Introduction Fra-1 is a member of the Fos family of proteins which can combine with Jun proteins to form the Activator Protein-1 (AP-1) transcription factor complex. Genes regulated by AP-1 include several associated with malignant transformation and it is now known that Fra-1 becomes the predominant Fos member of the AP-1 complex in several solid malignancies. Here, we have assessed the expression and immunolocalisation of Fra-1 in human muscle-invasive bladder cancer. Materials and Methods Initially, we assessed the human bladder cancer cells lines HT1376, J82, RT4, RT112, T24 and UMUC3 by Western Blot analysis for Fra-1 (Fra-1 R20, Santa Cruz Biotechnology). These cells were subsequently transfected with the pCGN-HAFra-1WT vector (generated in our laboratory), containing an in-frame N-terminal HA-tagged full length Fra-1 construct. Using standard immunocytostaining, these transfected cells were stained using anti-HA antibody (HA Y-11, Santa-Cruz Biotechnology) and visualised using fluorescence and confocal microscopy. Subsequently, paraffin sections from 104 muscle-invasive bladder tumours were stained for Fra-1 using the Fra-1 R20 antibody. Fra-1 staining was scored as absent, weak (+), moderate (++) or strong (+++). Results J82, T24 and UMUC3 cells demonstrated endogenous Fra-1 expression; HT1376, RT4 and RT112 did not. Western Blot analysis using anti-Fra-1 and anti-HA antibodies demonstrated expression of the transfected pCGN-HAFra-1WT vector in all six cell lines. Immunocytostaining for HA-tagged Fra-1 detected expression of HAFra-1WT in both the nucleus and cytoplasm of the transfected cells. 60/104 (58%) tumour sections stained positively for Fra-1. This staining was predominantly nuclear, but two tumours had unequivocal cytoplasmic staining and a further two had equivocal cytoplasmic immunoreactivity. Discussion To the best of our knowledge, this is the first report of Fra-1 expression in human bladder cancer. The data show that Fra-1, which regulates genes involved in tumour progression, is present in a significant proportion of muscle-invasive bladder tumours, and is also present in human bladder cancer cell lines. We also demonstrate that Fra-1, while assumed to be localised to the nucleus, is also present in the cytoplasm where it may have further, as yet undefined, functions.
[Proc Amer Assoc Cancer Res, Volume 46, 2005]