Orteguaza

3.2.1 Abstract

Heterogeneous mutations in the fibroblast growth factor receptor 2 gene (FGFR2) cause a range o f craniosynostosis syndromes. The specificity o f the Apert cranial phenotype reflects its narrow mutational range, in that 98% o f Apert cases result from the Ser252Trp or the Pro253Arg mutations in the FGFR2 — Ig llla extracellular subdomain. In contrast, a broad range o f mutations throughout the extracellular domain o f FG FR2 causes the overlapping cranial phenotypes o f Pfeiffer, Crouzon, and related craniofacial dysostoses.

The expression o f F G F R l, the Igllla/c and Igllla/b isoforms o f FGFR2, and FGFR3 is investigated in Apert (P253R) and Pfeiffer (C278F) foetal cranial tissue and contrasted with normal human controls. FGFR1 and FGFRJ are normally expressed at 14 — weeks in the cells o f the periosteum and osteoid, in domains overlapped by that o f FGFR2, which widely includes pre — osseous cranial mesenchyme. FGFR2 expression is, however, restricted to domains o f advanced osseous differentiation in both Apert and Pfeiffer cranial

skeletogenesis, in the presence o f FG F2 but n ot FG F4 or FGF7. Whereas expression o f the

FGFR2-IgIIIa/b (KGFR) isoform is restricted in normal human cranial osteogenesis, there is preliminary evidence that KGFR is ectopicaHy expressed in Pfeiffer cranial osteogenesis.

Contraction o f the FGFR2-IgIIIa/c (BEK) expression domain in cases o f A pert and Pfeiffer foetal cranial ossification suggests that the m utant activation o f this receptor, by ligand - dependent or ligand - independent means, results in negative - autoregulation. This

phenom enon, resulting from different mechanisms in the two syndromes, offers a model by which to explain differences in their cranial phenotypes.

3.2.2 Introduction

A range o f m utations in three hom ologous genes encoding the fibroblast grow th factor recep to r (FG FR ) proteins 1-3 cause related syndrom ic craniofacial dysostoses. T hese syndrom es, w hich variably feature craniosynostosis, facial dysm orphism , and extracranial m anifestations, were initially identified as frequent phenotypic clusters. A p e rt syndrom e (Apert, 1906) features a narrow range o f craniofacial dysm orphism including: pterional in drawing, a m arkedly foreshortened skull base, turribrachycep haly, severe m idfacial retrusion w ith hypertelorism ; and coronal sutural synostosis, w ith a widely unossified m edian sagittal diastem a in place o f m etopic and sagittal sutures (K reiborg and C ohen, Jr., 1990;K reiborg et al, 1993;Cohen and K reiborg, 1994). T h e characteristic A p ert cranial p h en otype reflects a narrow m utational range w ithin the F G F R 2 gene. T w o neighbouring ‘linker region’ m utations in the I g llla extracellular subdom ain o f F G F R 2 cause 98% A pert cases (Wilkie et al, 1995a) as a result o f Hgand - dependent ‘gain o f function’ (A nderson et al, 1998c).

By contrast, the related syndrom es o f C rouzon, Pfeiffer, and Jackson W eiss show a m ore variable and overlapping craniofacial dysm orphism . T he basicranial sutures generally fuse earlier than in A pert syndrom e, and the m etopic and sagittal sutures form and fuse w ithout the unossified m edian diastem a (K reiborg et al, 1993;CinaUi et al, 1995). F urth erm o re, facial retrusion may range from negligible to severe; and may variably effect the supraorbital or m idfacial skeleton, w ith m inimal pterional in drawing. T he phenotypic variability w ithin and betw een these syndrom es reflects their m utational base. A wide range o f m utations

encom passing the extracellular and transm em brane dom ains o f F G F R 2 causes the ‘C rouzon — P feiffer’ group o f syndrom es, w ith greatest frequency in the I g lllc subdom ain o f the F G F R 2 -Ig IIIa /c (BEK) splice variant (Burke et al, 1998). M any o f these m utations result in the creation o r rem oval o f an unpaired cysteine residue, o r affect a neighbouring site to cause conform ational change and confer Hgand independent functional gain (N eüson and Friesel, 1995;Neilson and Friesel, 1996;Galvin et al, 1996;Mangasarian et al, 1997;R obertson et al, 1998). Previous investigation o f m u tan t F G F R 2 signalling has been undertaken in m odel systems w hich lack the profile o f co factors and signalHng m olecules th at is endogenous to hum an tissue. T hese studies cannot, therefore, directly inform attem pts at cHnical

gen o ty p e /p h en o ty p e correlation.

A series o f F G F R expression studies in norm al hum an foetal and infant parietal osteogenesis is reported, and contrasted w ith cases o f A p ert (FG FR 2 — P250R) and Pfeiffer (F G F R 2 — C278F) cranial m orphogenesis. T he two different m utations, previously fo u n d by — w’/ro analyses to have different m odes o f activation, have similar effects u p o n the /« — siïu

expression o f FG FR2. Taken together, previous m — vitro and the current in — situ analyses in hum an tissues m ay provide a m olecular basis for their differences in craniofacial form .

3.2.3 Materials & Methods