The trypanocidal benzoxaborole AN7973 inhibits trypanosome mRNA processing
Fig 3
AN7973 affects mRNA processing.
A) Trypanosomes were incubated with AN7973 for different times (0–9 hours). RNA was subjected to denaturing agarose gel electrophoresis, blotted, and detected using radioactive probes. This panel shows Methylene blue-stained gel showing equal loading of total RNA. The three prominent bands are rRNA. B) As (A), blot probed with a [32P]-end-labelled probe complementary to the spliced leader. The signals representing the ~140nt SLRNA and mature mRNAs are indicated. The cluster of bands around 2 kb probably represent the abundant mRNAs encoding the Variant Surface Glycoprotein, alpha and beta tubulin (both ~1.9 kb including 100nt poly(A)), and EF-1 alpha (1.8 kb) [91]. C) The blot from (B) was stripped then incubated with a beta-tubulin probe. Shadows from the SL probe remain. The positions of monomer, dimer and multimers from the tubulin repeats are indicated. D) The effect of AN7973 is not due to inhibition of SLRNA synthesis. Trypanosomes were treated either with AN7973 for 1h, or with Actinomycin D for 30 min, or with AN7973 followed by Actinomycin D, as indicated above the lanes. The lower panel is the rRNA loading control.