FPHH is a rare autosomal dominant disorder with variable penetrance caused by mutations in the KITLG gene, which encodes the C-Kit ligand . Because FPHH is very rare with reduced penetrance, no clear incidence rate of this disease has been documented. Patients with FPHH often do not have systemic symptoms. Growth retardation and intellectual disability in some affected members were reported by Westerhof et al. . A 2-year-old Chinese female FPHH patient with frequent seizures, impaired temperature regulation and susceptibility to infection also had mild intellectual disability.
Nine publications in PubMed (https://www.ncbi.nlm.nih.gov/pubmed) were related to pathogenic mutations of the KITLG gene. Mutations in the KITLG gene are associated with autosomal dominant nonsyndromic deafness-69 (DFNA69, MIM 616697), Waardenburg syndrome-2 (WS2, MIM 193510), and FPHH. Seco Z et al.  reported that certain mutations of KITLG, including c.286_303delinsT (p.Ser96Ter), c.200_202del (p.His67_Cys68delinsArg), and c.310C>G (p.Leu104Val), cause asymmetric and unilateral hearing loss and Waardenburg syndrome type 2 (WS2). Ogawa Y et al.  reported a patient with WS2 who had the unusual complication of large, pigmented macules caused by a homozygous KITLG mutation (c.94G>A, p.Arg32Cys). It was speculated that the mechanism of the mutation underlying WS2 leading to membrane incorporation and reducing secretion of KITLG occurs via a gain-of-function or dominant-negative effect. A de novo mosaic KITLG variant (NM_000899.3:c.329A>G; p.Asp110Gly) was found in a 6-year-old boy who had congenital linear and mottled hyperpigmentation . However, all the phenotypes presented in these three publications with KITLG are not defined as clearly associated with FPHH; therefore, we only summarize all the other KITLG mutations associated with FPHH here in this study.
Including the novel mutation (c.104A>T, p.Asn35Ile) we reported in this study, eight different missense mutations in the KITLG gene responsible for FPHH have been identified (Supplementary Table 3, Fig. 4). Seven out of eight mutations were clustered in a short amino acid sequence (VTNNV, amino acids 33-37) in exon 2 (Fig. 4), except for c.337G>A, which was in exon 4 of the KITLG gene. Most pathogenic mutations in FPHH occur within the VTNNV domain of the KITLG protein (amino acids 33-37) and lie within the third b-strand of the protein. Only the p.Val37 change represents the first amino acid of the second a-helix (amino acids 37-46). To date, the reported mutations are only involved in 33V, 34T, 36N, and 37V but not 35N. We are the first to report the c.104A>T (p.Asn35Ile) mutation at 35N (Fig. 4) with FPHH patients.
Except for diffuse hyper- and hypopigmentation, vast CAL-like lesions have been detected as the most common skin problems present in FPHH patients. Vitiligo was found in one family. Sparse lateral eyebrows and malignancy (pharyngeal cancer, papillary thyroid cancer and melanoma) were found in two families. Short sutures were found in only one family, and mental retardation was not present in these FPHH patients (Supplementary Table 3).
KITLG, as KIT LIGAND, is produced locally in human skin by epidermal keratinocytes and endothelial cells, where it induces the migration, development and survival of melanocytes. The signaling of KITLG and its receptor KIT plays an important role in melanocyte proliferation and pigment production [15, 16]. The role of the KIT/KITLG system in melanogenesis has been experimentally confirmed using animal models [3, 17]. After KITLG binds the c-KIT receptor, dimerization is triggered. It initiates signal transduction via the RAS/MAPK pathway to upregulate melanoblast proliferation [16, 18]. The KITLG/C-KIT/RAS/MAPK signaling pathways play important roles in the regulation of hematopoiesis, stem cell survival, gametogenesis, mast cell development, migration and function, and skin color [19, 20]. Mutant alleles of the KITLG gene are lethal in homozygous mice and produce a variable level of coat-color dilution in heterozygous mice . It has been reported in human studies that variations in the KITLG gene are also associated with skin, hair, and eye pigmentation (MIM 611664), autosomal dominant nonsyndromic deafness-69, WS2 and FPHH.
Here, we reported a novel c.104A>T (p.Asn35Ile) mutation of KITLG in a Chinese FPHH family. According to the 2015 ACMG guidelines, the mutation was initially identified as a ‘likely pathogenic’ mutation. To the best of our knowledge, only eight different missense KITLG mutations have been reported to cause FPHH (Supplementary Table 3). Notably, seven known mutations were clustered in a highly conserved short amino acid sequence VTNNV (amino acids 33-37) (Fig. 4). The VTNNV domain of the KITLG protein (amino acids 33–37) lies within the third b-strand of the protein and is responsible for its binding functions. Both mutations c.104A>T (p.Asn35Ile) and c.101C>T (p.Thr34Ile) found in this study were located in the VTNNV domain and were predicted to be detrimental variations by the SIFT and Polyphen-2 tools. Using the Swiss-Model servers [22, 23], three-dimensional structures of mutant KITLG proteins were found to change compared with the wild type (Fig. 3). Both 35Asn and 34Thr are polar, hydrophilic amino acids, and the mutant became nonpolar, hydrophobic isoleucine; therefore, it might change the features of the protein and affect the ligand affinity to its receptor c-Kit, thus affecting the migration of melanoblasts, melanosome transfer and melanin synthesis, conferring a phenotype with hyper- and hypopigmentation. The precise mechanisms need to be elucidated by further experiments. Our findings revealed a novel KITLG mutation associated with FPHH and reinforced the evidence that VTNNV was a hot spot for mutation. However, definitive functional analyses of this mutation are needed to determine the structure-function relationship in patients with FPHH.