An EGR2/CITED1 Transcription Factor Complex and the 14-3-3σ Tumor Suppressor Are Involved in Regulating ErbB2 Expression in a Transgenic-Mouse Model of Human Breast Cancer

Abstract

Amplification and elevated expression of the ErbB2 receptor tyrosine kinase occurs in 20% of human breast cancers and is associated with a poor prognosis. We have previously demonstrated that mammary tissuespecific expression of activated ErbB2 under the control of its endogenous promoter results in mammary tumor formation. Tumor development was associated with amplification and overexpression of ErbB2 at both the transcript and protein levels. Here we demonstrate that the EGR2/Krox20 transcription factor and its coactivator CITED1 are coordinately upregulated during ErbB2 tumor induction. We have identified an EGR2 binding site in the erbB2 promoter and demonstrated by chromatin immunoprecipitation assays that EGR2 and CITED1 associate specifically with this region of the promoter. EGR2 and CITED1 were shown to associate, and expression from an erbB2 promoter-reporter construct was stimulated by EGR2 and was further enhanced by CITED1 coexpression. Furthermore, expression of the 14-3-3 tumor suppressor led to downregulation of ErbB2 protein levels and relocalization of EGR2 from the nucleus to the cytoplasm. Taken together, these observations suggest that, in addition to an increased gene copy number and upregulation of EGR2 and CITED1, an elevated erbB2 transcript level involves the loss of 14-3-3, which sequesters a key transcriptional regulator of the erbB2 promoter.

Loss of the 14-3-3σ Tumor Suppressor Is a Critical Event in ErbB2-Mediated Tumor Progression

“…One likely explanation for these observations is that 14-3-3σ negatively regulates transcription factors involved in regulating ErbB2 expression. Consistent with this concept, one previous study has demonstrated that 14-3-3σ is involved in sequestration of the EGR2 transcription factor that directly activates the erbB2 promoter (16). Given that the expression of the activated erbB2 allele is driven by the endogenous erbB2 promoter in the ErbB2 KI model, there is strong selective pressure for the loss of 14-3-3σ expression.…”

“…It has also been demonstrated that all human ErbB2-overexpressing breast cancer cell lines have no or reduced 14-3-3σ expression (25). Further molecular analyses suggested that selection for loss of 14-3-3σ may in part be related to its ability to sequester the EGR2/ CITED1 transcription factor complex in the cytoplasm and thus interfere with its capacity to transactivate the endogenous erbB2 promoter (16). Taken together, these data argue that, in addition to sequestration of key translation and cellcycle components, 14-3-3σ can directly modulate the transcription of ErbB2 through relocalization of key transcription factors involved in its transcriptional activation.…”

“…Given that the average tumor chromosome 4 (15). Refined CGH/BAC analyses revealed that in addition to the erbB2 amplicon, there was a frequent loss of a chromosomal region spanning the 14-3-3σ tumor suppressor locus (16,17). The region of loss on mouse chromosome 4 is syntenic with human chromosome 1p35-36, which is also frequently lost in human sporadic breast cancer (18,19).…”

“…Given the documented roles of 14-3-3σ in the regulation of epithelial polarity, ErbB2 expression, and ErbB2 tumor progression (16,17,28), we evaluated whether loss of 14-3-3σ in the mammary epithelium would impact ErbB2-driven tumor induction. To directly assess whether 14-3-3σ acts as a tumor suppressor in vivo, we assessed whether mammary epithelial disruption of 14-3-3σ could impact mammary tumor progression in the ErbB2 KI mouse strain.…”

“…with the effects of 14-3-3σ on ErbB2 transcription (16) and MAPK inactivation (Fig. 4) accounts for a strong selection for 14-3-3σ silencing during breast cancer progression.…”