Two translation initiation codons direct the expression of annexin VI 64kDa and 68kDa isoforms

Annexin A6 is a multicompetent, multifunctional protein involved in several biological processes within and outside of the cell. Whereas HeLa cells express annexin A6 only as a 68/67-kDa doublet, indicating alternative splicing (Smith PD et al. (1994) Proc Natl Acad Sci USA 91, 2713-2717), the GMO2784 human fibroblast cell line expresses two additional isoforms at 64 and 58kDa. In both cell lines, annexin A6 is located intracellularly and on the plasma membrane. In vitro eukaryotic protein synthesis of pIRESneoAnxA6 cDNA and pIRESneoAnxA6/Met^1- or Met^33- using a reticulocyte lysate coupled transcription/translation system revealed that this gene contains two translation start codons, Met¹ and Met³³. Immunoprecipitation of the products obtained from the transcription/translation system using various anti-annexin A6 antibodies confirmed the presence of several isoforms and suggested that this protein might be present in different configurations.