Proteome analysis of differentially expressed nuclear proteins in the phyA phyB cry1 cry2 quadruple mutant of Arabidopsis thaliana.

Phytochromes phyA and phyB, and cryptochromes cry1 and cry2 are the fourth main photoreceptors that control growth and development in plants by light. These photoreceptors are present in the nucleus and affect the expression levels of transcription factors and nuclear regulatory proteins that interact with light regulated gene promoters. DIGE techniques (Differential In Gel Electrophoresis) provides information not only of the abundance of proteins but also possible postranslational modifications when stained with Pro-Q Diamond. The main purpose of the work is to identify new nuclear proteins regulated by these photoreceptors. We constructed a proteome profile from nuclear enrich fractions of phyA phyB cry1 cry2 quadruple mutant and WT light grown Arabidopsis seedlings. Nuclear protein enrich fractions were obtained by a new modified protocol (based on Folta and Kaufman, 2006 and Calikowski and Meier, 2006)that enriched four times in nuclear proteins. Statistical analysis showed 39 differentially expressed spots where 15 spots were over expressed and 24 spots underexpressed in the quadruple mutant (p< 0.05, FDR 0.05). To study whether any of these 39 proteins were phosphorylated, we analyzed when the intensity of the Cy3+ ProQ Diamond fluorescence signal compared to the Cy3 signal. We found 2 potentially phosphorylated spots. The identity of all these nuclear proteins will provide relevant information toward elucidation on photoreceptor signaling pathways and their regulatory mechanisms.