Formation of cilia and flagella is a tightly controlled process
organized by intraflagellar transport (IFT), for which two
motor proteins, kinesin-2 and dynein, are responsible. The exact
functions of IFT and IFT-related proteins in spermiogenesis are poorly
understood in mammalian species. To investigate mechanisms of
IFT during mouse spermatogenesis, a male germ cell-specific
knockout mouse model for kinesin-2 subunit kinesin-like protein
3A (KIF3A) was generated. Depletion of KIF3A caused defects in
sperm tail development and the core structure, the axoneme,
failed to form. A transient microtubular platform, the manchette,
surrounds the sperm head during spermiogenesis. In
KIF3A-depleted mice, the manchette appeared constricted and its
clearance was delayed, causing abnormal head shape, suggesting that IFT
and KIF3A function are also important for manchette function. In
addition, we identified the KIF1-binding protein (KBP) and the
meiosis-specific nuclear structural protein 1 (MNS1) as a novel
interaction candidates for KIF3A.



Sperm flagellar protein 2 (SPEF2) has been shown to
localize in elongating spermatids, and its interaction with
IFT-related protein IFT20 has been established. To
characterize the function of SPEF2 in sperm tail formation, a
male germ cell-specific conditional knockout mouse model for
SPEF2 was generated (Spef2 cKO). Depletion of SPEF2 caused
defects in axoneme formation, and similar defects in the manchette
and head shaping were observed in KIF3A-depleted mice. We also
identified cytoplasmic dynein 1 and GOLGA3 as novel interaction
candidates for SPEF2 in the testis. Inhibition of dynein 1
activity in the manchette blocked the SPEF2 transport,
suggesting its role in manchette-related transport. IFT20 and SPEF2
localization in the Golgi complex and the delayed appearance of
IFT20 in the manchette in Spef2 cKO suggests that SPEF2 functions
as an adaptor in dynein 1-mediated transport in elongating spermatids.
This study increases our understanding of the protein transport
mechanisms required for the formation of functional spermatozoa.