Identification of mitogen-activated protein kinase kinase as a chemoresistant pathway in MCF-7 cells by using gene expression microarray.
BACKGROUND
Components of the mitogen-activated protein kinase (MAPK) cascade have been implicated in apoptotic regulation. This study used gene expression profiling analysis to identify and implicate mitogen-activated protein kinase kinase (MEK5)-BMK1 (big mitogen-activated kinase-1)/extracellular signal related protein kinase (ERK5) pathway as a novel target involved in chemoresistance.


METHODS
Differential gene expression between apoptotically sensitive (APO+) and apoptotically resistant (APO-) MCF-7 cell variants was determined by using microarray and confirmed by reverse transcriptase- polymerase chain reaction (RT-PCR). An apoptotic/viability reporter gene assay was used to deter-mine the effects of the transfection of a dominant-negative mutant of BMK1 (BMK1/DN) in conjunction with apoptotic-inducing agents (etoposide, tumor necrosis factor-alpha [TNF], or TNF-related apoptosis-inducing ligand [TRAIL]), with or without phorbol ester (PMA).


RESULTS
Of the 1186 genes detected through microarray analysis, MEK5 was increased 22-fold in APO- cells. Overexpression of MEK5 was confirmed by using RT-PCR analysis. Expression of BMK1/DN alone resulted in a dose-dependent increase in cell death versus control (P <.05). In addition, BMK1/DN enhanced the sensitivity of MCF-7 cells to treatment-induced cell death (P <.05). The ability of PMA to partially suppress TRAIL- and TNF-induced cell death was inhibited by BMK1/DN. However, only TRAIL-induced activity suppression reached statistical significance (P <.05).


CONCLUSIONS
The overexpression of MEK5 in APO- MCF-7 breast carcinoma cells shows that this MAPK signaling protein represents a potent survival molecule. Molecular inhibition of MEK5 signaling may represent a mechanism for sensitizing cancer cells to chemotherapeutic regimens.
Weldon
Christopher B
CB
Department of Surgery, Tulane University School of Medicine and Tulane University School of Public Health & Tropical Medicine, New Orleans 70112, USA.
Scandurro
Ali B
AB
Rolfe
Kevin W
KW
Clayton
John L
JL
Elliott
Steven
S
Butler
Nancy N
NN
Melnik
Lilia I
LI
Alam
Jawed
J
McLachlan
John A
JA
Jaffe
Bernard M
BM
Beckman
Barbara S
BS
Burow
Matthew E
ME
D000970:Antineoplastic Agents / Q000494:pharmacology; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D051017:Apoptosis Regulatory Proteins; D001943:Breast Neoplasms; D002273:Carcinogens / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D005047:Etoposide / Q000494:pharmacology; D005260:Female; D006801:Humans; D048671:MAP Kinase Kinase 5; D008562:Membrane Glycoproteins / Q000494:pharmacology; D048054:Mitogen-Activated Protein Kinase 7; D020929:Mitogen-Activated Protein Kinase Kinases / Q000235:genetics*; D020928:Mitogen-Activated Protein Kinases / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D010703:Phorbol Esters / Q000494:pharmacology; D053221:TNF-Related Apoptosis-Inducing Ligand; D014407:Tumor Cells, Cultured / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D000970:Antineoplastic Agents; D000972:Antineoplastic Agents, Phytogenic; D051017:Apoptosis Regulatory Proteins; D002273:Carcinogens; D008562:Membrane Glycoproteins; D010703:Phorbol Esters; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; D014409:Tumor Necrosis Factor-alpha; D005047:Etoposide; D048054:Mitogen-Activated Protein Kinase 7; D020928:Mitogen-Activated Protein Kinases; D048671:MAP Kinase Kinase 5; C482563:MAP2K5 protein, human; D020929:Mitogen-Activated Protein Kinase Kinases