Fig 3. Transcriptional and translational activities of Hm1^A during the seedling and mature stages.

(A) Reverse transcription (RT)-PCR assay showing no change in Hm1^A accumulation in leaves from week-1 through week-8 after planting. The actin gene was used as a control. (B) Quantitative real time PCR (qRT-PCR) measurements of the expression of Hm1^A also demonstrates no change in Hm1^A accumulation across the time period when APR is established. (C) Western blots showing that the level or stability of the HM1^A protein do not change over time during plant development. Equal amounts of protein were loaded following quantification with the Bradford method. (D) In vitro HC-toxin reductase (HCTR) assays showing that the relative enzymatic activity encoded by Hm1^A is less than Hm1^B73 but higher than hm1^Pr, the null allele. The specific activity of HCTR varies between alleles but not over time between weeks-3 and -7 in any genotype. The HCTR assay was based on the determination via LC-MS/MS of the amount of HC-toxin reduced by leaf protein extracts from the leaves of all genotypes. Different letters indicate significant differences between genotypes (p_adj < 0.05).