| The opportunistic fungal pathogen Candida albicans is both a benign gut commensal and a frequently fatal systemic pathogen.If the host’s immune system is compromised,C.albicans infects the host and causes inflammation.Ysy6 of Saccharomyces cerevisiae may suppress the secretion defect of a secY mutant in Escherichia coli.Its mammalian homologue is stress-associated endoplasmic reticulum protein 1(SERP1),also known as ribosome-associated membrane protein 4(RAMP4),which is a Sec61-associated polypeptide that is induced by ER stress.However,the function of Ysy6 remains to be investigated in C.albicans.In addition,Atg8 is a key protein for autophagosome formation during autophagy,which can also be activated by ER stress.In this study,the Ysy6 of C.albicans was identified,and the functions of Ysy6 and Atg8 were studied.The main results are showed as follows:(1)BLASTP analysis revealed the homologous protein of Saccharomyces cerevisiae Ysy6 protein in C.albicans genome databases.Their identity was 50.8%.The GFP localization revealed that Ysy6 is localized on the ER and vacuole.(2)YSY6 single deletion strain and double mutant of both YSY6 and ATG8 were constructed by PCR-mediated homologous recombination.Double mutant were found to be sensitive to the several transition metal ions and the glycosylation inhibitor TN.It shows that Ysy6 and Atg8 play a synergistic role in the process of ion and drug tolerance.(3)Under the treatment of TN,the double mutant had a high percentage of the PI –positive cells and VMP-positive cells.Their ROS levels and expression of OSR genes increased.Deletion of both YSY6 and ATG8 resulted in severe mitochondrial dysfunction under this stress,such as the morphologic changes of mitochondria,the decrease of membrane potential and the decrease of ATP contents.At the same time,the double mutation attenuated splicing of HAC1 mRNA,and decreased level of the unfolded protein response(UPR)activation.These results suggest that Ysy6 and Atg8 may affect the splicing of HAC1 mRNA in the face of ER stress,thereby activating the UPR.(4)Mutation of both YSY6 and ATG8 affected the morphogenesis of Candida albicans.The abilities of hyphae development,flocculation,embedded growth,adhesion,and biofilm formation in double mutant were significantly reduced.The morphogenesis also influenc the pathogenicity of Candida albicans.For example,the ability to secrete phospholipase and aspartic proteases was reduced in the Candida albicans.The ability to withstand macrophage attack was also significantly decreased,and the ability to infection of mice was lost completely,indicating that Ysy6 and Atg8 are critical to the virulence of C.albicans and they are potential virulence factors of C.albicans.In this study,we mainly found that Ysy6 and Atg8 palyed a significant role in ER stress response process,morphogenesis and pathogenicity,which enriched the content of mechanism in ER stress response,and provided a theoretical basis for developing new antifungal drugs in clinical research. |
