Abstract
TTC12 is a cytoplasmic and centromere-localized protein that plays a role in the proper assembly of dynein arm complexes in motile cilia in both respiratory cells and sperm flagella. This finding underscores its significance in cellular motility and function. However, the wide role of TTC12 in human spermatogenesis-associated primary ciliary dyskinesia (PCD) still needs to be elucidated. Whole-exome sequencing (WES) and Sanger sequencing were performed to identify potentially pathogenic variants causing PCD and multiple morphological abnormalities of sperm flagella (MMAF) in an infertile Pakistani man. Diagnostic imaging techniques were used for PCD screening in the patient. Real-time polymerase chain reaction (RT‒PCR) was performed to detect the effect of mutations on the mRNA abundance of the affected genes. Papanicolaou staining and scanning electron microscopy (SEM) were carried out to examine sperm morphology. Transmission electron microscopy (TEM) was performed to examine the ultrastructure of the sperm flagella, and the results were confirmed by immunofluorescence staining. Using WES and Sanger sequencing, a novel homozygous missense variant (c.C1069T; p.Arg357Trp) in TTC12 was identified in a patient from a consanguineous family. A computed tomography scan of the paranasal sinuses confirmed the symptoms of the PCD. RT–PCR showed a decrease in TTC12 mRNA in the patient’s sperm sample. Papanicolaou staining, SEM, and TEM analysis revealed a significant change in shape and a disorganized axonemal structure in the sperm flagella of the patient. Immunostaining assays revealed that TTC12 is distributed throughout the flagella and is predominantly concentrated in the midpiece in normal spermatozoa. In contrast, spermatozoa from patient deficient in TTC12 showed minimal staining intensity for TTC12 or DNAH17 (outer dynein arms components). This could lead to MMAF and result in male infertility. This novel TTC12 variant not only illuminates the underlying genetic causes of male infertility but also paves the way for potential treatments targeting these genetic factors. This study represents a significant advancement in understanding the genetic basis of PCD-related infertility.
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Data availability
All the data will be available on kindly request by contacting the corresponding author (qshi@ustc.edu.cn).
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Acknowledgements
This work was supported by the National Natural Science Foundation of China (82171599, U21A20204 and 82071709), the National Key Research and Developmental Program of China (2022YFC2702601, 2022YFA0806303, 2021YFC2700202 and 2019YFA802600), the Global Select Project (DJK-LX-2022010) of the Institute of Health and Medicine, Hefei Comprehensive National Science Center, and the Joint Fund for New Medicine of USTC (YD9100002034).
Funding
National Natural Science Foundation of China, 82171599, Qinghua Shi, U21A20204, Qinghua Shi, 82071709, Qinghua Shi, National Key Research and Developmental Program of China, 2022YFC2702601, Qinghua Shi, 2022YFA0806303, Qinghua Shi, 2021YFC2700202, Qinghua Shi, 2019YFA802600, Qinghua Shi, Global Select Project, DJK-LX-2022010, Qinghua Shi, Joint Fund for New Medicine of USTC, YD9100002034, Qinghua Shi.
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IA wrote the manuscript and analyzed the data. IA, HA, FR, AH and AZ collected patient samples and performed semen analysis. IA, KK, and TA: performed the experiments. HZ: performed WES and Sanger sequencing. HM, XJ, MAK, SD and MZ analyzed the data. BX, AU and WS: reviewed and edited the manuscript. QS: Conceived and supervised the study.
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All the data will be available on kindly request by contacting the corresponding author (qshi@ustc.edu.cn). The Institutional Ethical Committee of the University of Science and Technology of China (IEC-USTC) reviewed and approved the studies involving human participants with approval number 2019-KY-168. To be part of this study, the patients/participants provided written consent. Each participant completed an informed consent form before the study started.
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Communicated by Shuhua Xu.
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438_2024_2161_MOESM2_ESM.pptx
Supplementary file2 Figure S1: Whole exome sequencing data analysis pipeline. WES, whole exome sequencing; LOD, logarithm of odds; MAF, minor allele frequency. Figure S2: Chest X-rays and HRCT scan. Figure S3: Schematic representation of previously reported variants in TTC12 (PPTX 481 KB)
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Ali, I., Ali, H., Unar, A. et al. A novel homozygous missense TTC12 variant identified in an infertile Pakistani man with severe oligoasthenoteratozoospermia and primary ciliary dyskinesia. Mol Genet Genomics 299, 69 (2024). https://doi.org/10.1007/s00438-024-02161-2
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DOI: https://doi.org/10.1007/s00438-024-02161-2