Sumoylation of the Plant Clock Transcription Factor CCA1 Suppresses DNA Binding

Abstract

In plants, the circadian clock regulates the expression of one-third of all transcripts and is crucial to virtually every aspect of metabolism and growth. We now establish sumoylation, a posttranslational protein modification, as a novel regulator of the key clock protein CCA1 in the model plant Arabidopsis. Dynamic sumoylation of CCA1 is observed in planta and confirmed in a heterologous expression system. To characterize how sumoylation might affect the activity of CCA1, we investigated the properties of CCA1 in a wild-type plant background in comparison with ots1 ots2, a mutant background showing increased overall levels of sumoylation. Neither the localization nor the stability of CCA1 was significantly affected. However, binding of CCA1 to a target promoter was significantly reduced in chromatin-immunoprecipitation experiments. In vitro experiments using recombinant protein revealed that reduced affinity to the cognate promoter element is a direct consequence of sumoylation of CCA1 that does not require any other factors. Combined, these results suggest sumoylation as a mechanism that tunes the DNA binding activity of the central plant clock transcription factor CCA1.