Identification of bi-allelic KIF9 loss-of-function variants contributing to asthenospermia and male infertility in two Chinese families

Abstract

IntroductionAsthenozoospermia (AZS) is a leading cause of male infertility, affecting an estimated 18% of infertile patients. Kinesin proteins function as molecular motors capable of moving along microtubules. The highly conserved kinesin family member 9 (KIF9) localizes to the central microtubule pair in the flagella of Chlamydomonas cells. The loss of KIF9 expression in mice has been linked to AZS phenotypes.MethodsVariant screening was performed by whole exome sequencing from 92 Chinese infertile patients with AZS. Western blot was used to was used for analyzing of candidate proteins expression. Patients’ sperm samples were stained with immunofluorescent to visualise proteins localization and were visualised by transmission electron microscopy (TEM) to determine axoneme structures. Co-immunoprecipitation assay was used to verify the binding proteins of KIF9. In vitro fertilization (IVF) was used to evaluate the efficiency of clinical treatment.ResultsBi-allelic KIF9 loss-of-function variants were identified in two unrelated Chinese males exhibiting atypical sperm motility phenotypes. Both of these men exhibited typical AZS and suffered from infertility together with the complete absence of KIF9 expression. In contrast to these KIF9-deficient patients, positive KIF9 staining was evident throughout the flagella of sperm from normal control individuals. KIF9 was able to interact with the microtubule central pair (CP) component hydrocephalus-inducing protein homolog (HYDIN) in human samples. And KIF9 was undetectable in spermatozoa harboring CP deletions. The morphologicy of KIF9-deficient spermatozoa appeared normal under gross examination and TEM. Like in mice, in vitro fertilization was sufficient to overcome the fertility issues for these two patientsDiscussionThese findings indicate that KIF9 associates with the central microtubules in human sperm and that it functions to specifically regulate flagellar swinging. Overall, these results offer greater insight into the biological functions of KIF9 in the assembly of the human flagella and its role in male fertility.

The kinesin motor Kif9 regulates centriolar satellite positioning and mitotic progression

“…Its activity has been linked to mRNA transport 19 , macrophage podosome homeostasis 20 , and mitosis 21 . It has also appeared in screens related to breast cancer 22 , glioblastoma 23 , mouse inner ear development 24 , nuclear movement during skeletal muscle differentiation 25 , and sperm flagellum formation and maintenance 26,27 . However, Kif9’s fundamental cellular function remains poorly understood.…”

Characterizing the in vitro motor properties of two kinesin-9 family members from Tetrahymena

“…Kinesin-9A is an axonemal component protein localized along one of the central pair of microtubules (C2) [17][18][19] . The loss of kinesin-9A function causes defective ciliary motility in multiple species [20][21][22][23] . Structural studies have revealed that kinesin-9A adopts two states on the central microtubule, positioned 8 nm apart [17] , suggesting that kinesin-9A movement may regulate dynein-induced ciliary bending [17,18,24] .…”

Distinct Roles of Kif6 and Kif9 in Mammalian Ciliary Trafficking and Motility

“…Although there is no observed movement of KIF9B within flagella, it is speculated that it is involved in PFR transport or the recognition of axoneme asymmetry for PFR assembly in Trypanosoma (Demonchy et al, 2009). In mammals, mutations in Kif9 have been linked to defective sperm motility and impaired fertility (Meng et al, 2022;Miyata et al, 2020). However, its function in motile cilia remains unexplored.A frameshift mutation of KIF6 has been found in a patient with intellectual disability and megalocephaly (Konjikusic et al, 2018).…”

“…This raises the question of whether Kif6, in other species, also contributes to ciliary trafficking of essential materials involved in cilia beating and recognition or establishment of axonemal polarity. In mammals, mutations in Kif9 have been associated with defective sperm motility and impaired fertility (Meng et al, 2022; Miyata et al, 2020). A frameshift mutation of KIF6 has been found in a patient with intellectual disability and megalocephaly (Buchan et al, 2014; Konjikusic et al, 2018).…”