Human embryonic stem cells as a model for cardiac gene discovery : from chip to chap

Abstract

Here we described the use of human embryonic stem cells (hESCs) as a model to obtain insights into commitment to the mesoderm and endoderm lineages and the early steps in human cardiac cell differentiation by means of whole-genome temporal expression profiling. Furthermore, we used it as an approach to identify new genes that may be associated with (abnormal) human heart development. This was the first detailed temporal transcriptional profiling of hESCs differentiating towards a specific cell type to be reported, most likely due to the lack of efficient protocols for directed differentiation. It is important to profile the expression changes that occur before and during cell commitment and to identify the key genes involved during the critical time points of the differentiation process. Furthermore it allowed us to identify novel genes that are enriched in cardiomyocytes. One of several novel cardiac-enriched genes that were identified in the microarray analysis was cloned and studied further. We mainly focused on unravelling the function of this gene. We showed that this gene encodes a cytoskeletal protein that is actin-associated and binds to ?-actinin2 at the Z-disc of sarcomeres. Hence, we named the gene Cytoskeletal Heart-enriched Actin-associated Protein (CHAP). Furthermore, we identified and knocked down the CHAP orthologues in zebrafish which lead to impaired heart and skeletal muscle development and function, indicating an important role for chap in muscle. Since heart failure is the number one cause of mortality and morbidity worldwide, the identification of new components, the precise molecular mechanisms of the Z-disc and its role in signaling has become critical for understanding the regulation of cardiac function and disease. Furthermore, we describe the identification of another novel heart-enriched gene which was annotated as steroid 5?-reductase 2-like 2 (SRD5A2L2). We investigated the expression of this gene during embryonic development and provide the first indication that steroid metabolism might be involved in muscle development. Finally, we show that Asb2, a member of the Ankyrin-repeat and SOCS-box containing family which we also identified in our previous microarray study, is expressed during early mouse heart and skeletal muscle development. Our findings represent novel disease candidate genes that could be involved in (cardio) myopathies. Further studies on the function of CHAP, Srd5a4 and Asb-2 will shed more light on their possible role in disease. Since the molecular mechanisms of cardiogenesis are conserved in zebrafish, mouse and human, we could use the zebrafish as a model system to investigate the importance of novel genes such as Srd5a4 and Asb-2 in cardiac development. In addition, making use of DNA-databases of congenital heart disease patients such as the ”CONgenital COR vitia” registry (CONCOR), which contains DNA of over 4000 dutch patients, will allow us to screen for mutations in our discovered candidate genes