XB-ART-59753Genomics 2023 May 01;1153:110630. doi: 10.1016/j.ygeno.2023.110630.
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Identification of rare loss-of-function variants in FAM3B associated with non-syndromic orofacial clefts.
Orofacial clefts (OFCs) are the most common congenital craniofacial disorders and cause serious problems with the appearance, orofacial function and mental health of the patients. The fibroblast growth factor (FGF) signaling pathway is critical for several aspects of craniofacial development and loss-of-function mutations of coding genes for multiple FGFs and FGFRs can lead to OFCs. We recently characterized FAM3B as a novel ligand of FGF signaling, which, through binding to FGFRs and activating downstream ERK, regulates craniofacial development in Xenopus. In this study, we identify two rare variants in FAM3B (p.Q61R and p.D128G) via target region sequencing of FAM3B on 144 unrelated sporadic patients with non-syndromic OFCs (NSOFCs). Bioinformatic analysis predict that these two variants are likely to be damaging and biochemical experiments show that these two variants weaken the FGF ligand activity of FAM3B by decreasing its expression and thus secretion. In summary, our results indicate that FAM3B is a novel candidate gene for NSOFCs in humans.
PubMed ID: 37105387
Article link: Genomics
Species referenced: Xenopus
Genes referenced: fam3b tbl1x
GO keywords: Wnt signaling pathway
Article Images: [+] show captions
|Fig. 1. Identification of rare loss-of-function variants (p.Q61R and p.D128G) in FAM3B associated with non-syndromic orofacial clefts. (A-B) Identification of p.Q61R and p.D128G variant in FAM3B gene in two unrelated patients with NSOFCs. Sanger sequencing chromatograms of mutated loci are labeled with arrow. (C) Location of FAM3B variants in the transcript (top) and protein (below). UTR: untranslated region; SP: signal peptide.|
|Fig. 2. In silico prediction of the pathogenicity of p.Q61R and p.D128G variants of FAM3B. (A) Frequencies of p.Q61R and p.D128G of FAM3B gene in databases of general populations. (B) Predicted pathogenicity of p.Q61R and p.D128G variant of FAM3B by various bioinformatic softwares. (C) Alignment of FAM3B protein sequences among vertebrates showing the conservation of amino acids affected by patient missense alterations (arrow). (D) Human FAM3B protein structure predicted by AlphaFold (https://alphafold.ebi.ac.uk/) showing the hydrogen bonds (H-bond) formed by Q61 with K59 and D64 (left) and D128 with R107, Y130, E131 and N134 (right). The Q61 and D128 substitutions are likely to disrupt the hydrogen bonds and thus the protein structure. Dashed boxes indicate that Q61 and D128 locate at α-helix and flexible structure, respectively. (E) ACMG classification of p.Q61R and p.D128G variants.|
|Fig. 3. p.Q61R and p.D128G variants of FAM3B show impaired capacity to activate ERK. (A) Hela cells transfected with His-tagged wild-type or mutant FAM3B were serum starved for 24 h and harvested for immunoblotting with the indicated antibodies. TUBULIN served as a loading control, which was detected in cell lysate, but not medium. Note that p.Q61R and p.D128G variants of FAM3B display reduced expression and secretion. (B) A flow chart summarizing our research rationale.|