25KDa-Thiamine triphosphatase is essential for spermatozoid development in mice

For many years, our laboratory has been interested in thiamine triphosphate (ThTP), a vitamin B1 derivative whose metabolism and physiological role remain unclear. Regarding its production, we have shown that in E. coli and in rat brain mitochondria, ThTP is synthesized from thiamine diphosphate and inorganic phosphate through a chemiosmotic mechanism involving the FoF1- ATP synthase [1, 2]. In mammalian cells, its concentration is maintained at a low level through hydrolysis by a very specific cytosolic 25-kDa thiamine triphosphatase (ThTPase) [3]. In order to gain insight in the role of ThTP and ThTPase in mammalian tissues, we decided to generate a mouse strain invalidated in 25kDa-ThTPase with the hope that these mice will accumulate ThTP in their tissues. We obtained genetically modified embryonic stem (ES) cells from the Knockout Mouse Project (KOMP) repository. In those cells, one of the 25kDa-ThTPase alleles was replaced by a construction containing the lacZ and the neomycin resistance genes. Those ES cells were microinjected in blastocysts and the chimeric blastocysts were injected in a mouse uterus to generate chimerae. However, when we bred those mice with wild type mice, the construction was never transmitted to the pups. To explain this result, we selected those chimerae that presented a sex-reversal. In those mice, all the spermatozoids derive from the injected embryonic stem cells, so that half of the spermatozoids are expected to harbor the construction. However, after qPCR analysis, we observed that the spermatozoids with the construction were outnumbered by a factor of thousand. These results strongly suggest that the 25kDa-ThTPase is required for spermatozoid development. 1. Gangolf, M., Wins, P., Thiry, M., El Moualij, B. & Bettendorff, L. (2010) J. Biol. Chem. 285, 583-94. 2. Gigliobianco, T., Gangolf, M., Lakaye, B., Pirson, B., von Ballmoos, C., Wins, P. & Bettendorff, L. (2013) Scientific reports. 3, 1071. 3. Lakaye, B., Makarchikov, A. F., Antunes, A. F., Zorzi, W., Coumans, B., De Pauw, E., Wins, P., Grisar, T. & Bettendorff, L. (2002) J. Biol. Chem. 277, 13771-7