Different Ectopic Hoxa2 Expression Levels in Mouse Cranial Neural Crest Cells Result in Distinct Craniofacial Anomalies and Homeotic Phenotypes.

Taro Kitazawa,Sebastien Ducret,Maryline Minoux,Filippo M Rijli

doi:10.3390/jdb10010009

Abstract

Providing appropriate positional identity and patterning information to distinct rostrocaudal subpopulations of cranial neural crest cells (CNCCs) is central to vertebrate craniofacial morphogenesis. Hox genes are not expressed in frontonasal and first pharyngeal arch (PA1) CNCCs, whereas a single Hox gene, Hoxa2, is necessary to provide patterning information to second pharyngeal arch (PA2) CNCCs. In frog, chick and mouse embryos, ectopic expression of Hoxa2 in Hox-negative CNCCs induced hypoplastic phenotypes of CNCC derivatives of variable severity, associated or not with homeotic transformation of a subset of PA1 structures into a PA2-like identity. Whether these different morphological outcomes are directly related to distinct Hoxa2 overexpression levels is unknown. To address this issue, we selectively induced Hoxa2 overexpression in mouse CNCCs, using a panel of mouse lines expressing different Hoxa2 ectopic expression levels, including a newly generated Hoxa2 knocked-in mouse line. While ectopic Hoxa2 expression at only 60% of its physiological levels was sufficient for pinna duplication, ectopic Hoxa2 expression at 100% of its normal level was required for complete homeotic repatterning of a subset of PA1 skeletal elements into a duplicated set of PA2-like elements. On the other hand, ectopic Hoxa2 overexpression at non-physiological levels (200% of normal levels) led to an almost complete loss of craniofacial skeletal structures. Moreover, ectopic Hoxa5 overexpression in CNCCs, while also resulting in severe craniofacial defects, did not induce homeotic changes of PA1-derived CNCCs, indicating Hoxa2 specificity in repatterning a subset of Hox-negative CNCCs. These results reconcile some discrepancies in previously published experiments and indicate that distinct subpopulations of CNCCs are differentially sensitive to ectopic levels of Hox expression.

Highlights

The multipotent neural crest cells delaminate dorsally along the rostrocaudal axis of the forming neural tube and migrate towards various regions of the developing embryo, where they locally differentiate into a broad range of cell lineages [1]
We found that overexpression of Hoxa5 in cranial neural crest cells (CNCCs) induced a loss of proximal PA1 and PA2–PA4 structures without homeotic transformation, indicating specificity of Hoxa2 in the repatterning of a subset of PA1 into PA2-like CNCC derivatives
Wnt1-Hoxa2-medium fetuses displayed a duplication of the pinna, even though both normal and ectopic pinnae were mildly hypoplastic as compared to wild-type pinna

Summary

Introduction

The multipotent neural crest cells delaminate dorsally along the rostrocaudal axis of the forming neural tube and migrate towards various regions of the developing embryo, where they locally differentiate into a broad range of cell lineages [1]. Cranial neural crest cells (CNCCs) colonize the frontonasal process (FNP) and pharyngeal arches (PAs), providing the main source of skeletogenic head mesenchyme, central to craniofacial and pharyngeal morphogenesis. More posterior CNCCs fill the PAs, forming the cartilages and bones of the upper and lower jaws, middle ear ossicles, outer ear, hyoid and thyroid structures [1,2,3,4]. Defects of regional specification and patterning of different CNCC subpopulations lead to distinct craniofacial abnormalities [5]. CNCCs contributing to the FNP and first pharyngeal arch (PA1) do not express Hox genes, whereas CNCCs contributing to the second and more posterior arches display nested combinatorial Hox expression patterns, providing each PA with distinct regional identities along the rostrocaudal axis [3,4,6,7,8]. In Hoxa knockout mice, the PA2-derived stapes, styloid process and lesser horns of hyoid bone were absent and replaced by proximal PA1-like structures including the malleus, incus, gonial bone and tympanic ring in a mirror image pattern of their normal counterparts [9,10]

Methods

Results

Conclusion