Resistance Gene-Guided Genome Mining: Serial Promoter Exchanges in Aspergillus nidulans Reveal the Biosynthetic Pathway for Fellutamide B, a Proteasome Inhibitor.
Fungal genome projects are revealing thousands of cryptic secondary metabolism (SM) biosynthetic gene clusters that encode pathways that potentially produce valuable compounds. Heterologous expression systems should allow these clusters to be expressed and their products obtained, but approaches are needed to identify the most valuable target clusters. The inp cluster of Aspergillus nidulans contains a gene, inpE, that encodes a proteasome subunit, leading us to hypothesize that the inp cluster produces a proteasome inhibitor and inpE confers resistance to this compound. Previous efforts to express this cluster have failed, but by sequentially replacing the promoters of the genes of the cluster with a regulatable promotor, we have expressed them successfully. Expression reveals that the product of the inp cluster is the proteasome inhibitor fellutamide B, and our data allow us to propose a biosynthetic pathway for the compound. By deleting inpE and activating expression of the inp cluster, we demonstrate that inpE is required for resistance to internally produced fellutamide B. These data provide experimental validation for the hypothesis that some fungal SM clusters contain genes that encode resistant forms of the enzymes targeted by the compound produced by the cluster.
Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California , Los Angeles, California 90089, United States.
Hsu-Hua
HH
Yeh
Department of Molecular Biosciences, University of Kansas , Lawrence, Kansas 66045, United States.
Manmeet
M
Ahuja
Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California , Los Angeles, California 90089, United States.|Department of Pharmacy, Chia Nan University of Pharmacy and Science , Tainan City 71710, Taiwan.
Yi-Ming
YM
Chiang
Department of Molecular Biosciences, University of Kansas , Lawrence, Kansas 66045, United States.
C Elizabeth
CE
Oakley
Department of Molecular Biosciences, University of Kansas , Lawrence, Kansas 66045, United States.
Shauna
S
Moore
Department of Molecular Biosciences, University of Kansas , Lawrence, Kansas 66045, United States.
Olivia
O
Yoon
Department of Molecular Biosciences, University of Kansas , Lawrence, Kansas 66045, United States.
Heather
H
Hajovsky
Department of Bacteriology and Department of Medical Microbiology and Immunology, University of Wisconsin , Madison, Wisconsin 53706, United States.
Jin-Woo
JW
Bok
Department of Bacteriology and Department of Medical Microbiology and Immunology, University of Wisconsin , Madison, Wisconsin 53706, United States.
Nancy P
NP
Keller
Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California , Los Angeles, California 90089, United States.|Department of Chemistry, University of Southern California, Dornsife College of Letters, Arts, and Sciences , Los Angeles, California 90089, United States.
Clay C C
CC
Wang
Department of Molecular Biosciences, University of Kansas , Lawrence, Kansas 66045, United States.
Berl R
BR
Oakley
D001233:Aspergillus nidulans / Q000235:genetics*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016681:Genome, Fungal; D055666:Lipopeptides / Q000096:biosynthesis* / Q000494:pharmacology; D005810:Multigene Family; D011401:Promoter Regions, Genetic; D046988:Proteasome Endopeptidase Complex / Q000187:drug effects*; D066244:Proton Magnetic Resonance Spectroscopy; D021241:Spectrometry, Mass, Electrospray Ionization
D004791:Enzyme Inhibitors; D055666:Lipopeptides; C513652:fellutamide B; D046988:Proteasome Endopeptidase Complex