Abstract Body: Disrupting the tightly regulated process of cardiac development can lead to congenital heart diseases, yet the role of posttranslational mechanisms remains largely overlooked. Here, we uncover a novel role for Cullin-RING ubiquitin E3 ligases (CRLs) in perinatal cardiac development. In CRLs, RBX1 functions as the catalytic subunit, partnering with Cullin 1-4 proteins and various substrate-recognition receptors to mediate ubiquitination and degradation of intracellular proteins. RBX1 is highly expressed in fetal hearts but downregulated in adults. Surprisingly, deletion of a single Rbx1 allele in mouse hearts via Xmlc2^Cre, which is active from embryonic day (E) 7.5, was sufficient to cause heart failure and perinatal lethality by postnatal day 5. Heterozygous knockout mice exhibited myocardial hypoplasia in both the trabecular and compact layers, discernible at E13.5 and developed ventricular septal defects by E18.5. Similarly, deletion of one Rbx1 allele via cTnT^Cre, which is highly active between E7.5 and E10.5, caused postnatal cardiac dysfunction. In contrast, mice with heterozygous Rbx1 deletion via αMHC^Cre, which is highly active in cardiomyocytes during midgestation, were viable and exhibited no cardiac abnormalities. However, homozygous deletion via αMHC^Cre led to myocardial hypoplasia and biventricular non-compaction by E14.5, ultimately causing embryonic heart failure and lethality by E18.5. These data suggest that precise temporal regulation of RBX1 levels is essential for cardiac morphogenesis. In all three mutant hearts, the observed cardiac developmental abnormalities are attributed to deficits in cardiomyocyte proliferation and maturation. Transcriptomic analysis revealed disruptions in MEF2 transcriptional networks, which are essential for cardiogenesis, among others in RBX1-deficient hearts. Mechanistically, RBX1 deficiency inactivated CRLs, leading to the accumulation of Hippo kinases (LATS1/2, MST1/2, SAV, and MOB1), which inhibited YAP transactivation and proliferation gene expression. We further demonstrate that RBX1 promotes MOB1 ubiquitination and degradation. These findings highlight that RBX1 acts as a dosage-sensitive regulator of early cardiogenesis, ensuring proper cardiomyocyte proliferation, maturation, and ventricular chamber formation through modulation of the Hippo-YAP pathway.