Phosphatidylinositol 3-phosphate regulates iron transport via PI3P-binding CgPil1 protein

Iron homeostasis, which is pivotal to virulence, is regulated by the phosphatidylinositol 3-kinase CgVps34 in the human fungal pathogen Candida glabrata. Here, we identify CgPil1 as a phosphatidylinositol 3-phosphate (PI3P)-binding protein and unveil its role in retaining the high-affinity iron transporter CgFtr1 at the plasma membrane (PM), with PI3P negatively regulating CgFtr1-CgPil1 interaction. PI3P production and its PM localization are elevated in the high-iron environment. Surplus iron also leads to intracellular distribution and vacuolar delivery of CgPil1 and CgFtr1, respectively, from the PM. Loss of CgPil1 or CgFtr1 ubiquitination at lysines 391 and 401 results in CgFtr1 trafficking to the endoplasmic reticulum and a decrease in vacuole-localized CgFtr1. The E3-ubiquitin ligase CgRsp5 interacts with CgFtr1 and forms distinct CgRsp5-CgFtr1 puncta at the PM, with high iron resulting in their internalization. Finally, PI3P controls retrograde transport of many PM proteins. Altogether, we establish PI3P as a key regulator of membrane transport in C. glabrata. [Display omitted] •The phosphatidylinositol 3-kinase CgVps34 controls plasma membrane protein abundance•The eisosome core component protein CgPil1 binds to phosphatidylinositol 3-phosphate•High iron increases PI3P levels and vacuolar delivery of CgFtr1 (iron transporter)•CgPil1 aids CgFtr1 retention at the plasma membrane in response to iron availability Askari et al. uncover the mechanism underlying PI3K-dependent retrograde trafficking of the iron transporter CgFtr1 from the plasma membrane (PM). They find that the PI3P-binding eisosome protein CgPil1 retains CgFtr1 at the PM, with surplus environmental iron elevating synthesis and PM localization of PI3P, which releases CgFtr1 from CgPil1, followed by CgFtr1 ubiquitination.