Homozygous missense mutation in MSH5 identified in POI pedigree
Two sisters (III4 and III5) from the non-consanguineous Han Chinese family (Fig. 1A), aged 31 and 29 years, experienced oligomenorrhea since menarche (14 and 13 years old), and amenorrhea occurred approximately 10 years later (Table 1). Both of them have elevated serum FSH, infantile uteri, and atrophic ovaries devoid of follicles. Chromosomal abnormalities, FMR1 premutation, autoimmune disorders, previous ovarian surgery or chemo-/radiotherapy were absent in any of the family members.
Figure 1 Pedigree of a family with two daughters afflicted by POI and homozygous MSH5 p.D487Y variant. (A) The pedigree of the index family, ascertained through III5. WES was performed on the family members labeled with asterisk, and those labeled with genotypes were available for Sanger sequencing. “T” denotes the mutant MSH5 allele, and “G” wild type. Arrow indicates the proband. (B) The location of p.D487Y variant is in the DNA-binding domain of MSH5, and the residue is conserved from saccharomyces to human (C). (D) shows the mRNA level of MSH5 in fetal tissues, which is significantly higher in ovary than others. (E) The RT-PCR in fetal ovary, human granulosa cells (hGCs, obtained from one patient receiving in vitro fertilization treatment) and COV434 cells, shows that MSH5 is also highly expressed in adult granulosa cells. MT, mutant; and WT, wild type.
Table 1 Clinical features of familial and sporadic POI patients with mutations in MSH5 a1–2 mg daily dose of oral estradiol valerate tablet for 21 days plus oral micronized progesterone (200 mg/days) for 12 days each month.
In this POI pedigree, two affected siblings (III4, III5), both parents (II3, II4), and one unaffected daughter (III6) had been chosen to perform WES (Fig. 1A). In an autosomal-recessive inheritance model, 2 nonsynonymous homozygous variants in MSH5 (MIM 603382, chromosome 6p21.33) and ZNF391 (Zinc Finger Protein 391, chromosome 6p22.1) were revealed. Sanger sequencing confirmed that neither of the two homozygous variants were present in unaffected family members and both of the variants were absent in 400 fertile women. The ZNF391 variant (ENST00000244576: c.187A > G, p.S63G) was predicted to be benign by Polyphen2, and the Serine residue mutated was not conserved among species (Supplementary Material, Fig. S1). Furthermore, ZNF391 has not been related to any human disease and no mutation was found in 200 sporadic patients with POI. Therefore, the MSH5 variant (ENST00000375755: c.1459G > T, p.D487Y) remained as the only potential candidate for this POI family. Sanger sequencing for MSH5 in sporadic cases with POI identified 3 additional heterozygous mutations (ENST000 00375755: c.1057C > A, p.L353M; c.1459G > T, p.D487Y and c.2107 A > G, p.I703V), which had not been reported in either the Exome Variant Server or 1000 Genomes database. Among them, p.L353M and p.D487Y located in the DNA-binding domain and the original residues were highly conserved among species from yeast to human, while p.I703V occurred at the less conserved residue located at the ATPase domain (Supplementary Material, Fig. S2).