Gradient of Integrin α6A Distribution in the Myocardium During Early Heart Development

The interactions of cells with extracellular matrices (ECM)1 are likely to be key determinants of embryonic development. Integrin adhesion receptors are ideally positioned to mediate some of these interactions since, in addition to mechanical adhesion, they transduce signals affecting cell proliferation and differentiation. We investigated expression of the integrin αβ1, a receptor for the ECM component, laminin in the early mouse embryo. An intriguing feature of this integrin is the existence of α6 subunit isoforms. The A and B isoforms, which differ in the cytoplasmic tails., are expressed in cell-type specific fashion, and are likely to implement distinct cellular interactions with laminin. By RT-PCR, α6B but not α6A mRNA was detectable in embryo extracts from fertilized oocytes to 6.5 d.p.c. In subsequent stages, up to 11.5 d.p.c., α6A mRNA was observed in mRNA extracts from whole embryos, but still in significantly lower amounts than α6B. However, in extracts from isolated heart (9.5 to 11.5 d.p.c.), α6A was the predominant α6 isoform, while in extracts from other embryo parts no α6A mRNA was detectable. At the protein level, immunostaining with specific antibodies showed α6A protein in myocardial cells, at the early stage of heart tube development (8.5 d.p.c). Localization to the myocardium was tightly restricted, since other structures of the embryonic heart, e.g., endocardium, or of the remaining embryo did not stain with anti-α6A antibody. In the ventricular myocardium, expression of α6A appeared more intense than in the subendocardial layer. Quantitation by confocal microscopy unveiled a gradient of expression of α6A, increasing from the outer to the inner layers of the myocardium. This is the first demonstration of a gradient distribution of integrin molecules in a tissue, which appears to be directly connected with the process of organogenesis. The mechanism underlying our observations is not the turning on of a gene, rather it is the activation of a splicing mechanism that substitutes the cytoplasmic domain of a laminin receptor. Because integrin cytoplasmic domains are thought to be an important functional end of the molecule, this may be a mechanism to modulate cellular responses to laminin.