Characterization of the Pneumocystis jirovecii and Pneumocystis murina phosphoglucomutases (Pgm2s): a potential target for Pneumocystis therapy

Pneumocystis cyst life forms contain abundant beta-glucan carbohydrates, synthesized using beta-1,3 and beta-1,6 glucan synthase enzymes and the donor uridine diphosphate (UDP)-glucose. In yeast, phosphoglucomutase (PGM) plays a crucial role in carbohydrate metabolism by interconverting glucose 1-phosphate and glucose 6-phosphate, a vital step in UDP pools for beta-glucan cell wall formation. This pathway has not yet been defined in Pneumocystis. Herein, we surveyed the Pneumocystis jirovecii and Pneumocystis murina genomes, which predicted a homolog of the Saccharomyces cerevisiae major PGM enzyme. Furthermore, we show that PjPgm2p and PmPgm2p function similarly to the yeast counterpart. When both Pneumocystis pgm2 homologs are heterologously expressed in S. cerevisiae pgm2 Delta cells, both genes can restore growth and sedimentation rates to wild-type levels. Additionally, we demonstrate that yeast pgm2 Delta cell lysates expressing the two Pneumocystis pgm2 transcripts individually can restore PGM activities significantly altered in the yeast pgm2 Delta strain. The addition of lithium, a competitive inhibitor of yeast PGM activity, significantly reduces PGM activity. Next, we tested the effects of lithium on P. murina viability ex vivo and found the compound displays significant anti-Pneumocystis activity. Finally, we demonstrate that a para-aryl derivative (ISFP10) with known inhibitory activity against the Aspergillus fumigatus PGM protein and exhibiting 50-fold selectivity over the human PGM enzyme homolog can also significantly reduce Pmpgm2 activity in vitro. Collectively, our data genetically and functionally validate phosphoglucomutases in both P. jirovecii and P. murina and suggest the potential of this protein as a selective therapeutic target for individuals with Pneumocystis pneumonia.