Expression, Regulation and Function of Myoglobin in Breast Cancer
G Kristiansen, J Hu, D Stiehl, M Rose, J Gerhardt, FR Fritzsche, H Moch, JP Theurillat, E Gnaiger, T Hankeln, TA Gorr, E Dahl. University of Zurich, Zurich, Switzerland; University Hospital of the RWTH Aachen, Aachen, Germany; Medical University of Innsbruck, Innsbruck, Austria; Johannes Gutenberg University Mainz, Mainz, Germany
Background: Myoglobin is assumed to be among the best characterized proteins in humans and is widely believed to be restricted to muscle tissues. Non-muscle myoglobin expression has hardly been noticed by a wider public. Having accidentally observed myoglobin expression in solid tumors, we aimed to analyse expression, regulation and function of myoglobin (Mb) in breast cancer.
Design: A wide spectrum of morphological and biological techniques has been used in this study including immunohistochemistry/immunofluorescence, microdissection, transmission electron microscopy, Western blot, qPCR, cell culture (under normoxia and hypoxia), shRNA and siRNA knockdown, in silico data mining, high resolution respirometry, migration assays.
Results: Mb protein is expressed in 71% of invasive breast carcinomas (n=917) in association with the hypoxia inducible factor 2a (HIF-2 a), carbonic anhydrase IX and a significantly better prognosis. This expression of Mb in breast cancer was confirmed on mRNA level and occurred irrespective of rhabdomyoid differentiation and was inducible by prolonged hypoxia in breast cancer cell lines (MDA-MB231, MDA-MB468, MCF7) with a median change fold of 3.4 after 72 hours. This induction, mediated by HIF-1- and HIF-2-, originated not from the gene's constitutive promoter active in striated muscle but from an alternative start site proximal to a 5' utr exon that is located upstream the initial ATG codon of the commonly translated reading frame. Functionally, stable myoglobin knockdown in MDA-MB468 cells was associated with a stimulated O2 uptake during mild hypoxia, yet did not impact the O2 diffusion gradient in these small cells during limiting conditions. Knockdown of Mb also conferred a significant retardation of the cells' in vitro motility to close an inflicted wound within the normoxic culture.
Conclusions: Mb is commonly expressed in breast cancer at lower levels and irrespective of a metaplastic phenotype. Although generally inducible by hypoxia other functions of myoglobin, not directly related to the transport of oxygen, need to be analysed in further studies.
Monday, March 22, 2010 1:00 PM
Poster Session II # 65, Monday Afternoon