(A) TBX5 gene (top) and protein (bottom) with domains labeled. Patient mutation c.254C>G (p.P85R) is indicated by an arrowhead. (B) Anti-HA and wheatgerm agglutinin (WGA) immunofluorescence staining of FaDu cells transfected with HA-TBX5-WT or HA-TBX5-P85R (original magnification, ×40). Ratiometric quantification of nucleus-to-cytoplasm HA signal by box plot (n = 6). P value was determined by Welch’s 2-sample t test. (C) Volcano plot showing the distribution of differentially expressed genes (FDR <0.05 and |log₂fold change| >0.25) comparing HOS and control cardiomyocytes. (D) OR by Fisher’s exact test comparing the overlap of down- and upregulated genes identified in C and in published TBX5-KO iPSC-derived cardiomyocytes (2). (E) OR by Fisher’s exact test comparing the overlap of down- and upregulated genes identified in C and published TBX5 ChIP-Seq from iPSC-derived cardiomyocytes (Supplemental Ref 14). (F) Gene ontology (GO) term analysis of TBX5-dependent genes associated with TBX5 ChIP-Seq not previously reported in cardiomyocyte-derived iPSCs or mouse tissue.