Discussion

Trypanosoma brucei is held poised for transmission to it’s tsetse vector by the activity of a tyrosine phosphatase, TbPTP1. We have discovered that the substrate of TbPTP1 is a glycosomal phosphatase, TbPIP39, whose enzymatic activity is repressed by PTP1-mediated dephosphorylation. Interestingly, a feedback regulation operates whereby inactive PIP39 stimulates PTP1, reinforcing its own repression. The composition of trypanosome glycosome is very different in bloodstream (BSF) and procyclic-forms (PCF). Moreover, during differentiation there is rapid pexophagy, which quickly turns over existing glycosomes, promoting metabolic adaptation. We envisaged that PIP39 might (a) be directed to all glycosomes during differentiation, or (b) selectively target only the newly-synthesised PCF-type glycosomes, with BSF-type glycosomes remaining PIP39-negative and being rapidly degraded. We used PIP39-specific antibody to stain stumpy cells undergoing differentiation for the relative colocalisation of PIP39 with constitutive glycosomal proteins present in both BSF and PCF-type glycosomes. Cell samples were assayed by fluorescence and dual-label confocal microscopy. We found that the PIP39 localisation changes as stumpy cells differentiate to procyclic forms, moving from the proximity of the  flagellar pocket to a glycosomal location. This rapid PIP39 relocalisation is visible within 1 hour of cis-Aconitate exposure and provides a novel early differentiation marker.