EZH2 is required for parathyroid and thymic development through differentiation of the third pharyngeal pouch endoderm

Abstract

The Ezh2 gene encodes a histone methyltransferase of the Polycomb Repressive Complex 2 that methylates histone H3 lysine 27. In this work we asked whether EZH2 has a role in the development of the pharyngeal apparatus and whether it regulates the expression of the Tbx1 gene, which encodes a key transcription factor required in pharyngeal development. To these ends, we performed genetic in vivo experiments with mouse embryos and we used mouse embryonic stem cell (ESC)-based protocols to probe endoderm and cardiogenic mesoderm differentiation. Results showed that EZH2 occupies the Tbx1 gene locus in mouse embryos, and that suppression of EZH2 was associated with reduced expression of Tbx1 in differentiated mESCs. Conditional deletion of Ezh2 in the Tbx1 expression domain, which includes the pharyngeal endoderm, did not cause cardiac defects but revealed that the gene has an important role in the morphogenesis of the 3rd pharyngeal pouch (PP). We found that in conditionally deleted embryos the 3rd PP was hypoplastic, had reduced expression of Tbx1, lacked the expression of Gcm2, a gene that marks the parathyroid domain, but expressed FoxN1, a gene marking the thymic domain. Consistently, the parathyroids did not develop, and the thymus was hypoplastic. Thus, Ezh2 is required for parathyroid and thymic development, probably through a function in the pouch endoderm. This discovery also provides a novel interpretational key for the finding of Ezh2 activating mutations in hyperparathyroidism and parathyroid cancer.

EZH2 and POU2F3 Can Aid in the Distinction of Thymic Carcinoma from Thymoma

“…Two promising candidates for the immunohistochemical distinction of TETs are the histone methyltransferase Enhancer of Zeste Homolog 2 (EZH2) and the transcription factor POU class 2 homeobox 3 (POU2F3). EZH2 is involved in thymic development [ 10 ]. To our knowledge, only a single study has investigated EZH2 immunoreactivity for distinguishing thymic carcinoma from thymoma [ 4 ].…”

Chromatin regulators in the TBX1 network confer risk for conotruncal heart defects in 22q11.2DS and sporadic congenital heart disease

“…The H3k4me1 mark is for poised enhancers that are further modified to promote gene expression (55). Other proteins we identified with MDRVs are members of the Tbx1 for mouse cardiovascular development (58), supporting a connection to Tbx1. CHD7 is the gene for CHARGE syndrome and functions as an ATP-dependent chromatin remodeler that interacts with the BAF complex (56) as shown in Fig.…”

“…Other proteins we identified with MDRVs are members of the PRC (Polycomb) complex that represses transcription, the BAF complex that alters nucleosomes (SWI/SNF) (56) and NSD1 and NSD2, which are involved in lysine methylation (57). EZH2 is a member of the polycomb complex that methylates H3K27 and Ezh2 genetically interacts with Tbx1 for mouse cardiovascular development (58), supporting a connection to Tbx1. CHD7 is the gene for CHARGE syndrome and functions as an ATP-dependent chromatin remodeler that interacts with the BAF complex (56) as shown in Fig.…”

Fn1Regulates the Third Pharyngeal Pouch Patterning and Morphogenesis

“…Tbx1 , a known transcription factor expressed in the endoderm, distal ectoderm, and mesoderm, is important for the thymus/parathyroid fate specification and organogenesis (Xu et al 2005; Reeh et al 2014; Hasten and Morrow 2019; Caprio et al 2021). On the third pp, Tbx1 is restricted to the parathyroid domain and is coexpressed with Gcm2 and complementary to Foxn1 (Manley et al 2004; Liu et al 2007; Bain et al 2016; Caprio et al 2021). Ectopic Tbx1 in the thymus domain can repress Foxn1 expression (Reeh et al 2014; Bain et al 2016).…”

“…Splotch ( Pax3 null) mutant mouse embryos with severe NCC deficiency have also been shown to have variable defective thymus organogenesis (Conway et al 1997; Griffith et al 2009). Proper epithelial-mesenchymal interactions are also essential for thymus/parathyroid morphogenesis, which involves genes such as TBX1 (Hasten and Morrow 2019; Caprio et al 2021), BMP4 (Tsai et al 2003; Patel et al 2006; Gordon et al 2010; Neves et al 2012; Swann et al 2017), FGF7/10 (Revest et al 2001), HOXA3 (Su et al 2001; Kameda et al 2004; Chojnowski et al 2014; Chojnowski et al 2016; Gordon 2018), and SHH (Grevellec et al 2011; Bain et al 2016; Peissig et al 2018), among others. The BMP4 and SHH genes were proposed to function opposingly in patterning the third pp, whereas in the Shh null mutant, Bmp4 expression is expanded throughout the third endoderm (Moore-Scott and Manley 2005; Gordon and Manley 2011).…”