RESUMO
Yeast cells can extrude intracellular drugs through membrane-associated efflux pumps, such as ATP-binding cassette (ABC) transporters and members of the major facilitator superfamily. Gene expression of drug efflux pumps is regulated by several transcription factors involved in pleiotropic drug resistance (PDR). Salicylic acid (SA) possesses weak antifungal activity. Although the excretion mechanisms of some antifungal drugs have been revealed, the mechanism of SA extrusion remains unclear. To elucidate the mechanism of SA excretion, we screened SA-resistant mutants from random mutagenized Saccharomyces cerevisiae BY4741 cells. We successfully isolated 60 SA-resistant clones (KinSal001-060). KinSal052, one of the strongest SA-resistant clones, also exhibited resistance to 4-nitroquinoline-1-oxide and cycloheximide, indicating that it acquired the PDR phenotype. We identified a novel mutation in YRR1 conferring SA resistance to KinSal052. YRR1 encodes a Zn(II)2Cys6-type zinc-finger transcription factor that reportedly activates gene expression involved in PDR. Yeast cells carrying the yrr1 allele (yrr1-52) activated expression of several efflux pump-encoding genes, including YOR1, SNQ2, AZR1, and FLR1. These results suggested that SA resistance in KinSal052 is conferred by the overexpression of efflux pumps constitutively activated by the mutant form of Yrr1p.
Assuntos
Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/genética , Ácido Salicílico/farmacologia , Fatores de Transcrição/genética , 4-Nitroquinolina-1-Óxido/farmacologia , Transportadores de Cassetes de Ligação de ATP/genética , Sequência de Aminoácidos , Antifúngicos/metabolismo , Antifúngicos/farmacologia , Cicloeximida/farmacologia , Farmacorresistência Fúngica Múltipla/genética , Regulação Fúngica da Expressão Gênica , Genes Fúngicos , Proteínas de Membrana Transportadoras/genética , Dados de Sequência Molecular , Mutação , Transportadores de Ânions Orgânicos/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Ácido Salicílico/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Idiopathic pulmonary alveolar proteinosis (IPAP) is a rare disease characterized by excessive amounts of lipoproteinaceous material in the alveolus. This report presents an interim analysis of nationwide epidemiological data from Japanese patients with pulmonary alveolar proteinosis, and the roles of serum markers for IPAP. (i) The nationwide demographic data from 166 Japanese patients with IPAP are shown. The female to male ratio was 1:2, and the average age was 51 +/- 14 years old (age range: 15-79 years) at registration or diagnosis. A total of 30% of patients with IPAP have a poor clinical course. In total, 30% of patients were treated with whole lung lavage therapy (WLL). Under WLL, the patients significantly improved in the short term, but 40% of the patients who underwent WLL worsened again. A new strategy such as granulocyte-macrophage colony-stimulating factor (GM-CSF) therapy for intractable PAP is required. (ii) The correlation of serum KL-6, carcinoembryonic antigen, surfactant proteins D and A, and LDH with disease severity suggests their potential as disease markers. In contrast, serum anti-GM-CSF antibody did not correlate with disease severity, but is a specific marker for the diagnosis of IPAP. The combined measurements of the serum markers may well prove very useful for both the diagnosis and the management of IPAP patients.
