Inhibition of Transcription Factor GATA-4 Expression Blocks In Vitro Cardiac Muscle Differentiation

Abstract

Commitment of mesodermal cells to the cardiac lineage is a very early event that occurs during gastrulation, and differentiation of cardiac muscle cells begins in the presomite stage prior to formation of the beating heart tube. However, the molecular events, including gene products that are required for differentiation of cardiac muscle cells, remain essentially unknown. GATA-4 is a recently characterized cardiac muscle-restricted transcription factor whose properties suggest an important regulatory role in heart development. We tested the role of GATA-4 in cardiac differentiation, using the pluripotent P19 embryonal carcinoma cells, which can be differentiated into beating cardiac muscle cells. In this system, GATA-4 transcripts and protein are restricted to cells committed to the cardiac lineage, and induction of GATA-4 precedes expression of cardiac marker genes and appearance of beating cells. Inhibition of GATA-4 expression by antisense transcripts blocks development of beating cardiac muscle cells and interferes with expression of cardiac muscle markers. These data indicate that GATA-4 is necessary for development of cardiac muscle cells and identify for the first time a tissue-specific transcription factor that may be crucial for early steps of mammalian cardiogenesis.

Differentiation-dependent mechanisms of transcriptional regulation of the catalytic subunit of phosphorylase kinase

“…Of possibly special significance are the MEF-2 site (bp k215-207) and the heart progenitor cell-specific transcription-factor GATA-4 site (bp k563 to k555). These have been shown to be important for skeletal-and cardiac-muscle development [26,27]. The MEF-2 motif has been shown to regulate the cardiac troponin C gene [43] as well as the myogenin gene itself [44].…”

“…The third of these is within the primary PhK-γ promoter sequence between the CAAT and TATA boxes that are located at k144 to k140 and k32 to k26 bp respectively. Multiple other cis-regulatory sites that are apparent from the PhK-γ sequence (Figure 1, lower panel), include an MEF-2 site at k213 and three GATA-4 sites at k1027, k895 and k551, which in other systems have been shown to be important for skeletal-and cardiac-muscle development [26,27].…”

“…Given that p(k116\j357)Luc, even in the absence of the CAAT box, also supported modest, but differentiation-dependent, transcription, it is likely that the Ep E-box (bp k114 to k109) is providing the differentiation specificity, with general transcription enhanced by one or more positive elements between bp k115 and k484. Possible candidates for this include the MEF (bp k215) and GATA-4 (bp k551) sites, which have been shown in other systems to be important for skeletaland cardiac-muscle development [26,27]. Addition of sequences further upstream of bp k484 led to a potent down-regulation of the transcriptional activity localized to the region spanning k744 to k983.…”

Regulation of the rat cardiac troponin I gene by the transcription factor GATA-4

“…Two recent studies have explored the role of GATA-4 in early cardiocyte differentiation. Grepin et al [48] have suggested that GATA-4 plays a role in cardiac cell commitment, as evidenced by the lack of cardiocyte differentiation in P19 cells transfected with antisense GATA-4 constructs ; however, data from chimaeric mice created by blastocyst injection with embryonic stem cells with GATA-4 deleted indicates that these cells have the capacity to contribute to the myocardium [49]. By analogy with the roles of GATA and other transcription factors in haematopoietic lineages [50], GATA-4 and other GATA proteins and cofactors in the heart may play a role in both early cardiocyte differentiation, as well as later stages of developmental regulation.…”

In vitro organogenesis from undifferentiated cells in Xenopus

“…Thus, screening with these in vitro-induced hearts has contributed new knowledge as to when and how the components of the hyaluronan matrix function in cardiogenesis. These systems also permit more detailed analyses of the roles of Wnt signaling (Marvin et al, 2001;Schneider and Mercola, 2001;Tzahor and Lassar, 2001) and transcription factors, such as Nkx2.5 (Durocher et al, 1997;Reecy et al, 1999) and GATA4 (Grepin et al, 1995(Grepin et al, , 1997Jiang et al, 1999) in cardiogenesis. In the recent study, we examined a comprehensive set of cardiogenesis-related genes using an in vitro Xenopus-based system and microarrays.…”