Sp1 and AP-1 regulate expression of the human gene VIL2 in esophageal carcinoma cells.
Ezrin, encoded by VIL2, is a membrane-cytoskeletal linker protein that has been suggested to be involved in tumorigenesis. Ezrin expression in esophageal squamous cell carcinoma (ESCC) was described recently, but its clinical significance and the molecular mechanism underlying its regulated expression remain unclear. Thus, we retrospectively evaluated ezrin expression by immunohistochemistry in a tissue microarray representing 193 ESCCs. Ezrin overexpression in 90 of 193 tumors (46.6%) was associated with poor survival (p = 0.048). We then explored the mechanism by which ezrin expression is controlled in ESCC by assessing the transcriptional regulatory regions of human VIL2 by fusing deletions or site-directed mutants of the 5'-flanking region of the gene to a luciferase reporter. We found that the region -87/-32 containing consensus Sp1 (-75/-69) and AP-1 (-64/-58) binding sites is crucial for VIL2 promoter activity in esophageal carcinoma cells (EC109) derived from ESCC. AP-1 is comprised of c-Jun and c-Fos. Electrophoretic mobility shift and chromatin immunoprecipitation experiments demonstrated that Sp1 and c-Jun bound specifically to their respective binding sites within the VIL2 promoter. In addition, transient expression of Sp1, c-Jun, or c-Fos increased ezrin expression and VIL2 promoter activity. Use of selective inhibitors revealed that VIL2 transactivation required the MEK1/2 signal transduction pathway but not JNK or p38 MAPK. Taken together, we propose a possible signal transduction pathway whereby MEK1/2 phosphorylates ERK1/2, which phosphorylates Sp1 and AP-1 that in turn bind to their respective binding sites to regulate the expression of human VIL2 in ESCC cells.
Gao
Shu-Ying
SY
Department of Biochemistry and Molecular Biology, Shantou University, Shantou, China.
Li
En-Min
EM
Cui
Lei
L
Lu
Xiao-Feng
XF
Meng
Ling-Ying
LY
Yuan
Hua-Min
HM
Xie
Jian-Jun
JJ
Du
Ze-Peng
ZP
Pang
Jian-Xin
JX
Xu
Li-Yan
LY
D000328:Adult; D000368:Aged; D045744:Cell Line, Tumor; D003598:Cytoskeletal Proteins / Q000096:biosynthesis* / Q000235:genetics; D018572:Disease-Free Survival; D004938:Esophageal Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000401:mortality; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D048369:MAP Kinase Kinase 1 / Q000235:genetics / Q000378:metabolism; D048370:MAP Kinase Kinase 2 / Q000235:genetics / Q000378:metabolism; D048670:MAP Kinase Kinase 4 / Q000235:genetics / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000235:genetics; D008297:Male; D046228:Microarray Analysis; D008875:Middle Aged; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D018307:Neoplasms, Squamous Cell / Q000235:genetics / Q000378:metabolism* / Q000401:mortality; D010766:Phosphorylation / Q000235:genetics; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051076:Proto-Oncogene Proteins c-fyn / Q000235:genetics / Q000378:metabolism; D016755:Proto-Oncogene Proteins c-jun / Q000235:genetics / Q000378:metabolism; D020218:Response Elements / Q000235:genetics; D012189:Retrospective Studies; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D015996:Survival Rate; D018808:Transcription Factor AP-1 / Q000235:genetics / Q000378:metabolism*; D048051:p38 Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism; D000099149:Ezrin
D003598:Cytoskeletal Proteins; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; D048670:MAP Kinase Kinase 4; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D047428:Protein Kinase Inhibitors; D051076:Proto-Oncogene Proteins c-fyn; D016755:Proto-Oncogene Proteins c-jun; D016329:Sp1 Transcription Factor; D018808:Transcription Factor AP-1; D048051:p38 Mitogen-Activated Protein Kinases; D000099149:Ezrin; C482128:MAP2K2 protein, human; C482125:MAP2K1 protein, human