The transcription factor Fra-2 promotes mammary tumour progression by changing the adhesive properties of breast cancer cells
Introduction
Activating protein-1 (AP-1) is a transcription factor complex composed of members of the Jun, Fos, and activating transcription factor (ATF) family that bind as hetero or homodimers to the regulatory sequences of various target genes. Inhibition of AP-1 activity results in blocked proliferation, migration, invasion and experimental metastasis of tumour cells in various systems.1
The transcription factor Fra-2 (Fos-related antigen 2) was identified as a serum-inducible gene with homologies to c-Fos, FosB and Fra-1, but with significantly lower transforming activity compared with c-Fos.2, 3 Like the other members of this Fos family, Fra-2 forms heterodimers with Jun family members. In contrast to c-Fos and FosB, but similar to Fra-1, Fra-2 proteins lack a C-terminal transactivating domain and do not stimulate artificial AP-1-responsive promoters in vitro.4, 5 Animal models have shown that Fra-2 is required for proper bone and cartilage development, and Fra-2 −/− pups exhibit growth retardation and die within one week after birth.6 On the other hand, ectopic expression of Fra-2 in transgenic mice results in extensive fibrosis, predominantly in the lung, leading to premature mortality.7 Investigations on human clinical tissues indicate that Fra-2 might play an important role in the progression of various human tumour types in vivo.8 Fra-2 overexpression was found in salivary gland tumours9, colorectal cancer10 and adult T-cell leukaemia11, and in anaplastic large cell lymphomas. In the latter case, Fra-2 is amongst the genes which are affected by the characteristic t2;5 translocation leading to enhanced Fra-2 expression in the tumour cells.12
In breast cancer cell lines and tumour tissues, Fra-2 is detectable in variable amounts and phosphorylation states.13 In a Western blot study with 75 breast cancer samples, we found a significant association of strong Fra-2 expression with a high frequency of recurrence.14 In experimental models, Fra-2 expression is associated with an increased invasive potential and motility, whereas proliferation was hardly influenced at all.14, 15 In mouse mammary adenocarcinoma cell lines (CSMLO) with different metastatic potential, strong Fra-1 and Fra-2 expression was present in the metastatic cells, whereas c-Fos and FosB were undetectable.16 In transfection experiments with these cell lines, overexpression of Fra-2 resulted in activation of osteopontin, thrombospondin and CD44 which are all known to be involved in metastasis.17
In order to further investigate the role of Fra-2 in breast cancer progression, Fra-2 mRNA expression was analysed in a cohort of 164 mammary carcinomas with long-term follow-up data. In addition, we performed stable transfection leading to Fra-2 overexpression in the ER-negative MDA-MB231 cells. Amongst the genes which are differentially regulated in these cells, we identified a number of genes involved in cell–cell or cell–ECM interaction which suggested that the effect of Fra-2 is at least partly due to its influence on cell adhesion, especially during extravasation from the bloodstream. This hypothesis was further confirmed by ECM adhesion assays and analysis of attachment to E-selectin-coated surfaces in a laminar flow system. The correlation of Fra-2 expression with two relevant Fra-2 target genes, the cell adhesion molecules ICAM-1 and L1-CAM, was further analysed in our cohort of clinical breast cancer samples.
Section snippets
Patients
For array-based Fra-2 mRNA detection, samples from 167 patients (mean age 56.2 years; range 29–85 years) were analysed. All patients were treated for breast cancer at the University Medical Center Hamburg-Eppendorf, Germany, Department of Gynecology, between the years 1991 and 2002. Patient selection was based upon availability of tumour tissue. All patients gave written informed consent to access their tissues and review their medical records in accordance with the principles of the
Fra-2 expression in clinical tumour tissues
Fra-2 (FOSL2) mRNA expression in 167 breast cancer samples was evaluated by microarray analysis. Since the Affymetrix genchips harboured three probesets for FOSL2 (205409_at, 218880_at and 218881_s_at), we first compared the expression data of these sets and found highly significant correlations between all of them (p < 0.001; not shown). Since similar correlations with established prognostic markers were found for all probesets, we chose one of them (218881_s_at) for further analysis. Expression
Discussion
This study was undertaken to examine the role of Fra-2 expression in malignant progression of breast cancer in an experimental system and clinical tumours. Similar to our prior western blot study with a smaller cohort14, the present analysis of RNA expression data of 167 breast cancer samples has shown that high Fra-2 expression is associated with nodal involvement and early relapse, suggesting that Fra-2 might regulate target genes which are involved in the metastatic cascade in breast cancer.
Conflict of interest statement
Ralph M. Wirtz is employed at Siemens Healthcare Diagnostics Products GmbH, Cologne, Germany. For the other authors, there are no conflicting interests.
Acknowledgements
We gratefully acknowledge the excellent technical assistance of Katrin Beck and Sylke Krenkel and we thank the department directors Fritz Jänicke and Thomas Löning for generous support. Ulrich Richter is supported by a scholarship from the Werner Otto Foundation, which is gratefully acknowledged. This work was supported by the Deutsche Krebshilfe, Grant No. 106110.
References (39)
The role of the Fos family of transcription factors in tumorigenesis
Eur J Cancer
(2005)- et al.
The breast cancer beta 4 integrin and endothelial human CLCA2 mediate lung metastasis
J Biol Chem
(2001) - et al.
L1-CAM in a membrane-bound or soluble form augments protection from apoptosis in ovarian carcinoma cells
Gynecol Oncol
(2007) - et al.
ICAM-1 signaling in endothelial cells
Pharmacol Rep
(2009) - et al.
Transcription factors control invasion: AP-1 the first among equals
Oncogene
(2007) - et al.
Isolation of human fos-related genes and their expression during monocyte–macrophage differentiation
Oncogene
(1990) - et al.
Isolation and characterization of fra-2, an additional member of the fos gene family
Proc Natl Acad Sci USA
(1990) Fos family members: regulation, structure and role in oncogenic transformation
Histol Histopathol
(2000)- et al.
Transformation by fos proteins requires a C-terminal transactivation domain
Mol Cell Biol
(1993) - et al.
Simultaneous generation of fra-2 conditional and fra-2 knock-out mice
Genesis
(2007)
Development of pulmonary fibrosis through a pathway involving the transcription factor Fra-2/AP-1
Proc Natl Acad Sci USA
Gene expression screening of salivary gland neoplasms
J Mol Diagn
Differential expression of the AP-1 transcription factor family members in human colorectal epithelial and neuroendocrine neoplasms
Am J Clin Pathol
Aberrant expression of Fra-2 promotes CCR4 expression and cell proliferation in adult T-cell leukemia
Oncogene
Gene deregulation and spatial genome reorganization near breakpoints prior to formation of translocations in anaplastic large cell lymphoma
Proc Natl Acad Sci USA
Expression pattern of the AP-1 family in breast cancer: association of fosB expression with a well-differentiated, receptor-positive phenotype
Int J Cancer
The role of the AP-1 transcription factors c-Fos, FosB, Fra-1 and Fra-2 in the invasion process of mammary carcinomas
Breast Cancer Res Treat
Role of Fra-2 in breast cancer: influence on tumor cell invasion and motility
Breast Cancer Res Treat
Fra-1 induces morphological transformation and increases in vitro invasiveness and mobility of epithelioid adenocarcinoma cells
Mol Cell Biol
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