Specificity of Membrane-Associated J-Domain Protein, Caj1, in Amphotericin B Tolerance in Budding Yeast

Hsp70:J-domain protein (JDP) machineries play pivotal roles in maintaining cellular proteostasis and governing various aspects of fungal physiology. While Hsp70 is known for its involvement in conferring tolerance to diverse antifungal drugs, the specific contribution of JDPs remains unclear. In this study, we examined the sensitivity of cytosolic JDP deletion strains of budding yeast to amphotericin B (AmB), a polyene antifungal agent widely utilized in fungal disease treatment due to its ability to disrupt the fungal plasma membrane (PM). Deleting Caj1, a PM-associated class II JDP, heightened susceptibility to AmB, and the protection conferred by Caj1 against AmB necessitated both its N-terminal J-domain and C-terminal lipid binding domain. Moreover, Caj1 deficiency compromised PM integrity as evidenced by increased phosphate efflux and exacerbated AmB sensitivity, particularly at elevated temperatures. Notably, phytosphingosine (PHS) addition as well as overexpression of PMP3, a positive PM integrity regulator, significantly rescued AmB sensitivity of caj1Δ cells. Our results align with the notion that Caj1 associates with the PM and cooperates with Hsp70 to regulate PM proteostasis, thereby influencing PM integrity in budding yeast. Loss of Caj1 function at the PM compromises PM protein quality control, thereby rendering yeast cells more susceptible to AmB.