| Aspergillus fumigatus is a widespread saprophytic fungus in the natural environment,and it is also a conditional pathogenic fungus that can infect immunodeficient patients.Due to the wide use of immunosuppressants,the number of patients with immune deficiency is on the rise,which leads to the increasing number of invasive aspergillosis caused by A.fumigatus infection.At present,azole drugs are the most widely used antifungal drugs in clinic.However,due to the long-term and extensive application of azole drugs,a large number of azole drug-resistant strains lead to the decline of drug efficacy.Among the drug-resistant strains of A.fumigatus in clinic,the most common is caused by the mutation of Cyp51A,but there are still some drug-resistant strains caused by non-Cyp51A mutations.So far,many of these drug resistance mechanisms are not clear.Some studies have shown that A.fumigatus evolved some stress response pathways to external environmental stimulation,such as the pathway to deal with drug stress stimulation,the oxidative stress pathway of host immunity and so on,all of which play an important role in its drug resistance and survival.According to the genomic information of A.fumigatus and the functional protein comparison of homologous proteins in model yeast,two potential stress response proteins OrmA and MtmA,were selected to analyze their biological functions through the comprehensive means of molecular biology and cell biology.The results were summarized as follows:1.It is clarified that OrmA is involved in the drug susceptibility and fungal virulence by regulating sphingolipid synthesis.Deletion of ormA increases the sensitivity of A.fumigatus to azole drugs,and it is found that OrmA can regulate the metabolism of the sphingolipid by lipidomics.Deletion of ormA resulted in a significant increase of sphingolipid ceramide components,while adding the sphingolipid synthesis inhibitor myriocin can rescue the azole susceptibility induced by lack of ormA.These data indicate that OrmA is related to drug susceptibility by affecting the synthesis of sphingolipid.2.The expression of OrmA depends on the HacA,a major transcription factor in an unfolded protein response pathway（UPR）.OrmA can respond to not only drug stimulation but also endoplasmic reticulum pressure reagent dithiothreitol（DTT）or tunicamycin（TM）stimuli.Further studies have shown that the deletion of HacA led to a significant decrease in the expression of OrmA,indicating that HacA is very important for the expression of OrmA.3.The results of the mouse pulmonary infection model shows that the lack of OrmA decreases the mortality of immunodeficient mice,which indicating that OrmA is required for A.fumigatus virulence.4.It is confirmed that mitochondrial transporter MtmA is necessary for oxidative stress in A.fumigatus.The mtmA is an essential gene in A.fumigatus and the inhibition of MtmA expression by constructing the conditional strain causes extremely sensitive to oxidative stress reagents menadione and H2O2,which mainly result from a significant decrease in Sod2 enzyme activity.The overexpression of Sod2 can rescue the sensitivity to oxidative stress caused by inhibition of MtmA expression,indicating that MtmA can regulate Sod2-involved oxidative stress pathway.5.It Is found that the inhibition of MtmA expression affected the intracellular Zn ion balance of A.fumigatus,while Zn ion chelating agent TPEN can rescue the growth defects caused by the inhibition of MtmA expression.This suggests that the disorder of Zn ion metabolism is the key factor for the death caused MtmA deletion.6.The inhibition of MtmA expression significantly increase the tolerance of A.fumigatus to various antifungal agents.However,the low expression of MtmA leads to the significant decrease in the expression of the drug target Erg11A,Erg11B and Erg5,which is contradictory to its resistance.Further studies have found that the low expression of MtmA causes resistance to drug due to the overexpression of multiple drug pumps,especially Mdr1.However,the overexpression of the drug pump results from the continued activation of the transcription factor CrzA due to the low expression of MtmA.In conclusion,this article illustrates the stress response protein OrmA involved in azole drugs tolerance,and the deficiency of OrmA results in drug susceptibility and the over-expression of OrmA leading to drug resistance.OrmA is also required for the virulence of A.fumigatus.The mtmA in A.fumigatus is an essential gene and participates in oxidative stress response,ion balance,drug tolerance,etc.These findings of this study further broaden people’s understanding of the important proteins involved in antifungal drugs stress response in A.fumigatus,and and may also provide a theoretical basis for the future discovery of new antifungal drug target. |
