Abstract Body: Haploinsufficiency of the RNA splicing regulator, RBFOX2, has been linked to congenital heart disease (CHD), including hypoplastic left heart syndrome (HLHS), yet the underlying pathogenic mechanisms remain unclear. Here, we generated human induced pluripotent stem cells (iPSCs) that were either heterozygous or null for RBFOX2 and differentiated them into cardiomyocytes (iPSC-CMs). Heterozygous iPSC-CMs exhibited intermediate defects in cell size, adhesion, sarcomere organization, respiration, and calcium handling relative to null cells, consistent with stalled differentiation. Engineered heart tissues (EHTs) also showed dose-dependent reductions in systolic force. At the molecular level, we found that RBFOX2 promotes CM differentiation by driving transcriptome maturation, shifting exon usage profiles to more mature states in genes encoding sarcomere, cytoskeleton, and focal adhesion components, including the critical Z-disc factor alpha-actinin 2 (ACTN2). This transition is initiated when RBFOX2 levels reach a critical threshold that enables auto-splicing of mutually exclusive exons encoding early and late RBFOX2 isoforms. Specifically, RBFOX2 binds its own transcript just upstream of the early/fetal exon to repress its inclusion during transcriptome maturation. In heterozygous CMs, this threshold is never reached, resulting in retention of the early exon. This, in turn, causes RBFOX2 isoform imbalance and leads to co-inclusion of both exons, generating an aberrant transcript containing a premature stop codon. Consequently, functional RBFOX2 transcripts are reduced to well below 50% of WT. Rescue experiments indicate that the early RBFOX2 isoform supports cellular differentiation and transcriptome maturation, whereas the late isoform promotes functional maturation. Finally, we demonstrate that ACTN2 overexpression rescues heterozygous, but not null, phenotypes by restoring contractility, which activates a mechanosensitive feedback loop involving upregulation of RBFOX2 from the wild-type allele that matures the transcriptome. Together, these findings identify RBFOX2 dosage-sensitive autoregulatory splicing as a key regulator of cardiomyocyte differentiation and suggest that myocardial-intrinsic defects contribute to RBFOX2-mediated CHD pathogenesis and heart failure susceptibility.