Study on the role of MYH9 and its interacting proteins in esophageal cancer cells

Abstract

Objective This paper aims to study the effect of MYH9 on the motility of esophageal squamous cell carcinoma (ESCC) cells, and to identify the interaction protein of the tail domain (TD) of MYH9. Methods MYH9 was knocked down by siRNA in ESCC KYSE510 cells, and the ability of cell invasion and migration was detected by transwell assay, besides the ability of cell wound healing was detected by wound healing assay. The recombinant plasmid pGEX-KG-MYH9(TD) was constructed to induce the expression and affinity purification of GST-MYH9 (TD) fusion protein. MYH9 (TD) interaction proteins were identified and analyzed by GST pull-down combined with liquid chromatography-tandem mass spectrometry (LC-MS/MS). ClusterProfiler R language data processing package was used for functional annotation of potential interacting proteins.Furthermore, Co-immunoprecipitation(Co-IP) was exploited to verify the interaction of MYH9 and its interacting proteins. Results MYH9 knockdown significantly inhibited the invasion and migration(P<0.001) and wound healing of ESCC cells. A total of 72 potential interacting proteins were identified in KYSE510 cells by MS. Gene Oncology (GO) enrichment analysis revealed that potential interacting proteins were involved in a variety of biological processes, including the JAK-STAT signaling pathway, cell adhesion regulation, NIK/NF-κB signaling pathway, cell cycle checkpoints, lipid raft formation, DNA transcription initiation, and mRNA processing, etc. Co-IP results confirmed that PSIP1 and BRG1 interacted with MYH9. Conclusion MYH9 promotes the motility of ESCC cells, and MYH9 tail domain mediates the interaction of various motion-related proteins. Key words: Esophageal squamous cell carcinoma(ESCC); MYH9 tail domain; Invasion and migration; Interacting protein