Pneumocystis phosphoglucomutase links glucose-phosphate metabolism to β-1,3-glucan synthesis and cell-wall integrity

The fungal cell wall is a dynamic, carbohydrate-rich structure whose composition is closely coordinated with cellular metabolism. Phosphoglucomutase (PGM) interconverts glucose-1-phosphate and glucose-6-phosphate, linking carbon metabolism with UDP-glucose-dependent cell-wall biosynthesis. We previously identified functional Pgm2 homologs from Pneumocystis jirovecii and Pneumocystis murina. Here, using a Saccharomyces cerevisiae pgm2Δ complementation model, we examined the relationship between Pgm2 activity, β-1,3-glucan, and cell-wall integrity. Pgm2 deficiency reduced β-1,3-glucan content and altered zymolyase sensitivity without major changes in cell-wall thickness. Bulk RNA sequencing revealed that the pgm2Δ mutant exhibited coordinated metabolic and cell-wall transcriptional responses, including increased SLT2/MPK1, RLM1, FKS2/GSC2, GFA1, CHS1, CRH1, SED1, and CWP1 expression and decreased FKS1, GAS1, GAS3, and CHS2 expression. Expression of either Pjpgm2 or Pmpgm2 broadly shifted these responses toward wild-type levels. Selected transcriptional changes were further evaluated by qRT-PCR, which supported the major stress and cell-wall responses identified by RNA sequencing. FKS2/GSC2 showed the same directional pattern, although differences were not statistically significant by qRT-PCR. Despite this compensatory response, Pgm2-deficient cells were markedly more susceptible to sublethal caspofungin, whereas either Pneumocystis Pgm2 homolog restored tolerance toward wild-type levels. These findings link Pgm2-dependent glucose-phosphate metabolism with β-1,3-glucan levels and cell-wall function. Given the importance of β-1,3-glucan to the Pneumocystis cyst wall and its limited cell-wall biosynthetic pathways, these findings further support Pgm2 as an important component of Pneumocystis cell-wall biology and a potential antifungal target.