Characterization of the ARF7/ARF19 mediated signal cascade regulating root hair growth.

Two kinases and one kinase-interacting protein were identified using 2 microarray datasets of root hair mutants coupled to a comprehensive reverse genetics approach. The project aims to reveal how they are regulated by auxin and how they regulate tip-growth in root hairs. This can be summaried by the following objectives:1) The genes are auxin-regulated in an auxin response factor-dependent manner (ARF7/ARF19). Chromatin ImmunoPrecipitation (ChIP) followed by gene-specific PCRs will reveal whether they are direct or indirect targets of these ARFs (Dr. Hill, Nottingham Univ.) and qPCR will quantify their expression levels in several auxin-signaling mutants, further unraveling their auxin-regulation. 2) The involvement of the genes in NADPH oxidase-dependent, auxin-regulated ROS accumulation in root hairs will be monitored. This study involves molecular biological techniques and different forms of microscopy. 3) With the use of ion-specific vibrating probes and ion-sensitive dyes coupled to ratio-imaging, the effect of the gene knock-outs on extracellular ion-fluxes and intracellular ion-gradients at the growing tip root hairs - a conditio sine qua non for sustaining tip-growth - will be quantified (Prof. Feijó, Lisbon Univ.).4) The interaction partners/targets of the proteins will be identified using tandem affinity purification (Dr. De Jaeger, Gent), followed by kinase assays. T-DNA insertion lines for the identified interaction partners will be screened for root hair (and pollen tube growth) phenotypes.Together, these objectives will clarify how auxin regulates these genes and how these genes, in turn, regulate tipgrowth in root hairs (and pollen tubes)/

Keywords:PHYSIOLOGY (PLANT), ARABIDOPSIS THALIANA L.

Disciplines:General biology, Plant biology