Small GTPase Rab37 targets tissue inhibitor of metalloproteinase 1 for exocytosis and thus suppresses tumor metastasis

  • 蔡 宗翰

Student thesis: Doctoral Thesis

Abstract

Background & Aims: The five year survival rate for lung cancer patients remains at 13-15% over the past years However the mechanisms involved in lung cancer progression remain unclear Recent publications show alterations of Rab small GTPase-controlled vesicle traffic during tumorigenesis However whether any of the Rabs plays a metastasis suppressive role is least explored Our previous data demonstrates for the first time that low mRNA expression and promoter hypermethyaltion of human Rab37 (hRAB37) gene correlates with lung cancer progression Therefore the current study aims to investigate the functions of hRAB37 to regulate vesicle trafficking and its signal pathways involved in cell migration/invasion and tumor metastasis Methods: We performed a proteomics screen of conditioned medium to identify one of the cargos of hRAB37 namely tissue inhibitor of metalloproteinase 1 (TIMP1) The hRAB37-mediated TIMP1 exocytosis was determined by confocal immuno-electron microscopy (EM) and total internal reflection fluorescence imaging (TIRF) The biological effects of hRAB37 on tumor metastasis and TIMP1-metalloproteinase 9 (MMP9) signaling were detected in cultured cells and nude mice by overexpression or knockdown of hRAB37 We analyzed protein expression of hRAB37 and TIMP1 by immunohistochemistry and fluorescence-immunohistochemistry in 165 lung cancer patients and mRNA expression of hRAB37 and TIMP1 in 450 lung cancer patients derived from the The Cancer Genome Atlas (TCGA) dataset Results: Our confocal analysis of trafficking markers indicated that hRAB37 co-localized with secretory granule markers and involved in the exocytotic pathway Co-localization of TIMP1 in hRAB37-containing vesicles was confirmed by immuno-EM assay Our TIRF image analysis provided the dynamic views of hRAB37-mediated trafficking and docking of TIMP1-containing vesicles to the plasma member Overexpression of hRAB37 resulted in loss of migration/invasion ability and MMP9 activity in CL1-5 lung cancer cells-based assays whereas knockdown of hRAB37 in H460 lung cancer cells increased the migration/invasion ability and MMP9 activity Notably inactive or knockdown of hRAB37 remarkably promoted lung tumor metastasis in tail vain injection and lung-to-lung tumor metastasis animal models Interestingly TIMP1 knockdown increased the migration/invasion ability and MMP9 activity in hRAB37 expression cells which were confirmed by animal metastasis assays These data suggested that TIMP1 is essential to hRAB37-mediated metastasis suppression Clinically lung cancer patients with low hRAB37 showed concordantly reduced TIMP1 (P
Date of Award2016 Feb 16
Original languageEnglish
SupervisorYi-Ching Wang (Supervisor)

Cite this

Small GTPase Rab37 targets tissue inhibitor of metalloproteinase 1 for exocytosis and thus suppresses tumor metastasis
宗翰, 蔡. (Author). 2016 Feb 16

Student thesis: Doctoral Thesis