Novel mutations in PMFBP1, TSGA10 and SUN5: Expanding the spectrum of mutations that may cause acephalic spermatozoa

To the Editor: Acephalic spermatozoa is a rare form of teratozoospermia, and the vast majority of cases are idiopathic. Furthermore,53.45% (51.22%, 21/41 SUN5; 50%, 7/14 PMFBP1; 100%, 1/1 TSGA10;100%, 1/1 BRDT; and 100%, 1/1 DNAH6) have been associated with a definitive genetic etiology. Only a few studies on this condition have been reported, and previous study populations were mainly from Hefei and Xiamen. Here, we performed a whole-exome analysis of 11 unrelated individuals with acephalic spermatozoa syndrome in Hunan, China, and subsequent variant filtering identified one compound heterozygous variant (c.361C>T[p.Gln121Ter] and c.2089-1G>T) in PMFBP1, one homozygous mutation (c.1739A>C[p.Gln580Pro]) in TSGA10, and one homozygous variant (c.772C>T[p.Arg258Cys]) in SUN5 in three unrelated infertile men (Figure 1A). The remaining mutations were novel gene results that should be further validated by animal models. We evaluated the influence of these mutations on sperm morphology and pregnancy outcomes. Although all patients exhibited acephalic spermatozoa, the phenotypes and proportions were slightly different (Figure 1B). Patient II-1 in family 1: None of his spermatozoa exhibited a normal morphology; ~100% were headless tails, and a few were tailless heads, which were all separated from the sperm flagella. The c.361C>T mutation in PMFBP1leads to a premature termination codon (PTC), which results in a very short N-terminus lacking functional structural maintenance of the chromosome (Smc) domain. The c.2089-1G>T[p.Ile697Leufs*257] mutation leads to the skipping of exon 15 and causes protein truncation resembling the knockout (Figure 1C). The splicing variation was classified as pathogenic according to the Association for Molecular Pathology guidelines. The subject's parents carried a heterozygous mutation. The c.361C>T heterozygous mutation from his mother was present in the ExAC (0) and GnomAD (5.43715e-05) databases, while the c.2089-1G>T mutation was absent in the ExAC and GnomAD databases. RT-PCR of the blood samples revealed that the mutation leads to aberrant splicing of mRNA. As of the date of this submission, the patient's wife had not yet entered the ICSI cycle. Patient II-1 in family 2: Spermatozoa with normal morphology comprised 1% of the population, while ~99% were headless tails. The subject had a homozygous mutation, c.1739A>C[p.Gln580Pro in exon 18] in TSGA10, which was found in the ExAC (0.0007), 1000G (0.001) and GnomAD (0.001) databases. Protein alignments showed that this variant in a C-terminal COG4372 domain (exons 16-19) was highly conserved among different species. SIFT, PolyPhen2 and PROVEAN predicted that this variant was a disease-causing mutation. This study represents a second confirmation report and implicates TSGA10 mutations as a cause of acephalic spermatozoa. The patient's wife underwent in vitro fertilization (IVF) treatment at our hospital. After 1 cycle of IVF-assisted reproductive treatment, 14 oocytes and 7 IVF + 7 ICSI.9 fertilized ova underwent cleavage and eventually formed four 8C embryos. Two embryos were subsequently implanted, and the patient's wife finally succeeded in becoming pregnant. Patient II-1 in family 3: Spermatozoa with a normal morphology comprised 5% of the total population, while ~70% were headless tails. A very small proportion of intact spermatozoa had an abnormal headtail junction, and a few tailless heads were observed, whereas the remaining 25% had other head deformities, such as microcephalic spermatozoa with or without a small acrosome structure, which differ from the so-called round-headed spermatozoa from Sun5 male mice but do not contain chromatin or acrosomes.SUN5 is the major gene involved in acephalic spermatozoa, and many mutations have been reported. A variant in SUN5 did not cause microcephalic spermatozoa in this patient, and no pleiotropic effects were obvious. Other pathogenic genes may be responsible for the multiple malformations in the patient's spermatozoa, which need to be further explored. The c.772C>T mutation in SUN5 is present in the ExAC (0) and GnomAD (0) databases and is predicted to be damaging by multiple in silico pathogenic prediction tools. The three-dimensional structure of the SUN5 protein was constructed by de novo modeling techniques. Mutation of Arg258 to cysteine may disturb the surrounding hydrogen bonds, affecting the overall structure of the SUN5 conserved region. Only one egg was obtained from the patient's wife, and no transplantable embryos or blastocysts were obtained. PMFBP1 interacts with SUN5 but does not directly bind to SUN5 or SPATA6, and PMFBP1 and TSGA10 have the same Smc superfamily domain. They may associate with the centrosome and basal body and participate in completing the coupling apparatus, ultimately resulting in acephalic spermatozoabut not delayed or stagnated embryonic development. We herein summarized known data on the genetic contribution of genes to acephalic spermatozoa (Figure 1D) and broadened the Received: 7 February 2020 Revised: 19 March 2020 Accepted: 20 March 2020