Deficiency in class III PI3-kinase confers postnatal lethality with IBD-like features in zebrafish

Shaoyang Zhao,Jean Agnetti,Heying Li,Jianhong Xia,Leilei Zhang,Leilei Zhang,Leilei Zhang,Xiaodong Shu,Xiaodong Shu,Xiaodong Shu,Yinxiong Li,Yinxiong Li,Haiyun Wang,Haiyun Wang,Haiyun Wang,Jianhong Xia,Xiuhua Wu,Yan Chen,Yan Chen,Pengtao Wang,Pengtao Wang,Pengtao Wang,Xiaoshan Wang,Xiaoshan Wang,Duanqing Pei,Duanqing Pei,Duanqing Pei,Jean Agnetti,Heying Li,Jianhong Xia

doi:10.1038/s41467-018-05105-8

Abstract

The class III PI3-kinase (PIK3C3) is an enzyme responsible for the generation of phosphatidylinositol 3-phosphate (PI3P), a critical component of vesicular membrane. Here, we report that PIK3C3 deficiency in zebrafish results in intestinal injury and inflammation. In pik3c3 mutants, gut tube forms but fails to be maintained. Gene expression analysis reveals that barrier-function-related inflammatory bowel disease (IBD) susceptibility genes (e-cadherin, hnf4a, ttc7a) are suppressed, while inflammatory response genes are stimulated in the mutants. Histological analysis shows neutrophil infiltration into mutant intestinal epithelium and the clearance of gut microbiota. Yet, gut microorganisms appear dispensable as mutants cultured under germ-free condition have similar intestinal defects. Mechanistically, we show that PIK3C3 deficiency suppresses the formation of PI3P and disrupts the polarized distribution of cell-junction proteins in intestinal epithelial cells. These results not only reveal a role of PIK3C3 in gut homeostasis, but also provide a zebrafish IBD model.

Highlights

The class III PI3-kinase (PIK3C3) is an enzyme responsible for the generation of phosphatidylinositol 3-phosphate (PI3P), a critical component of vesicular membrane
Mutations in tetratricopeptide repeat domain-7A (TTC7A) gene were identified from patients of very early onset inflammatory bowel disease (IBD) (VEO-IBD) or multiple intestinal atresia (MIA), which result in reduced cell adhesion, elevated apoptosis, defective epithelial barrier and polarity[8,9]
Among the 200+ independent genetic risk loci for IBD identified by GWAS10,11, there are loci that have nearby genes involved in bacterial sensing and autophagy, such as nucleotide-binding oligomerization domain containing 2 (NOD2), autophagy-related 16 like 1 (ATG16L1) and immunity-related GTPase M (IRGM) or inflammatory response, such as interleukin 23 receptor (IL23R), caspase recruitment domain family member 9 (CARD9), matrix metallopeptidase 9 (MMP9), tumor necrosis factor (TNF) and interleukin 1 beta (IL1B) and they are suggested to be causal genetic variants[12,13,14]

Summary

Introduction

The class III PI3-kinase (PIK3C3) is an enzyme responsible for the generation of phosphatidylinositol 3-phosphate (PI3P), a critical component of vesicular membrane. We show that PIK3C3 deficiency suppresses the formation of PI3P and disrupts the polarized distribution of cell-junction proteins in intestinal epithelial cells These results reveal a role of PIK3C3 in gut homeostasis, and provide a zebrafish IBD model. Intestinal specific knockout of adherens junction protein E-Cadherin (encoded by the Cdh[1] gene) in mice disrupts the barrier function of intestine and induces epithelial cell death and defective bacterial defensing[3,4]. Conditional knockout of this gene in the intestine of adult mice induces down-regulation of the tight junction protein zonula occludens 1 protein (ZO-1), cytoplasmic mis-localization of E-Cadherin and destabilization of epithelial cell–cell junctions[5] Both CDH1 and HNF4A were identified as susceptibility loci in UC patients by genome-wide association studies (GWAS)[6]. We propose that the zebrafish pik3c3 mutant may serve as a disease model for mechanistic studies of epithelial damage-induced IBD and a platform for developing and evaluating potential therapeutic interventions

Methods

Results

Conclusion