Stage-specific expression of the glycine cleavage complex subunits in Leishmania infantum

The mitochondrial glycine cleavage complex (GCC) is an important part of cellular metabolism due to its role in the maintenance and balance of activated one-carbon units for a wide range of biosynthetic processes. In the protozoan parasite Leishmania, little is known about these metabolic processes. However, the importance of amino acid catabolism, especially for the clinically relevant amastigote form of this parasite, is becoming increasingly clear. Using a bioinformatics approach, we have identified orthologs of the genes encoding the four loosely associated GCC subunits (GCVP, GCVT, GCVH, and GCVL) in the visceral species Leishmania infantum. We report here that all GCC genes are expressed in L. infantum and that several are enriched in the intracellular amastigote stage. To further assess the regulation of GCC components throughout the life cycle of Leishmania, we focused on the T-protein component GCVT. GCVT is encoded by two almost identical tandemly arranged gene copies that have very divergent 3'UTRs. Using two different reporter gene systems, we demonstrate that the divergent GCVT 3'UTRs are responsible for the differential regulation of GCVT-1 and GCVT-2 isogenes at the protein level in both developmental forms of L. infantum. The GCVT-1 3'UTR is responsive to heat stress, resulting in higher expression of GCVT-1 in promastigotes, whereas the GCVT-2 3'UTR harbors a SIDER2 retroposon, which contributes to the amastigote-specific expression of GCVT-2 protein. Interestingly, our data indicate that expression of most GCC genes is inducible upon excess glycine and that this regulation is not conferred by 5'- or 3'-untranslated regions. Altogether, these data suggest a complex and multilayered regulation of the GCC both at the mRNA and protein levels throughout the L. infantum life cycle.