MORITA, M.; Yamato, K. T.

Bryophytes, pteridophytes and some gymnosperm species produce motile, ciliated spermatozoids that navigate themselves to the egg by regulating ciliary motility in response to a concentration gradient of attractants released from the egg and/or surrounding cells. However, the structural components of spermatozoid cilia in the land plants are largely unknown. In this study, we investigated MpCAFA (combined CAPS and FAP115; Mp1g04120) in the liverwort Marchantia polymorpha. The N-terminal and near C-terminal regions of MpCAFA show similarity to calcyphosine (CAPS), a mammalian EF-hand protein, and to FAP115, a ciliary protein of the green alga Chlamydomonas reinhardtii, respectively. MpCAFA is expressed specifically in antheridia, and its orthologs are found in some algae, bryophytes, pteridophytes and cycads, but not in most of seed plants. Spermatozoids of mutants that lack functional MpCAFA exhibited significant decrease in swimming speed. Notably, these mutants showed no obvious morphological defect, including the 9+2 axoneme arrangement, and retained chemotactic capability and fertility, forming normal spores. This suggests that MpCAFA is required for spermatozoid motility but not for sperm chemotaxis or subsequent reproductive processes. Introduction of MpCAFApro:MpCAFA-mCitrine fully complemented the mutant phenotype and revealed MpCAFA-mCitrine was localized along the entire length of the two spermatozoid cilia. Both the CAPS-like and FAP115-like regions were essential for MpCAFA function and subcellular localization in spermatozoid, whereas the C-terminal proline-rich region was dispensable. These findings indicate that MpCAFA is a major ciliary protein in land plants and can serve as a marker to visualize spermatozoid ciliary movement.