Loss of CXCL12/CXCR4 signalling impacts several aspects of cardiovascular development but does not exacerbate Tbx1 haploinsufficiency

Mahalia Page,Liam Ridge,Peter J Scambler,Diana Gold Diaz,Sarah Ivins,Tsolmon Tsogbayar

doi:10.1371/journal.pone.0207251

Abstract

The CXCL12-CXCR4 pathway has crucial roles in stem cell homing and maintenance, neuronal guidance, cancer progression, inflammation, remote-conditioning, cell migration and development. Recently, work in chick suggested that signalling via CXCR4 in neural crest cells (NCCs) has a role in the 22q11.2 deletion syndrome (22q11.2DS), a disorder where haploinsufficiency of the transcription factor TBX1 is responsible for the major structural defects. We tested this idea in mouse models. Our analysis of genes with altered expression in Tbx1 mutant mouse models showed down-regulation of Cxcl12 in pharyngeal surface ectoderm and rostral mesoderm, both tissues with the potential to signal to migrating NCCs. Conditional mutagenesis of Tbx1 in the pharyngeal surface ectoderm is associated with hypo/aplasia of the 4th pharyngeal arch artery (PAA) and interruption of the aortic arch type B (IAA-B), the cardiovascular defect most typical of 22q11.2DS. We therefore analysed constitutive mouse mutants of the ligand (CXCL12) and receptor (CXCR4) components of the pathway, in addition to ectodermal conditionals of Cxcl12 and NCC conditionals of Cxcr4. However, none of these typical 22q11.2DS features were detected in constitutively or conditionally mutant embryos. Instead, duplicated carotid arteries were observed, a phenotype recapitulated in Tie-2Cre (endothelial) conditional knock outs of Cxcr4. Previous studies have demonstrated genetic interaction between signalling pathways and Tbx1 haploinsufficiency e.g. FGF, WNT, SMAD-dependent. We therefore tested for possible epistasis between Tbx1 and the CXCL12 signalling axis by examining Tbx1 and Cxcl12 double heterozygotes as well as Tbx1/Cxcl12/Cxcr4 triple heterozygotes, but failed to identify any exacerbation of the Tbx1 haploinsufficient arch artery phenotype. We conclude that CXCL12 signalling via NCC/CXCR4 has no major role in the genesis of the Tbx1 loss of function phenotype. Instead, the pathway has a distinct effect on remodelling of head vessels and interventricular septation mediated via CXCL12 signalling from the pharyngeal surface ectoderm and second heart field to endothelial cells.

Highlights

TBX1 haploinsufficiency is the major contributing factor in the development of congenital cardiovascular defects in the 22q11.2 deletion syndrome (22q11.2DS)
The authors of this paper proposed that Cxcl12/Cxcr4 are genetically downstream of Tbx1 during pharyngeal neural crest cells (NCCs) development and that reduction of CXCR4 signalling causes misrouting of pharyngeal NCCs in chick
In addition to previously described expression in the outflow tract, we detected Cxcl12 transcripts in the pharyngeal surface ectoderm of E9.0 embryos i.e. at the time when signalling from the pharyngeal surface ectoderm would be most likely to impact the underlying, migrating NCCs (Fig 1A and 1C)

Summary

Introduction

TBX1 haploinsufficiency is the major contributing factor in the development of congenital cardiovascular defects in the 22q11.2 deletion syndrome (22q11.2DS). Conditional mutagenesis experiments have determined the tissue specific and temporal requirements for this transcription factor in the mouse (reviewed in [1]). Tbx is required in the pharyngeal surface ectoderm for the formation and remodelling of the embryonic pharyngeal arch artery (PAA) system into the great vessels. The defects observed in mice correlate well with abnormalities observed in human patients. Interrupted artic arch type B (IAA-B), which represents a left 4th pharyngeal arch artery (PAA) abnormality is quite specific for 22q11.2 deletion syndrome in that ~50% of patients presenting with this defect will test positive for the 22q11.2 deletion [2]

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Conclusion