The PEA3 Ets transcription factor is a downstream target of the HER2/Neu receptor tyrosine kinase

The HER2/neu gene, which is overexpressed in 20-30% of human breast tumors, encodes a receptor tyrosine kinase that functions through multiple signaling pathways to regulate the activity of nuclear transcription factors. We have reported that PEAS, an Ets family transcription factor, is overexpressed in HER2/Neu-induced breast tumors and their metastases. To account for the increased levels of PEA3 in these turners we have suggested that HER2/Neu enhances PEAS transcriptional activity, which then acts to stimulate expression of the PEAS gene. This hypothesis is consistent with the occurrence of PEAS binding sites in the PEAS promoter and with the ability of PEA3 to transactivate this promoter. To learn whether HER2/Neu indeed regulates PEAS activity we measured the capacity of constitutively-activated HER2/Neu to affect PEAS-dependent reporter gene expression. Coexpression of PEA3 and HER2/Neu stimulated PEA3-dependent reporter gene expression to a much greater extent than did either protein alone suggesting that HER2/Neu upregulates the transcriptional activity of PEAS. To define the pathway whereby HER2/Neu functions we employed dominant-negative mutants of signaling proteins known to be downstream of HER2/Neu. Overexpression of Rap1a, a Pas-related protein capable of antagonizing Pas function, completely inhibited the ability of HER2/Neu to stimulate PEAS-dependent gene expression. Pas is known to stimulate at least two mitogen-activated protein kinase (MAPK) cascades, the extracellular-regulated kinase (ERK) cascade and the stress-activated kinase (SAPK) or Jun kinase (JNK) cascade. Similarly, HER2/Neu activated both ERKs and SAPKs/JNKs in a Pas-dependent fashion. Dominant-inhibitory mutants in either the ERK or SAPK/JNK cascades partially inhibited HER2/Neu activation of PEAS-dependent gene expression. These findings suggest that HER2/Neu regulates PEA3 activity through two different Ras-dependent MAPK pathways.