Three distinct mechanisms, Notch instructive, permissive, and independent, regulate the expression of two different pericardial genes to specify cardiac cell subtypes

Abstract

The development of a complex organ involves the specification and differentiation of diverse cell types constituting that organ. Two major cell subtypes, contractile cardial cells (CCs) and nephrocytic pericardial cells (PCs), comprise the Drosophila heart. Binding sites for Suppressor of Hairless [Su(H)], an integral transcription factor in the Notch signaling pathway, are enriched in the enhancers of PC-specific genes. Here we show three distinct mechanisms regulating the expression of two different PC-specific genes, Holes in muscle ( Him ), and Zn finger homeodomain 1 ( zfh1 ). Him transcription is activated in PCs in a permissive manner by Notch signaling: in the absence of Notch signaling, Su(H) forms a repressor complex with co-repressors and binds to the Him enhancer, repressing its transcription; upon alleviation of this repression by Notch signaling, Him transcription is activated. In contrast, zfh1 is transcribed by a Notch-instructive mechanism in most PCs, where mere alleviation of repression by preventing the binding of Su(H)-co-repressor complex is not sufficient to activate transcription. Our results suggest that upon activation of Notch signaling, the Notch intracellular domain associates with Su(H) to form an activator complex that binds to the zfh1 enhancer, and that this activator complex is necessary for bringing about zfh1 transcription in these PCs. Finally, a third, Notch-independent mechanism activates zfh1 transcription in the remaining, even skipped -expressing, PCs. Collectively, our data show how the same feature, enrichment of Su(H) binding sites in PC-specific gene enhancers, is utilized by two very distinct mechanisms, one permissive, the other instructive, to contribute to the same overall goal: the specification and differentiation of a cardiac cell subtype by activation of the pericardial gene program. Furthermore, our results demonstrate that the zfh1 enhancer drives expression in two different domains using distinct Notch-instructive and Notch-independent mechanisms.

Single-cell sequencing of theDrosophilaembryonic heart and muscle cells during differentiation and maturation

“…The segmental Drosophila embryonic heart consists of 104 cardioblasts (CBs), distributed over eight segments (T3, A1-A7), as well as several types of pericardial cells (Panta et al, 2020;Reim and Frasch, 2009;Rotstein and Paululat, 2016). Post-specification the CBs express a common set of transcription factors (TFs), including Tail-up, H15 and Mef2 (Gajewski et al, 1997;Griffin et al, 2000;Mann et al, 2009).…”

“…The formation of the heart is a finely controlled process of specification and differentiation events thought to be under tight control of cardiac transcription factors and signaling pathways, in vertebrates as well as in the fruit fly (Ahmad, 2017; Bodmer and Frasch, 2010; Cripps and Olson, 2002). The segmental Drosophila embryonic heart consists of 104 cardioblasts (CBs), distributed over eight segments (T3, A1-A7), as well as several types of pericardial cells (Panta et al, 2020; Reim and Frasch, 2009; Rotstein and Paululat, 2016). Post-specification the CBs express a common set of transcription factors (TFs), including Tail-up, H15 and Mef2 (Gajewski et al, 1997; Griffin et al, 2000; Mann et al, 2009).…”