SPECC1L regulates palate development downstream of IRF6

Abstract

SPECC1L mutations have been identified in patients with rare atypical orofacial clefts and with syndromic cleft lip and/or palate (CL/P). These mutations cluster in the second coiled-coil and calponin homology domains of SPECC1L and severely affect the ability of SPECC1L to associate with microtubules. We previously showed that gene-trap knockout of Specc1l in mouse results in early embryonic lethality. We now present a truncation mutant mouse allele, Specc1lΔC510, that results in perinatal lethality. Specc1lΔC510/ΔC510 homozygotes showed abnormal palate rugae but did not show cleft palate. However, when crossed with a gene-trap allele, Specc1lcGT/ΔC510 compound heterozygotes showed a palate elevation delay with incompletely penetrant cleft palate. Specc1lcGT/ΔC510 embryos exhibit transient oral epithelial adhesions at E13.5, which may delay shelf elevation. Consistent with oral adhesions, we show periderm layer abnormalities, including ectopic apical expression of adherens junction markers, similar to Irf6 hypomorphic mutants and Arhgap29 heterozygotes. Indeed, SPECC1L expression is drastically reduced in Irf6 mutant palatal shelves. Finally, we wanted to determine if SPECC1L deficiency also contributed to non-syndromic (ns) CL/P. We sequenced 62 Caucasian, 89 Filipino, 90 Ethiopian, 90 Nigerian and 95 Japanese patients with nsCL/P and identified three rare coding variants (p.Ala86Thr, p.Met91Iso and p.Arg546Gln) in six individuals. These variants reside outside of SPECC1L coiled-coil domains and result in milder functional defects than variants associated with syndromic clefting. Together, our data indicate that palate elevation is sensitive to deficiency of SPECC1L dosage and function and that SPECC1L cytoskeletal protein functions downstream of IRF6 in palatogenesis.

SPECC1L-deficient primary mouse embryonic palatal mesenchyme cells show speed and directionality defects

“…We also show that SPECC1L deficiency impaired the ability of palatal mesenchyme cells to move collectivelya result that we posit partially explains the observed delayed and abnormal palate elevation in Specc1l cGT/ΔC510 mutants (Fig. 1) 19 . The defect in collective migration was also validated in U2OS cells, an independent cell line, which strongly supports a primary role for SPECC1L in collective movement.…”

“…However, palatal shelves in most mutants recover and fuse by E15.5 (Fig. 1) 19 . Analysis of E14.5 sections at mid-palate region indicated that the tongue in the mutant is usually adequately depressed, but the palatal shelves are acutely angled and do not immediately move inwards to occupy the space above the tongue (Fig.…”

“…Specc1l cGT/ΔC510 mutant embryos show abnormal palatogenesis. Specc1l cGT/ΔC510 compound heterozygotes are perinatal lethal and show a delay in palate elevation at E14.5 19 . However, palatal shelves in most mutants recover and fuse by E15.5 (Fig.…”

“…Analysis of E14.5 sections at mid-palate region indicated that the tongue in the mutant is usually adequately depressed, but the palatal shelves are acutely angled and do not immediately move inwards to occupy the space above the tongue (Fig. 2) 19 . Importantly, palate elevation in these mutants follows an abnormal sequence where posterior palate advances prior to middle and anterior palates, as in wildtype (WT) embryos (Fig.…”

A novel p.Pro871Leu missense mutation in SPECC1L gene causing craniosynostosis in a patient

“…SPECC1L cGT/ΔC510 compound mutant mice also exhibit palatal shelf elevation delay, and SPECC1L is a downstream cytoskeletal effector molecule in the palate that is regulated by the interferon regulatory transcription factor 6 (IRF6). SPECC1L regulates the palate development downstream of IRF6 29 . The collective data indicate that SPECC1L functions are necessary for normal facial morphogenesis and of its vital role in cell migration and adhesion.…”