Microarray transcriptomic profiling of Arabidopsis root stem cells within the BRL3 expression domain treated with Brassinolide (BL) [microarray]

Brassinosteroids (BRs) are steroid hormones involved in multiple processes of plant growth and development, and the adaptation to the environment. Some of the processes regulated by BRs are meristem activity and stem cell divisions. At the core of the root stem cell niche, it is placed the quiescent center (QC), that act as a cell reservoir. QC cells only trigger their divisions when need to replenish the stem cells, for example, after a DNA damage. BR signaling is in charge of triggering these QC divisions, but the exact mechanisms of how this process is regulated is still unknown. Here, we use an interdisciplinary approach, using Arabidopsis thaliana as a model system, including molecular genetics, physiology and bioinformatics to decipher the role of BR receptors upon DNA damage regulating the QC divisions. The results uncover novel roles for the BR-receptor kinase BRL3 (BRI1-like 3) receptor in DNA damage response (DDR) in plants, by modulating the DNA repair and the cell-cycle progression. We identified candidate tissue-specific transcriptional regulator, specifically expressed in the QC cells, the RNR2A (RIBONUCLEOTIDE REDUCTASE 2A), in charge of maintaining dNTPs (deoxynucleotide triphosphates) supply during DNA synthesis that is modulated by BRL3 downstream signaling events. Considering the importance of plant stem cells and their tissues for biomass accumulation and constantly exposed to adverse environmental stresses that can cause DNA damage or cell-cycle arrest in the RAM, here we stablished the mechanism linking root meristematic activity to the DDR through the cell-specific steroid receptor kinase BRL3. One factor experimental design (BL-treated vs. Control). Primary root tips of 6-day-old transgenic Arabidopsis seedlings expressing the reporter pBRL3:nlsYFP were treated with 10 nM of BL for 2h (in liquid media). Then the roots were submitted to protoplasting and the cells expressing YFP (BRL3 expression domain) selected using Fluorescence- Activated Cell Sorting (FACS). mRNA from selected cells in BL-treated or control conditions was extracted, converted to cDNA and hybridized to Affymetrix ATH1 GeneChip.