Subcellular localization of the oncoprotein MTG8 (CDR/ETO) in neural cells

Abstract

The t(8;21) translocation associated with acute myeloid leukemia (AML) disrupts two genes, the AML1 gene also known as the core binding factor A2 (CBFA2) on chromosome 21, and a gene on chromosome 8, hereafter referred to as MTG8, but also known as CDR and ETO. Extensive information is available on AML1, a member of the CBF family of transcription factors, containing a highly conserved domain, the runt box, of the Drosophila segmentation gene runt. This gene is essential for the hematopoietic development and is found disrupted in several leukemias. In contrast, the function of the MTG8 gene is poorly understood. The predicted protein sequence shows two unusual, putative zinc-fingers, three proline-rich regions, a PEST domain and several phosphorylation sites. In addition, we found a region encompassing aa 443–514 predicted to have a significant propensity to form coiled coil structures. MTG8 displays a high degree of similarity with nervy, a homeotic target gene of Drosophila, expressed in the nervous system. Human and mouse wild-type MTG8 are also highly expressed in brain relative to other tissues. For these reasons, we set out to investigate the expression and subcellular localization of the MTG8 protein in neural cells. Immunohistochemical experiments in a 12.5-day-old mouse embryo clearly showed that the protein was expressed in the neural cells of the developing brain and the spinal cord. In primary cultures of hippocampal neurons of 2–3 day-old mice, MTG8 was found in the nucleus, in the cytoplasm and as fine granules in the neurites. Cytoplasmic localization of the protein was observed in Purkinje cells of both human and mouse cerebellum. The molecular mass of MTG8 in total human and mouse brain was analysed by immunoblotting and determined to be between 70 and 90 kDa. Isoforms with the same molecular mass were demonstrated in synaptosomes isolated from mouse forebrain. The evidence of MTG8 in the nucleus and cytoplasm of neural cells suggests a specific mechanism regulating the subcellular localization of the protein.