CADM1 Interacts with Tiam1 and Promotes Invasive Phenotype of Human T-cell Leukemia Virus Type I-transformed Cells and Adult T-cell Leukemia Cells

Yoshinori Murakami,Eric J. Stanbridge,Mari Masuda,Tomoko Maruyama,Toshiki Watanabe,Hiroo Hoshino,Kunihiko Tsukasaki,Shimeru Kamihira,Tsutomu Ohta,Tomayoshi Hayashi,Akihiko Ito,Teruhiko Yoshida,Shoji Yamaoka

doi:10.1074/jbc.m109.076653

Abstract

CADM1 encodes a multifunctional immunoglobulin-like cell adhesion molecule whose cytoplasmic domain contains a type II PSD95/Dlg/ZO-1 (PDZ)-binding motif (BM) for associating with other intracellular proteins. Although CADM1 lacks expression in T lymphocytes of healthy individuals, it is overexpressed in adult T-cell leukemia-lymphoma (ATL) cells. It has been suggested that the expression of CADM1 protein promotes infiltration of leukemic cells into various organs and tissues, which is one of the frequent clinical manifestations of ATL. Amino acid sequence alignment revealed that Tiam1 (T-lymphoma invasion and metastasis 1), a Rac-specific guanine nucleotide exchange factor, has a type II PDZ domain similar to those of membrane-associated guanylate kinase homologs (MAGUKs) that are known to bind to the PDZ-BM of CADM1. In this study, we demonstrated that the cytoplasmic domain of CADM1 directly interacted with the PDZ domain of Tiam1 and induced formation of lamellipodia through Rac activation in HTLV-I-transformed cell lines as well as ATL cell lines. Our results indicate that Tiam1 integrates signals from CADM1 to regulate the actin cytoskeleton through Rac activation, which may lead to tissue infiltration of leukemic cells in ATL patients.

Highlights

CADM1 encodes a multifunctional immunoglobulin-like cell adhesion molecule whose cytoplasmic domain contains a type II PSD95/Dlg/ZO-1 (PDZ)-binding motif (BM) for associating with other intracellular proteins
Amino acid sequence alignment revealed that Tiam1 (T-lymphoma invasion and metastasis 1), a Rac-specific guanine nucleotide exchange factor, has a type II PDZ domain similar to those of membrane-associated guanylate kinase homologs (MAGUKs) that are known to bind to the PDZ-binding motif (PDZ-BM) of CADM1
In an attempt to elucidate the role of CADM1 in adult T-cell leukemia-lymphoma (ATL) cells, we performed data base homology searches of all NCBI sequences looking for cytoplasmic proteins with type II PDZ domains similar to the PDZ domains of MAGUKs and found T-lymphoma invasion and metastasis 1 (Tiam1), which is a Rac-specific guanine nucleotide exchange factor (GEF)

Summary

EXPERIMENTAL PROCEDURES

Cells—Jurkat, Molt-4, CCRF-CEM, T-all, and CEM/C2 cell lines derived from acute lymphoblastic T-cell leukemia patients and H9 and Hut derived from cutaneous T-cell lymphoma were obtained from the American Type Culture Collection (ATCC). All T cell lines described above were maintained in RPMI medium (Sigma) supplemented with 10% Tet system-approved fetal bovine serum (Takara Bio), 100 units/ml penicillin, and 100 ␮g/ml streptomycin (Invitrogen). A rabbit pAb to Tiam (C16) was purchased from Santa Cruz Biotechnology, Inc. Secondary antibodies used for immunoblot analysis were from GE Healthcare. The constructs for the N-terminal glutathione S-transferase (GST)-tagged cytoplasmic domain of CADM1 (GSTCADM1-C) and the truncated mutant lacking the C-terminal 3 amino acids (GST-CADM1-C⌬3) were described previously [15]. Transfection—To obtain Jurkat Tet-Off cell clones expressing full-length CADM1 or the mutant lacking the cytoplasmic domain of CADM1, cells were transfected with pTRE2/CADM1 or pTRE2/⌬C-HA using Fugene 6 (Roche Applied Science) and selected in a medium containing 300 ␮g/ml hygromycin (Invitrogen). Immunohistochemical analyses of formalin-fixed and paraffin-embedded tissue sections were performed as described previously [31, 32]

RESULTS

Induces Formation of Lamellipodia through Rac Activation in Jurkat

DISCUSSION