Keratin 8-deletion induced colitis predisposes to murine colorectal cancer enforced by the inflammasome and IL-22 pathway.

Keratins (K) are intermediate filament proteins important in protection from cellular stress. K8, K18 and K19 are the main components of keratin filaments in colonic epithelia but their role in intestinal diseases remains ambiguous. A function for keratins in intestinal health is supported by the K8-knock-out (K8(-/-)) mouse which manifests an early chronic ulcerative colitis-like inflammatory bowel disease and epithelial hyperproliferation. We tested whether K8(-/-) mice are more susceptible to colorectal cancer (CRC) compared to K8 wild type (K8(+/+)), and K8 heterozygote (K8(+/-)) mice showing increased proliferation but no inflammation. K8(-/-) mice did not develop CRC spontaneously, but had dramatically increased numbers of tumors in the distal colon in the azoxymethane (AOM) and Apc(Min/+) CRC models while neither K8(+/+) nor K8(+/-) mice were susceptible. Upregulation of IL-22 in combination with a complete loss of its negative regulator IL-22BP, and increased downstream STAT3-signaling in K8(-/-) and K8(-/-)Apc(Min/+) colonic epithelia confirmed that the IL-22 pathway, important in inflammation, proliferation and tissue regeneration, was activated. The nearly total loss of IL-22BP correlated with an activated inflammasome leading to increased cleaved caspase-1, and the putative IL-22BP inhibitor, IL-18, as well as a decrease in ALDH1/2. Ablation of K8 in a colorectal cancer cell line similarly resulted in increased IL-18 and decreased ALDH1/2. K8/K18 co-immunoprecipitated with pro-caspase-1, a component of the inflammasome in the colon, which suggests that keratins modulate inflammasome activity and protect the colon from inflammation and tumorigenesis. The K8-null mouse models also provide novel epithelial-derived robust colon-specific CRC models.

[1]  M. Omary,et al.  Keratin 20 helps maintain intermediate filament organization in intestinal epithelia. , 2003, Molecular biology of the cell.

[2]  H. Thompson,et al.  A Method for Serial Tissue Processing and Parallel Analysis of Aberrant Crypt Morphology, Mucin Depletion, and Beta-Catenin Staining in an Experimental Model of Colon Carcinogenesis , 2010, Biological Procedures Online.

[3]  K. Hung,et al.  Molecular Mechanisms of Colorectal Carcinogenesis , 2013 .

[4]  V. Karantza,et al.  Keratins in health and cancer: more than mere epithelial cell markers , 2011, Oncogene.

[5]  Francis J. Huber,et al.  IL-22BP is regulated by the inflammasome and modulates tumorigenesis in the intestine , 2012, Nature.

[6]  D. Kamp,et al.  Vimentin regulates activation of the NLRP3 inflammasome , 2015, Nature Communications.

[7]  Stefan Wirtz,et al.  Chemically induced mouse models of intestinal inflammation , 2007, Nature Protocols.

[8]  Bert Vogelstein,et al.  APC mutations occur early during colorectal tumorigenesis , 1992, Nature.

[9]  T. Magin,et al.  Regulation of keratin network organization. , 2015, Current opinion in cell biology.

[10]  IL-22 is related to development of human colon cancer by activation of STAT3 , 2013, BMC Cancer.

[11]  P. Rutgeerts,et al.  Unique Gene Expression and MR T2 Relaxometry Patterns Define Chronic Murine Dextran Sodium Sulphate Colitis as a Model for Connective Tissue Changes in Human Crohn’s Disease , 2013, PloS one.

[12]  E. Lane,et al.  Keratin mutations and intestinal pathology , 2004, The Journal of pathology.

[13]  M. Parkes,et al.  Human keratin 8 mutations that disturb filament assembly observed in inflammatory bowel disease patients , 2004, Journal of Cell Science.

[14]  Judy H. Cho,et al.  Analysis of keratin polypeptides 8 and 19 variants in inflammatory bowel disease. , 2007, Clinical gastroenterology and hepatology : the official clinical practice journal of the American Gastroenterological Association.

[15]  P. Boor,et al.  Keratins in health and disease. , 2015, Current opinion in cell biology.

[16]  A. Takayanagi,et al.  Interleukin-22, a member of the IL-10 subfamily, induces inflammatory responses in colonic subepithelial myofibroblasts. , 2005, Gastroenterology.

[17]  T. P. Pretlow,et al.  Aberrant crypt foci: what we know and what we need to know. , 2007, Clinical gastroenterology and hepatology : the official clinical practice journal of the American Gastroenterological Association.

[18]  M. Omary,et al.  Keratin 8 absence down-regulates colonocyte HMGCS2 and modulates colonic ketogenesis and energy metabolism , 2015, Molecular biology of the cell.

[19]  M. Merad,et al.  Interleukin-22 binding protein (IL-22BP) is constitutively expressed by a subset of conventional dendritic cells and is strongly induced by retinoic acid , 2013, Mucosal Immunology.

[20]  I. Petersen,et al.  Cytokeratin Profiles Identify Diagnostic Signatures in Colorectal Cancer Using Multiplex Analysis of Tissue Microarrays , 2006, Cellular oncology : the official journal of the International Society for Cellular Oncology.

[21]  C. Alam,et al.  The Amount of Keratins Matters for Stress Protection of the Colonic Epithelium , 2015, PloS one.

[22]  T. Takayama,et al.  Aberrant crypt foci: detection, gene abnormalities, and clinical usefulness. , 2005, Clinical gastroenterology and hepatology : the official clinical practice journal of the American Gastroenterological Association.

[23]  P. Coulombe,et al.  The expanding significance of keratin intermediate filaments in normal and diseased epithelia. , 2013, Current opinion in cell biology.

[24]  Joachim L. Schultze,et al.  Keratin 1 maintains skin integrity and participates in an inflammatory network in skin through interleukin-18 , 2012, Journal of Cell Science.

[25]  M. Omary,et al.  Keratin-8-deficient mice develop chronic spontaneous Th2 colitis amenable to antibiotic treatment , 2005, Journal of Cell Science.

[26]  M. Silverberg,et al.  Higher Activity of the Inducible Nitric Oxide Synthase Contributes to Very Early Onset Inflammatory Bowel Disease , 2014, Clinical and Translational Gastroenterology.

[27]  Takuji Tanaka,et al.  Dextran sodium sulfate strongly promotes colorectal carcinogenesis in ApcMin/+ mice: Inflammatory stimuli by dextran sodium sulfate results in development of multiple colonic neoplasms , 2006, International journal of cancer.

[28]  S. Rutz,et al.  Interleukin-22 induces interleukin-18 expression from epithelial cells during intestinal infection. , 2015, Immunity.

[29]  H. Baribault,et al.  Mid-gestational lethality in mice lacking keratin 8. , 1993, Genes & development.

[30]  G. Barber,et al.  Diverse roles of STING-dependent signaling on the development of cancer , 2015, Oncogene.

[31]  Shengdi Hu,et al.  An Efficient Genotyping Method for Genome-modified Animals and Human Cells Generated with CRISPR/Cas9 System , 2014, Scientific Reports.

[32]  M. Omary,et al.  Intermediate filaments take the heat as stress proteins. , 2010, Trends in cell biology.

[33]  R. Iozzo,et al.  Colorectal hyperplasia and inflammation in keratin 8-deficient FVB/N mice. , 1994, Genes & development.

[34]  S. Cross,et al.  Inflammation decreases keratin level in ulcerative colitis; inadequate restoration associates with increased risk of colitis-associated cancer , 2015, BMJ open gastroenterology.

[35]  T. Magin,et al.  Cytokines as genetic modifiers in K5–/– mice and in human epidermolysis bullosa simplex , 2009, Human mutation.

[36]  R. Sabat,et al.  Interleukin-22: a cytokine produced by T, NK and NKT cell subsets, with importance in the innate immune defense and tissue protection. , 2010, Cytokine & growth factor reviews.

[37]  R Fodde,et al.  A targeted chain-termination mutation in the mouse Apc gene results in multiple intestinal tumors. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[38]  M. Lukáš.  Inflammatory Bowel Disease as a Risk Factor for Colorectal Cancer , 2010, Digestive Diseases.

[39]  H. Knutsen,et al.  The min mouse on FVB background: susceptibility to spontaneous and carcinogen-induced intestinal tumourigenesis. , 2011, Anticancer research.

[40]  Y. Dou,et al.  IL-22(+)CD4(+) T cells promote colorectal cancer stemness via STAT3 transcription factor activation and induction of the methyltransferase DOT1L. , 2014, Immunity.

[41]  M. Omary,et al.  Toward unraveling the complexity of simple epithelial keratins in human disease. , 2009, The Journal of clinical investigation.

[42]  W. Ouyang,et al.  Deciphering the crosstalk among IL-1 and IL-10 family cytokines in intestinal immunity. , 2015, Trends in immunology.

[43]  M. Neurath,et al.  STAT3 links IL-22 signaling in intestinal epithelial cells to mucosal wound healing , 2009, The Journal of experimental medicine.

[44]  Satish K. Singh,et al.  Keratins modulate colonocyte electrolyte transport via protein mistargeting , 2004, The Journal of cell biology.

[45]  G. Duester.  Families of retinoid dehydrogenases regulating vitamin A function: production of visual pigment and retinoic acid. , 2000, European journal of biochemistry.

[46]  F. Powrie,et al.  Innate lymphoid cells sustain colon cancer through production of interleukin-22 in a mouse model , 2013, The Journal of experimental medicine.

[47]  R. Moll,et al.  The human keratins: biology and pathology , 2008, Histochemistry and Cell Biology.

[48]  M. Omary,et al.  Absence of keratin 8 confers a paradoxical microflora-dependent resistance to apoptosis in the colon , 2011, Proceedings of the National Academy of Sciences.

[49]  D. Hanahan,et al.  Hallmarks of Cancer: The Next Generation , 2011, Cell.

[50]  D. Haro,et al.  Ketogenic HMGCS2 Is a c-Myc Target Gene Expressed in Differentiated Cells of Human Colonic Epithelium and Down-Regulated in Colon Cancer , 2006, Molecular Cancer Research.

[51]  David A. Scott,et al.  Genome engineering using the CRISPR-Cas9 system , 2013, Nature Protocols.

[52]  D. Artis,et al.  Border patrol: regulation of immunity, inflammation and tissue homeostasis at barrier surfaces by IL-22 , 2011, Nature Immunology.

[53]  C. Rao,et al.  Adoptive transfer of regulatory T cells promotes intestinal tumorigenesis and is associated with decreased NK cells and IL‐22 binding protein , 2015, Molecular carcinogenesis.

[54]  E. Mizoguchi,et al.  Animal models of IBD: linkage to human disease. , 2010, Current opinion in pharmacology.

[55]  Markus F Neurath,et al.  An inducible mouse model of colon carcinogenesis for the analysis of sporadic and inflammation-driven tumor progression , 2007, Nature Protocols.

[56]  I. Singer,et al.  Cyclooxygenase 2 is induced in colonic epithelial cells in inflammatory bowel disease. , 1998, Gastroenterology.