Post-transcriptional Regulation Of Nkx2-5 In Heart By RHAU

Congenital heart disease is the leading cause of infant death all over the world. Many congenital heart defects result from spontaneous genetic mutations. Numerous mutations in the core cardiac transcription factors have been identified in patients with some of the most common forms of congenital heart defects. Heart is the first organ to form during development of the body; the development of the heart is tightly controlled by a group of core cardiac transcription factors. Nkx2-5 is one of the core cardiac transcriptional factors that are expressed as early as heart progenitor cells specification in both the primary and secondary heart field. Nkx2-5 is highly expressed in embryonic and neonatal heart, but at a relatively low level in adult mice. The transcriptional regulation of Nkx2-5 by both transcription factors and cis-regulatory elements is reported in many studies, but the post-transcriptional regulation on Nkx2-5 mRNA, such as translational control and mRNA decay, has not been reported so far. Through examining the expression of Nkx2-5 in RNA and protein level, we found that the RNA and protein level do not match. This indicates post-transcriptional regulation plays an important role in Nkx2-5 expression. Through a serial analysis, we found that the 5’UTR of Nkx2-5 mRNA particular the middle 70nt harbors a great translational inhibitory function. We found RNA helicase, RHAU, physically interacts with Nkx2-5 mRNA and promotes the Nkx2-5 mRNA translation and decay. Deleting RHAU in cardiomyocytes results in the accumulation of Nkx2-5 mRNA in the cytoplasm; however, Nkx2-5 protein levels are even lower due to the low translation efficiency of Nkx2-5 mRNA without RHAU. The changed Nkx2-5 mRNA levels can be restored after reintroducing RHAU into the RHAU deficient cardiomyocytes.Nkx2-5 is critical for normal heart development and is also needed for the survival of mature cardiomyocytes in the adult heart. The post-transcriptional regulation of Nkx2-5 by RHAU is expected to have great impact both on heart development and adult heart function. To explore the in vivo function of this regulation, we generated cardiac specific RHAU deficient mice by mating RHAU flox mice with Mespl-cre which is expressed in cardiogenic mesoderm at a later gastrulation stage. Targeted deletion of RHAU in embryonic heart resulted in severe heart hypoplasia and embryonic lethality. Downstream target genes of Nkx2-5 including βMHC, MLC2V and connexin45, were dysregulation in RHAU knockout mice. We deleted RHAU in the postnatal heart using αMHC-cre which is mainly expressed in postnatal cardiomyocyte to study the role of RHAU in postnatal heart. Loss of RHAU in the postnatal heart increased cell apoptosis and resulted in progressive heart failure.In conclusion, our findings revealed the importance of the post-transcriptional regulation on Nkx2-5 gene expression. RHAU as a critical post-transcriptional regulator of Nkx2-5, is essential for heart development and function. Studies on the post-transcriptional regulation of other core cardiac gene can help us better understand the normal heart formation and congenital heart disease.