Abstract P1073: The Mir-200 Family Modulates Expression Of Tbx5 , Gata 4, And Pitx2 During Early Cardiac Development.

Abstract

Congenital heart defects (CHDs) may be caused by mutations in genes that drive cardiac development. Cardiac development is also regulated at the transcription level. MicroRNAs (miRs) target the transcripts of genes in numerous cardiac cell types during embryonic development. We and others have shown that the miR-200 family modulates the transcripts of cardiogenic transcription factors (TFs) Tbx5 , Gata4 , and Pitx2 ( TGP ). However, relationship between these miRs and cardiogenic TFs during cardiac development is poorly understood. To this relationship, we cultured cardiomyocytes (CMs) with reduced miR-200 expression and found increased expression of Tbx5, Gata4, and Pitx2, providing evidence for targeting in cell-autonomous role. In vivo, the miR-200 family members, miR-200a and -200c , are highly expressed in the developing heart (e14.5) and reduced in adults (2mo) (Ct Value- 200a: 24.3 ± 0.59 v 38.3 ± 0.21; 200c: 25.0 ± 0.64 v 32.5 ± 0.16). Using our miR-200 family inhibitor mice models ( I-200 ), we have found these miRs are required for cardiac development. I-200 embryos are found with a membranous ventral septal defect and poor ventricle wall development, which is lethal by e16.5. At e14.5, I-200 hearts have a significant increase in expression fold change of Tbx5 (5.98 ± 1.51 p ≤ 0.05), Gata4 (3.29 ± 0.61 p ≤ 0.05), and Pitx2 (4.67 ± 0.23 p ≤ 0.05) compared to Wild-Type. With further investigation we discovered the miR-200 family is required for CMs differentiation. I-200 CMs have increased expression of cardiac progenitor markers Nkx2.5 and Mef2c compared to Wild-Type. As these cardiogenic transcription factors target DNA elements required for the maintenance of the CM transcriptome and identity, our data suggests the miR-200 family acts in this process. Concurrent single nuclei multiomics sequencing revealed changes in CM transcription and open chromatin to provided corresponding evidence to support this conclusion. Conclusions: The miR-200 family is a modulator of cardiogenic TFs expression and activity during development. Future directions will determine the role of miR- 200 in adult cardiac disease, such as ischemic injury. Our work provides new insights into gene dosage, modulated by miRs, that is required and necessary during cardiac development.