Transcriptional events during initial vertebrate heart development in vivo remain poorly understood. Mesp1, a bHLH transcription factor, has been described as the earliest transcriptional regulator of cardiac progenitors in multiple species, and represents an excellent candidate for the investigation of relevant targets during cardiovascular development. We report here that both depletion and mutation of Mespaa, the zebrafish homolog of mammalian Mesp1, lead to randomization of cardiac looping, together with significant cardiac morphogenesis defects. These disruptions are preceded by a defect in cardiac left-right asymmetry. Surprisingly, the defect in asymmetry was found to occur independently of defects in the transient organ of laterality, the Kupfferâ€™s vesicle (KV). We show that Mespaa regulates miR-430 expression during gastrulation to control the levels of Nodal signaling, and that this regulation is required for asymmetric laterality signaling in the prospective heart field. Ectopic expression of miR-430 is sufficient to induce cardiac laterality defects, and consistent with Mespaa over-expression in this system, the reduction of miR-430 leads to cardia bifida. This study reveals a novel transcriptional regulation of miR-430 by Mespaa and a role for this pathway in cardiac laterality during gastrulation.