC properties (tumorspheres). The expression of Hippo/YAP/TAZ components we evaluated in gastric tissues (immunohistochemistry) of patients infected or not with H. pylori, and with gastric carcinoma.
Results: H. pylori via CagA induced a rapid activation of YAP/TAZ and upregulation of their target
genes, which remained under the tight control of LATS2. YAP/LATS2 overexpression was also detected in H. pylori-infected gastric mucosa. In vitro, YAP/TAZ inhibition reduced EMT/CSC markers expression, and efficiently reduced the pool of CD44+ gastric CSCs forming tumorspheres. In gastric carcinoma, YAP and LATS2 were co-overexpressed in 5-30% of cells, which may correspond to gastric CSCs.
Conclusion: YAP/TAZ and LATS2 are activated in H. pylori-infected gastric epithelial cells and their
sustained activity controls EMT and CSC properties, constituting a promising target to inhibit gastric carcinogenesis.
Interleukin-18 mediates immune responses in Campylobacter jejuni infected secondary abiotic mice Heimesaat Markus M., Alutis Marie E., Grundmann Ursula,
Fischer André, Bereswill Stefan
Charité, University Medicine Berlin, Institute of Microbiology and Hygiene, Gastrointestinal Microbiology Research Group,
Berlin, Germany
Background: Human Campylobacter jejuni infections are progressively rising worldwide.
Informations about the molecular mechanisms underlying campylobacteriosis, however, are limited. In the present study we investigated whether cytokines such as IL-23, IL-22 and IL-18 sharing pivotal functions in host immunity were involved in mediating intestinal and systemic immunopathological responses upon C. jejuni infection.
Methodology/Results: To assure stable infection, secondary abiotic IL-23p19-/-, IL-22-/- and IL-18-
/- mice were generated by broad-spectrum antibiotic treatment. Following peroral C. jejuni strain 81-176 infection, mice of either genotype harbored comparably high pathogenic loads in their intestines. As compared to wildtype controls, however, IL-18-/- mice displayed less distinct C.
jejuni induced sequelae as indicated by less pronounced large intestinal shrinkage and lower
numbers of apoptotic cells in the colonic epithelial layer at day 8 postinfection (p.i.), that were accompanied by lower colonic numbers of adaptive immune cells including regulatory T cells and B lymphocytes and less distinct secretion of pro-inflammatory cytokines such as TNF, IFN-gamma, and IL-17A in colonic ex vivo biopsies at day 8 p.i. Upon C. jejuni infection, colonic IL-23p19 expression was up-regulated in IL-18-/- mice only, whereas IL-22 mRNA levels were lower in naive and infected IL-23p19-/- as well as infected IL-18-/- as compared to respective wildtype control mice. Remarkably, not only intestinal, but also systemic infection-induced immune responses were less pronounced in IL-18-/- mice as indicated by lower TNF, IFN-gamma, and IL-6 serum levels as compared to wildtype mice.
Conclusion: We here show for the first time that IL-18 is essentially involved in mediating C. jejuni
infection in the secondary abiotic mouse model. Future studies need to further unravel the underlying regulatory mechanisms orchestrating pathogenic-host interaction.
Campylobacter jejuni induces autoimmune peripheral neuropathy via Siglec-1 and IL-4 axes
in a mouse model of Guillain Barré Syndrome Ankit Malik, Jean M. Brudvig, Barbie J. Gadsden,
Alexander Ethridge, Linda S. Mansfield
Comparative Enteric Diseases Laboratory, College of Veterinary Medicine, Michigan State University, East Lansing, MI, USA
Campylobacter jejuni is a Gram-negative bacterium that is the most common bacterial cause of
gastroenteritis worldwide. C. jejuni infection has also been causally linked with development of the peripheral neuropathy called Guillain Barré Syndrome (GBS). We have previously shown that
C. jejuni isolates from human enteritis patients induce a Type1/17 cytokine dependent colitis
response in IL-10-/- mice. In contrast, isolates from human GBS patients colonize the IL-10-/- mice without inducing colitis but instead induce autoantibodies targeted against peripheral nerve gangliosides. We hypothesized that C. jejuni from GBS patients induce autoimmune responses and nerve lesions dependent upon IL-4 and Siglec-1 axes. C57BL/6 IL-10-/- mice were gavaged orally with C. jejuni strain HB93-13 or 260.94 from GBS patients, half given blocking antibodies for IL-4 or Siglec-1 and all assessed for clinical neurological signs/phenotypes, anti-ganglioside antibodies, cellular immune responses and lesions in gut and peripheral nerve tissues. Vehicle inoculated mice served as infection controls, while mice given isotype control antibody served as controls for IL-4 and Siglec-1 blocking antibody treatments. C. jejuni HB93-13 and 260.94 infection elicited mild GBS in mice. Antiganglioside antibody responses were dependent upon blunted Type1/17 but enhanced Type2 cytokine production by T helper cells. Autoantibody production correlated with enhanced macrophage infiltration in sciatic nerves and their dorsal root ganglia. Autoantibodies and histological lesions were significantly decreased in mice depleted of IL-4, without leading to induction of colitis. Peripheral nerve lesions were mild in infected mice, but were associated with abnormal gait and hind limb movements consistent with this syndrome’s manifestation in humans. Moreover, Siglec-1 served as a central antigen presenting cell receptor mediating GBS but not colitogenic isolate uptake, T cell differentiation and autoantibody elicitation. Thus, this is the first mouse model of an autoimmune disease induced directly by a bacterium that is dependent upon Siglec-1 and IL-4 axes.
Funded by NIAID, NIH, Department of Health and Human Services under ERIN CRC grant U19AI090872
Downregulation of USF1 transcription factors impacts p53 during Helicobacter pylori infection
and exacerbates gastric carcinogenesis Costa Lionel1, Corre Sébastien2, Michel Valérie1, JouvionGrégory3, Mouchet Nicolas2, De Reuse Hilde1,
Galibert Marie-Dominique2, Touati Eliette1
1
Institut Pasteur, Dpt of Microbiology, Unit of Helicobacter Pathogenesis, Paris, France
2
CNRS UMR6290, University of Rennes 1, Rennes, France
3
Institut Pasteur, Dpt of Infection and Epidemiology, Unit of Human Histopathology and Animal Model,
Paris, France
Introduction: Helicobacter pylori is the major risk factor for gastric cancer. It induces genetic
instabilities in gastric epithelial cells associated to chronic inflammation and impairs host DNA repair systems. Its oncoprotein CagA promotes the proteasomal degradation of p53, a central regulator of the DNA damage response. We previously reported that H. pylori inhibits the expression of the genes coding for the upstream stimulating factors USF1 and USF2 by DNA hypermethylation. USF belong to the basic helix-loop-helix-leucine zipper (bHLH-zip) class of transcription factors. USF1 regulates DNA damage response and the expression of Tp53. In addition, in response to a genotoxic stress USF1 binds to p53 and prevents its proteasomal degradation. In the present study, we investigate the consequences of the deregulation of USF1 on p53 level and DNA damage and repair response during H. pylori infection. We also address the impact of loss of USF1 on the severity of gastric cancer lesions induced by H. pylori.
Methods: Gastric epithelial cells (MKN45/AGS) were infected with H. pylori (7.13/26695) for
2h/24h and/or treated with camptothecin (CPT) a genotoxic compound. USF1 and p53 levels, their cellular localization and USF1-p53 interaction were analysed by immunofluorescence and Duolink proximity ligation assay. The consequences of USF1-deficiency were investigated on gastric lesions-induced by H. pylori in Usf1-/- and Usf1+/+ mice after 9/12 months.
Results: H. pylori inhibits USF1 nuclear level and translocates USF1 in the cytoplasm.
Concomitantly, p53 is depleted in the nuclei. H. pylori inhibits USF1-p53 interaction induced by CPT, and consequently favors p53 degradation. In addition, the severity of H. pylori–induced gastric lesions is exacerbated in Usf1-/- mice compared to Usf1+/+.
Conclusions: USF1 is a central regulator of the DNA damage response to H. pylori infection. Its
deficiency is directly associated with p53 degradation and the promotion of the gastric carcinogenesis process.