TMEM67 encodes the transmembrane protein meckelin localizing to the primary cilium and to the plasma membrane. TMEM67 regulates canonical Wnt/β-catenin signaling pathway in the developing cerebellum(14). Pathogenic mutations in TMEM67 gene result in dysfunction of primary cilia during early embryogenesis. Primary cilia play an important role in tissue development and signal transduction, with mutations in ciliary-associated genes resulting a broad spectrum of human disease phenotypes (15). Biallelic variants in the TMEM67 gene cause COACH syndrome 1 (#216360), Joubert syndrome 6 (#610688), Meckel syndrome 3 (#607361), Nephronophthisis 11 (#613550), RHYNS syndrome (#602152) and Bardet-Biedl syndrome 14, modifier of (#615991). MKS3 causes by TMEM67 gene is the most severe ciliopathy with almost 100% mortality rate(16), central nervous system abnormalities (occipital encephalocele, hydrocephalus, anencephaly, holoprosencephaly, Dandy–Walker syndrome), bilateral renal polycystic kidneys with cystic dysplasia and polydactyly. Other related conditions are lip/palate cleft, heart defects, genital anomalies, liver fibrosis, and skeletal defects. The presence of earlier-onset oligohydramnios may lead to intrauterine death(17). MKS3 has a clinical and genetic overlap with other viable ciliopathies, especially Joubert syndrome and Joubert syndrome-related disorders(17, 18).
TMEM67 gene encodes meckelin protein with an extracellular N-terminus containing a signal peptide and a cysteine rich domain, a transmembrane portion and an intracellular C-terminus including a coiled-coil domain(18). Genotype-phenotype correlation analysis based on literatures show that biallelic null variants in TMEM67 gene is more common in lethal MKS3 than milder Joubert syndrome(19). The distribution of pathogenic missense variants along TMEM67 gene mainly clustered in the extra-cellular region, such as cysteine rich region, and the transmembrane regions(19). In our report, c.2314del (p.M772*) generates a premature stop codon that is expected to induce nonsense-mediated mRNA decay (NMD) mechanisms. c.1415T > G (p.V472G) locates in extra-cellular region with unknown function. The prediction of protein structure show that missense variant lead to changes in the hydrogen bond distance between amino acid residues, which may affect the stability of the meckelin. Pathogenic missense variants from exon 8 to exon 15 of TMEM67 gene occur simultaneously with null variants at the trans position is common in lethal MKS3 cases(18).
The process of clarifying the genetic cause of RPL in this family is very sad. This family selected for genetic diagnosis, until experienced forth times spontaneous abortions and intrauterine death caused by fetal structural malformations. In this process, the mother suffered great physical and psychological trauma. If prenatal WES testing and corresponding genetic counseling can be carried out early in the family, it would be of great help to prevent recurrence of adverse pregnancies.
In conclusion, this report shows the importance of exome sequencing in fetal with RPL, expands TMEM67 gene variant spectrum related to MKS3. The family in the study experienced multiple adverse pregnancy outcomes. TMEM67 gene explains the genetic cause of fetus with RPL, provides the opportunity for preimplantation genetic testing and avoids the same adverse pregnancy outcomes in next pregnancy. Furthermore, detailed prenatal phenotypes provide evidence for prenatal WES testing.