hy 19354  (MedChemExpress)

Journal: eLife

Article Title: A parasitic fungus employs mutated eIF4A to survive on rocaglate-synthesizing Aglaia plants

doi: 10.7554/eLife.81302

Figure Lengend Snippet: Summary of K d (µM) between eIF4A protein and RNAs. A fluorescence polarization assay between FAM-labeled RNA ([AG] 10 ) and the indicated recombinant proteins was conducted to measure K d in the presence of DMSO, RocA, or aglafoline. ND, not determined.

Article Snippet: Chemical compound, drug , Aglafoline , MedChemExpress , Cat. #:HY-19354 , .

Techniques: Fluorescence, Recombinant

Journal: eLife

Article Title: A parasitic fungus employs mutated eIF4A to survive on rocaglate-synthesizing Aglaia plants

doi: 10.7554/eLife.81302

Figure Lengend Snippet: ( A ) RocA-mediated translational repression recapitulated by an in vitro reconstitution system with human factors. Recombinant proteins of H. sapiens eIF4A1 WT or Phe163Gly were added to the reaction with RocA. Reporter mRNA with CAA repeats or polypurine motifs was translated in the reaction. The data are presented as the mean and s.d. values (n = 3). ( B ) Translation of complex-preformed mRNAs to test the RocA gain of function. Recombinant proteins of Ophiocordyceps sp. BRM1 eIF4A1 WT or the Gly172His mutant were preincubated with the reporter mRNA possessing polypurine motifs in the presence or absence of RocA. After removal of free RocA by gel filtration, the protein-mRNA complex was added to RRL to monitor protein synthesis. The data are presented as the mean and s.d. values (n = 3). ( C ) MA (M, log ratio; A, mean average) plot of the translation efficiency changes caused by 3 µM aglafoline treatment in C. orbiculare eIF4A WT conidia. Resistant and sensitive mRNAs (FDR < 0.05) are highlighted. ( D ) Cumulative distribution of the translation efficiency changes in aglafoline-sensitive mRNAs (defined in C ) in C. orbiculare eIF4A WT conidia treated with 0.3 or 3 µM aglafoline. ( E ) Cumulative distribution of the translation efficiency changes in aglafoline-sensitive mRNAs (defined in C ) induced by 3 µM aglafoline treatment in C. orbiculare eIF4A WT and eIF4A His153Gly conidia. ( F ) Cumulative distribution of the global translation alterations, which are footprint changes normalized to mitochondrial footprints, in aglafoline-sensitive mRNAs (defined in C ) induced by 3 µM aglafoline treatment in C. orbiculare eIF4A WT and eIF4A His153Gly conidia. ( G ) Box plot of the translation efficiency changes caused by 3 µM aglafoline treatment in conidia across mRNAs with or without an [A/G] 6 motif in the 5′ UTR. The p values in ( D–G ) were calculated by the Mann–Whitney U test. Figure 3—source data 1. Files for the primary data corresponding to . Figure 3—source data 2. Files for the primary data corresponding to .

Article Snippet: Chemical compound, drug , Aglafoline , MedChemExpress , Cat. #:HY-19354 , .

Techniques: In Vitro, Recombinant, Mutagenesis, Filtration, MANN-WHITNEY

Journal: eLife

Article Title: A parasitic fungus employs mutated eIF4A to survive on rocaglate-synthesizing Aglaia plants

doi: 10.7554/eLife.81302

Figure Lengend Snippet: ( A ) Schematics of eIF4A recombination in C. orbiculare. NPTII , neomycin phosphotransferase II. ( B ) PCR-based screening of the recombined strains. The primer sets used for screening are depicted in ( A ). ( C, D ) Colony formation of the indicated C. orbiculare strains cultured in PDA for 5 days ( C ). The measured colony diameters are shown in the box plot (n = 5) ( D ). ( E–G ) Fluorescence polarization assay for FAM-labeled RNA ([AG] 10 ) (10 nM). WT and mutated Ophiocordyceps sp. BRM1 eIF4A proteins were used. To measure ATP-independent RNA clamping induced by aglafoline (50 µM), ADP and Pi were included in the reaction. The data are presented as the mean and s.d. values (n = 3). ( H ) Summary of the K d values in ( E–G ) under treatment with aglafoline. The data are presented as the mean and s.d. values. Figure 3—figure supplement 1—source data 1. Files for the full and unedited gel images corresponding to . Figure 3—figure supplement 1—source data 2. Files for the primary data corresponding to . Figure 3—figure supplement 1—source data 3. Files for the primary data corresponding to .

Article Snippet: Chemical compound, drug , Aglafoline , MedChemExpress , Cat. #:HY-19354 , .

Techniques: Cell Culture, Fluorescence, Labeling

Journal: eLife

Article Title: A parasitic fungus employs mutated eIF4A to survive on rocaglate-synthesizing Aglaia plants

doi: 10.7554/eLife.81302

Figure Lengend Snippet: ( A ) GO term analysis of aglafoline-sensitive mRNAs (defined in ). GO terms associated with yeast homologs were analyzed by DAVID ( ; ). ( B ) MA plot of the translation efficiency changes induced by 3 µM aglafoline treatment in C. orbiculare eIF4A WT mycelia. Resistant and sensitive mRNAs (false discovery rate [FDR] < 0.05) are highlighted. ( C ) Cumulative distribution of the translation efficiency changes in aglafoline-sensitive mRNAs (defined in B ) in C. orbiculare eIF4A WT mycelia treated with 0.3 or 3 µM aglafoline. ( D ) Cumulative distribution of the translation efficiency changes in aglafoline-sensitive mRNAs (defined in B ) induced by 3 µM aglafoline treatment in C. orbiculare eIF4A WT and eIF4A His153Gly mycelia. ( E ) Cumulative distribution of the global translation alterations, which are footprint changes normalized to mitochondrial footprints, in aglafoline-sensitive mRNAs (defined in B ) induced by 3 µM aglafoline treatment in C. orbiculare eIF4A WT and eIF4A His153Gly conidia. ( F ) Box plot of translation efficiency changes caused by 3 µM aglafoline treatment in mycelia across mRNAs with or without an [A/G] 6 motif in the 5′ UTR. ( G ) Venn diagram of the overlap between aglafoline-sensitive mRNAs in conidia (defined in ) and mycelia (defined in B ). The p values in ( C–F ) were calculated by the Mann–Whitney U test.

Article Snippet: Chemical compound, drug , Aglafoline , MedChemExpress , Cat. #:HY-19354 , .

Techniques: MANN-WHITNEY

Journal: eLife

Article Title: A parasitic fungus employs mutated eIF4A to survive on rocaglate-synthesizing Aglaia plants

doi: 10.7554/eLife.81302

Figure Lengend Snippet: ( A ) Workflow for monitoring the biomass of C. orbiculare eIF4A WT or eIF4A His153Gly strains on cucumber leaves under treatment with aglafoline. ( B ) Comparison of in planta fungal biomass of C. orbiculare eIF4A WT or eIF4A His153Gly strains with or without treatment with 1 µM aglafoline. Relative expression levels of the C. orbiculare 60 S ribosomal protein L5 gene (GenBank: Cob_v012718) normalized to that of a cucumber cyclophilin gene (GenBank: AY942800.1) were determined by RT–qPCR at 3 dpi (n = 8). The relative fungal biomasses of C. orbiculare were normalized to those of eIF4A WT without aglafoline. Significance was calculated by Student’s t- test (two-tailed). Three independent experiments showed similar results. Figure 4—source data 1. Files for the primary data corresponding to .

Article Snippet: Chemical compound, drug , Aglafoline , MedChemExpress , Cat. #:HY-19354 , .

Techniques: Comparison, Expressing, Quantitative RT-PCR, Two Tailed Test

Journal: eLife

Article Title: A parasitic fungus employs mutated eIF4A to survive on rocaglate-synthesizing Aglaia plants

doi: 10.7554/eLife.81302

Figure Lengend Snippet: ( A ) C. sativus leaves were sprayed with DMSO or aglafoline (1 µM) in water and incubated for 3 days using the same method as C. orbiculare inoculation. Figure 4—figure supplement 1—source data 1. Files for the full and unedited pictures corresponding to .

Article Snippet: Chemical compound, drug , Aglafoline , MedChemExpress , Cat. #:HY-19354 , .

Techniques: Incubation

Journal: eLife

Article Title: A parasitic fungus employs mutated eIF4A to survive on rocaglate-synthesizing Aglaia plants

doi: 10.7554/eLife.81302

Figure Lengend Snippet:

Article Snippet: Chemical compound, drug , Aglafoline , MedChemExpress , Cat. #:HY-19354 , .

Techniques: Recombinant, Plasmid Preparation, Sequencing, Reverse Transcription, Luciferase, Pore Size, Membrane, Ligation, Software, Chromatography, High Performance Liquid Chromatography, Nucleic Acid Purification