Characterization of novel proteins affected by the o2 mutation and expressed during maize endosperm development

The effect of the o2 mutation on protein expression during grain development was examined by two-dimensional electrophoresis (2-D PAGE) in seven different pairs of near-isogenic maize lines. The aim was to identify a set of proteins that are consistently affected in mutants, and which could be the products of new genes that are direct or indirect targets of the transcriptional activator O2. The abundance of 36 polypeptides was found to be modified in the seven backgrounds. Seventeen polypeptides were present in greater amounts in wild types than in mutants, and most of these were affected early. The remaining polypeptides were expressed at higher levels in mutants than in the wild types and were generally affected later in development, suggesting that they might be products of indirect targets of O2. Products of known direct target genes such as zeins, b-32 protein and a pyruvate orthophospate dikinase were included in the first set of polypeptides. Microsequencing of internal stretches of 15 amino acids was performed for thirteen polypeptides and homologies with sequences stored in databases were found for nine of them. Enzymes belonging to various metabolic pathways were tentatively identified, most of which were not previously known to be affected by the o2 mutation. These results confirm that the O2 gene could act as a connecting regulatory gene for different pathways of grain metabolism.