Functional analyses of the conserved Cysteine-rich with EGF-like domains (Creld) protein family in <em>Mus musculus</em>

Abstract

The Cysteine-rich with EGF-like domains (Creld) protein family is highly conserved between species. However, its physiological function remained elusive. Using Creld1 knock-out (<em>Creld1KO</em>) mice, Creld1 was identified as an important regulator of the calcineurin/nuclear factor of activated - cells c1 (NFATc1) signaling pathway during heart valve formation. <em>Creld1KO</em> embryos die at embryonic day E11.5 due to cardiac dysfunction. At E10.5, <em>Creld1KO</em> embryos display defects in the formation of the atrioventricular cushion, the precursor of the heart valve. Heart-valve formation crucially relies on the calcineurin/NFATc1 cascade. The results showed that in the <em>Creld1KO</em> endocardium, from where the heart valves originate, nuclear translocation of NFATc1 is impaired. This results in a decrease of NFATc1 target-gene expression thereby, proliferation within the atrioventricular cushions is impaired. We could demonstrate that Creld1 directly interacts with calcineurin B, the regulatory subunit of calcineurin, thus controlling NFATc1 translocation to the nucleus. In a heterologous system, expression of Creld1 is sufficient to endorse NFATc1 translocation to the nucleus. Sequential deletion of the different functional domains or the introduction of various point mutations indicate that the conserved WE domain of Creld1 is important for regulating the calcineurin phosphatase activity.