Location: Plant, Soil and Nutrition Research
Project Number: 1907-21000-036-00
Start Date: Jun 28, 2013
End Date: Jun 27, 2018
1) Study the role of sorghum AlMBP in regulating aluminum (Al) activated citrate transport via the sorghum Al tolerance protein, SbMATE. Will use a combination of ESI-Q-TOF MS/ ion mobility spectrometry and metal-ion chromatography to determine kinetics and specificity of Al binding by AlMBP. 2) Determine if Al binding by AlMBP causes this protein to disassociate from SbMATE using in vitro pull down assays, in vivo BiFC assays, and chemical cross-linking followed by LC-MS/MS analysis. 3) Determine the functional role of the SbMBP-SbMATE interaction by expressing both proteins in heterologous systems (oocytes and yeast) to determine if this confers Al activated of citrate exudation.4) Study the role of phosphorylation in regulation of SbMATE transport function via electrophysiological analysis of citrate efflux based on co-expression of SbMATE and candidate kinase proteins (CIPKs and calcineurin B-like [CBL] proteins) in oocytes.5) Investigate the role of protein structure in transport function for the plant MATE proteins that mediate citrate efflux and are involved in Al tolerance. Will determine the 3D crystal structure of SbMATE and use this structural model to direct functional analysis of SbMATE transport in oocytes. 6) After identifying altered SbMATE-type transporters that show enhanced function, the effects of these variants in plants will be determined by expressing SbMATE variants in transgenic Arabidopsis seedlings, and determining changes in Al tolerance. 7) In studies on rice Al tolerance, we will mine genome-wide association (GWA) data to identify/test candidate rice Al tolerance genes by a combination of high resolution mapping, molecular analysis in rice, expression of candidate Al tolerance genes in transgenic rice, and functional analysis of candidate transporter genes such as the Nrat1 Al transporter in heterologous systems (oocytes and yeast). 8) For research on root system architecture, we will mine data from joint linkage-GWA analysis on rice RSA traits to identify regions of the rice genome controlling root traits that play a role in nutrient acquisition (P, water and N) under limiting conditions. This will involve a combination of fine scale mapping, mRNA seq analysis of candidate genes, expression of candidate RSA trait genes in transgenic rice, and the verification of functionality of different root architectures by looking at performance in soil under limiting (low water, N or P) conditions.