Research On The Biological Characteristics Of A Novel RON Splice Variant Lacking Exon 2

Background:Abnormal expression of the Recepteur d’Origine Nantais(RON)receptor tyrosine kinase is accompanied by the generation of multiple splice or truncated variants,which mediates many critical cellular functions that contribute to tumor progression and metastasis.Here,we report a new RON splice variant in the human colorectal cancer(CRC)cell line HT29.This variant is a 165 kda protein generated by alternative pre-mRNA splicing that eliminates exon 2,causing an in-frame deletion of 63 amino acids in the extracellular domain of the RON β chain.The deleted transcript is a single chain expressed in the intracellular compartment,lacking tyrosine phosphorylation activity.Objective and methods:By constructing the wild type or the new RON variant plasmid vector,and then transfecting them into HEK293 cell,we established two kinds of protein expression systems.Using immunoprecipitation assay,immunoblotting assay,cell wound scratch assay and animal experiments model,we learned more about the biological characteristics of the novel variant RONA165E2.All results in this experiment will be an important step for disclosing the mysteries mechanisms of the targeted therapy failure in CRC patients.Results:Compared with the wild type RON protein,the RON△165E2 variant could phosphorylate phosphatase and tensin homolog(PTEN),thereby activate the PI3K/Akt pathway in HEK293 cell.In addition,in vitro and in vivo experiments showed that the RON△165E2 promoted cell migration and tumor growth.Finally,in an investigation of 67 clinical CRC samples,the variant was highly expressed in about 58%of the samples,and was positively correlated with the invasive depth of the tumor(P<0.05).These results demonstrate that the novel RON△165E2 variant promoted tumor progression while activating the PI3K/Akt pathway via PTEN phosphorylation.