Critical roles of Sertad1 during mouse heart development

Congenital heart diseases (CHDs) occur in as many as 1% of newborns and remain the leading cause of infant morbidity and mortality. Bone Morphogenic Protein (BMP) pathways play crucial roles in many aspects of cardiogenesis, and mutations that disturb BMP signaling cause various forms of congenital heart defects both in animal models and in human patients. BMP receptor-activated Smads (R-Smads), including Smad1, 5 and 8, are nuclear mediators of BMP ligands. Upon activation by the BMP-receptor complex, Smad1/5/8 oligomerize with Smad4 and translocate into the nucleus to regulate expression of target genes. Our long term goal is to reveal the molecular mechanisms whereby BMP-Smad pathways regulate normal cardiogenesis, and to apply this knowledge to the development of novel diagnostic/therapeutic strategies for CHDs. BMP R-Smad activities are critically regulated by their interaction partners. We screened a mouse embryonic heart yeast 2-hybrid library using Smad1 as the bait, and identified Sertad1 as a novel Smad1 interaction partner. Sertad1 encodes a transcriptional co-activator expressed in embryonic hearts. The Smad1-Sertad1 interaction was confirmed with molecular and biochemical studies. Overexpression of Sertad1 enhanced expression of several BMP responsive reporters. Significantly, expression of endogenous Nkx2.5 in embryonic cardiomyocytes was also increased by overexpression of Sertad1. We hypothesize that Sertad1 is a BMP R-Smad co-activator that is critical for mediating cardiogenic activities of BMP signaling. Two specific aims are outlined. In aim 1, we will further characterize the Sertad1-Smad1 interaction with biochemical and molecular approaches to gain fundamental information regarding this novel interaction. We will test whether Sertad1 directly interacts with Smad1, map the domains within Smad1 and Sertad1 responsible for their interaction, and determine whether Sertad1 specifically interacts with Smad1 or can also interact with other R-Smads. In aim 2, we will apply both gain-of-function and loss-of-function strategies to examine roles of Sertad1 during cardiogenesis using P19CL6 stem cells, which can efficiently differentiate into cardiomyocytes upon DMSO stimulation. Successful accomplishment of this project will significantly advance our insights into the molecular mechanisms by which BMP signaling mediates its complex cardiogenic activities. (AHA Program: Grant-in-Aid)