Abstract
The protozoan parasite Trypanosoma cruzi, the causative agent of Chagas disease, causes severe morbidity and mortality in afflicted individuals. About 30% of T. cruzi-infected individuals present with cardiac, gastrointestinal tract, and/or neurological disorders. Megacolon, one of the major pathologies of Chagas disease, is accompanied by gastrointestinal motility disorders. The molecular mechanism of T. cruzi-mediated megacolon in Chagas disease is currently unknown. To decipher the molecular mechanism of T. cruzi-induced alteration in the colon during the early infection phase, we exposed primary human colonic epithelial cells (HCoEpiC) to invasive T. cruzi trypomastigotes at multiple time points to determine changes in the phosphoprotein networks in the cells following infection using proteome profiler Human phospho-kinase arrays. We found significant changes in the phosphorylation pattern that can mediate cellular deregulations in colonic epithelial cells after infection. We detected a significant increase in the levels of phosphorylated heat shock protein (p-HSP) 27 and transcription factors that regulate various cellular functions, including c-Jun and CREB. Our study confirmed significant upregulation of phospho (p-) Akt S473, p-JNK, which may directly or indirectly modulate CREB and c-Jun phosphorylation, respectively. We also observed increased levels of phosphorylated CREB and c-Jun in the nucleus. Furthermore, we found that p-c-Jun and p-CREB co-localized in the nucleus at 180 minutes post infection, with a maximum Pearson correlation coefficient of 0.76±0.02. Increased p-c-Jun and p-CREB have been linked to inflammatory and profibrotic responses. T. cruzi infection of HCoEpiC induces an increased expression of thrombospondin-1 (TSP-1), which is fibrogenic at elevated levels. We also found that T. cruzi infection modulates the expression of NF-kB and JAK2-STAT1 signaling molecules which can increase pro-inflammatory flux. Bioinformatics analysis of the phosphoprotein networks derived using the phospho-protein data serves as a blueprint for T. cruzi-mediated cellular transformation of primary human colonic cells during the early phase of T. cruzi infection.
Highlights
The protozoan parasite Trypanosoma cruzi is the causative agent of Chagas disease, a neglected tropical disease which causes severe morbidity and mortality worldwide
We challenged primary human colonic epithelial cells with T. cruzi and evaluated changes in the phosphorylated kinases and phosphoprotein levels that may induce cellular and molecular alterations leading to cellular transformations during the early phase of infection
The parasite enhanced the levels of p-AKT, p-HSP27, p-JNK, and downstream transcription factors like p-c-Jun and p-CREB during the early infection phase
Summary
The protozoan parasite Trypanosoma cruzi is the causative agent of Chagas disease, a neglected tropical disease which causes severe morbidity and mortality worldwide. It is generally agreed that the enteric neurons [7, 8] and interstitial cells of Cajal [8, 9] decrease in numbers in megacolon, it is unclear what roles they play in the pathophysiology of chagasic megacolon. The fibrotic lesions observed in megacolon tissue sections can be caused by increased deposition of extracellular matrix (ECM) and matricellular proteins including TSP-1. The interactions between T. cruzi and colon cells including colon epithelium cells can deregulate cell signaling pathways leading to increased expression of transcription factors that upregulate the synthesis of ECM proteins [10, 11] causing fibrogenesis and cellular transformation reported in megacolon tissue sections. The role played by colon epithelium in the onset of chagasic megacolon remains unknown
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