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A Novel PAK1-Notch1 Axis Regulates Crypt Homeostasis in Intestinal Inflammation
Adrian Frick1, Vineeta Khare1, Kristine Jimenez1
1Division of Gastroenterology and Hepatology, Department of Internal Medicine III, Medical University of Vienna, Vienna, Austria.
Background & Aims:
p21-activated kinase-1 (PAK1) belongs to a family of serine-threonine kinases and contributes to cellular pathways such as nuclear factor-κB (NF-κB), mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT), and Wingless-related integration site(Wnt)/β-catenin, all of which are involved in intestinal homeostasis. Overexpression of PAK1 is linked to inflammatory bowel disease as well as colitis-associated cancer (CAC), and similarly was observed in interleukin (IL)10 knockout (KO) mice, a model of colitis and CAC. Here, we tested the effects of PAK1 deletion on intestinal inflammation and carcinogenesis in IL10 KO mice.
Methods:
IL10/PAK1 double-knockout (DKO) mice were generated and development of colitis and CAC was analyzed. Large intestines were measured and prepared for histology or RNA isolation. Swiss rolls were stained with H&E and periodic acid-Schiff. Co-immunoprecipitation and immunofluorescence were performed using intestinal organoids, SW480, and normal human colon epithelial cells 1CT.
Results:
When compared with IL10 KO mice, DKOs showed longer colons and prolonged crypts, despite having higher inflammation and numbers of dysplasia. Crypt hyperproliferation was associated with Notch1 activation and diminished crypt differentiation, indicated by a reduction of goblet cells. Gene expression analysis indicated up-regulation of the Notch1 target hairy and enhancer of split-1 and the stem cell receptor leucin-rich repeat-containing G-protein-coupled receptor 5 in DKO mice. Interestingly, the stem cell marker olfactomedin-4 was present in colonic tissue. Increased β-catenin messenger RNA and cytoplasmic accumulation indicated aberrant Wnt signaling. Co-localization and direct interaction of Notch1 and PAK1 was found in colon epithelial cells. Notch1 activation abrogated this effect whereas silencing of PAK1 led to Notch1 activation.
Conclusions:
PAK1 contributes to the regulation of crypt homeostasis under inflammatory conditions by controlling Notch1. This identifies a novel PAK1-Notch1 axis in intestinal pathophysiology of inflammatory bowel disease and CAC.
Insights
Deleting p21-activated kinase-1 (PAK1) exacerbates intestinal inflammation and cancer in IL10 knockout mice by activating Notch1 signaling. This study reveals a novel PAK1-Notch1 axis in inflammatory bowel disease and colitis-associated cancer.
Area of Science:
- Gastroenterology
- Molecular Biology
- Oncology
Background:
- p21-activated kinase-1 (PAK1) is implicated in cellular pathways crucial for intestinal homeostasis.
- PAK1 overexpression is linked to inflammatory bowel disease and colitis-associated cancer (CAC).
- Interleukin (IL)10 knockout mice serve as a model for colitis and CAC.
Purpose of the Study:
- To investigate the role of PAK1 deletion in intestinal inflammation and carcinogenesis within the IL10 knockout mouse model.
- To elucidate the molecular mechanisms underlying PAK1's contribution to intestinal pathophysiology.
Main Methods:
- Generation of IL10/PAK1 double-knockout (DKO) mice.
- Analysis of colitis and CAC development, including colon measurements and histology.
- Gene expression analysis, co-immunoprecipitation, and immunofluorescence studies on intestinal organoids and cell lines.
Main Results:
- DKO mice exhibited prolonged colons and crypts with increased inflammation and dysplasia compared to IL10 KO mice.
- Crypt hyperproliferation in DKO mice was associated with Notch1 activation and reduced goblet cell differentiation.
- Aberrant Wnt signaling and stem cell markers (LGR5, olfactomedin-4) were observed in DKO mice.
- Direct interaction between Notch1 and PAK1 was identified in colon epithelial cells.
Conclusions:
- PAK1 regulates crypt homeostasis during inflammation by controlling Notch1 activation.
- A novel PAK1-Notch1 signaling axis is identified in the pathophysiology of inflammatory bowel disease and CAC.
- Targeting the PAK1-Notch1 pathway may offer therapeutic strategies for intestinal diseases.
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