In search of a physiological role for thiamine triphosphate and the 25-kDa thiamine triphosphatase

In search of a physiological role for thiamine triphosphate and the 25-kDa thiamine triphosphatase
L. BETTENDORFF, B. LAKAYE, G. KOHN AND P. WINS
GIGA-Neurosciences, University of Liège, 4000-Liège, Belgium
Thiamine triphosphate (ThTP) was discovered over 60 years ago. Although it is present in most organisms from bacteria to mammals, its possible biological functions remain unclear. In contrast to thiamine diphosphate (ThDP), it is not a coenzyme. In E. coli cells, ThTP is transiently produced in response to amino acid starvation, while in mammalian cells, it is constitutively produced at a low rate. In some animal tissues, ThTP was able to phosphorylate proteins and activate a high-conductance anion channel in vitro. These observations raised the possibility of ThTP being part of a still uncharacterized cellular signaling pathway. Though it was long thought that ThTP is synthesized by a specific ThDP:ATP phosphotransferase, more recent studies indicate that two main mechanisms are involved: (1) in the cytosol adenylate kinase 1 can catalyze ThTP production from ThDP and ADP and (2) in brain mitochondria FoF1-ATP synthase can catalyze ThTP production from ThDP + Pi. The latter reaction is energized by the respiratory chain through a chemiosmotic mechanism analogous to oxidative phosphorylation. Both mechanisms are conserved from bacteria to mammals. While ThTP synthesis does not seem to require a specific enzyme, its hydrolysis in mammalian tissues is catalyzed by a very specific cytosolic 25 kDa thiamine triphosphatase (ThTPase). Because of this activity, steady-state ThTP levels are kept low in mammalian cells. ThTPase belongs to the CYTH superfamily of proteins which has representatives in all superkingdoms of life acting on tripolyphosphate and various triphosphorylated substrates. Although the whole chromosome region containing the ThTPase gene was lost in birds, orthologs of the ThTPase gene were found in all other known metazoan genomes. It seems that ThTPase activity appeared as a secondary acquisition of the CYTH proteins in the lineage leading from cnidarians to vertebrates. In particular, the Trp-53 residue of mammalian ThTPases plays a key role in substrate recognition and specificity by interacting with the thiazole part of ThTP. This residue is conserved in metazoan CYTH proteins with ThTPase activity. In order to gain insight into the physiological function(s) of the ThTP-ThTPase couple, we tried to produce a mouse invalidated in 25-kDa ThTPase. Surprisingly, we were unable to obtain any knockout animal, apparently because ThTPase seems to be required for spermatogenesis. As we previously showed that the enzyme is much more abundant in differentiated versus undifferentiated cells, we suspect that 25-kDa ThTPase might play a more general and important role during cell differentiation.

Acknowledgments
This work was supported by the F.R.S.-FNRS. LB and BL are respectively Research Director and Research Associate at the F.R.S.-FNRS.