Illumination is necessary and sufficient to induce histone acetylation independent of transcriptional activity at the C4-specific phosphoenolpyruvate carboxylase promoter in maize.

Expression of the C4-specific phosphoenolpyruvate carboxylase (C4-PEPC) gene in maize (Zea mays) is regulated in a tissue-specific manner, but affected by light and nutrient availability. We manipulated these stimuli in a combinatorial manner and analyzed concomitant changes in histone acetylation of the nucleosomes associated with the C4-PEPC gene in relation to transcriptional activity and steady-state mRNA levels. Whereas the transition from the lowest activity to an intermediate activity was observed in the absence of histone acetylation, the light-induced boost to full activity was associated with strong enhancement of the acetylation of both histones H3 and H4 limited to the gene region. Once activated by light, prolonged darkness was necessary to reduce both transcription and, in parallel, histone acetylation. Unexpectedly, histone acetylation was also induced in bundle sheath cells, although the transcriptional activity did not respond to illumination in this tissue. Furthermore, we were able to down-regulate the promoter by nitrogen depletion in the light without any decrease in the hyperacetylation of histone H4. When plants kept in prolonged darkness were nitrogen depleted and then exposed to light, transcription was not induced, but the promoter chromatin became hyperacetylated. We suggest a model where inhibition of a histone deacetylase in the light triggers H4 hyperacetylation at the C4-PEPC gene promoter regardless of the transcriptional activity of the gene. Our data indicate that an understanding of the interplay between histone modification and transcription requires analysis of signal integration on promoters in vivo.

Restriction accessibility in isolated nuclei reveals light-induced chromatin reorganization at the PEPC promoter in maize.

Expression of genes necessary to perform C4 photosynthesis in maize is activated by light. It is not known how this activation is regulated on the chromatin level in vivo. We analysed alterations in the chromatin structure of the promoter of the C4-specific isoform of phosphoenolpyruvate carboxylase (PEPC) after illumination of seedlings. A protocol was established that facilitates the preparation of nuclei from maize leaves with intact chromatin structure and resistance to DNA degradation during prolonged incubation at high temperatures. The presence of non-spliced transcripts from the C4-PEPC gene in the nuclei was demonstrated by RT-PCR. The chromatin was partially digested with restriction endonucleases. Quantitative PCR analyses revealed a clear increase in the accessibility of the promoter chromatin to restriction dependent on illumination of the seedlings. The data indicate chromatin reorganization at the C4-PEPC promoter during activation.