Phenotypic responses to nitrate availability in medicago truncatula under a range of nodulation conditions

Abstract

Nitrogen (N) is a major element required for plant growth. Plants obtain N mainly as inorganic N forms such as nitrate and ammonia from soil and show plasticity in root and shoot phenotypes in response to N availability in the soil to minimise carbon (C) costs and maximise N capture. Legumes are able source N from N2 fixation through symbiotic relationship with soil N2-fixing bacteria, collectively known as rhizobia. During this symbiosis, legumes supply C to rhizobia housed in nodules, in return for N obtained through N2 fixation in nodules. These additional C sinks and N sources are expected to change the phenotypic responses of nodulated legumes to external N. This thesis examined the influence of nodulation on the phenotypic responses of legumes to N availability. It was hypothesised that the presence of N2-fixing nodules would alleviate the plasticity responses to an external N gradient while reducing the available C to invest into root branching or shoot growth. To test these hypotheses, nodule numbers were altered in three ways: 1) by comparing three legume species differing in nodulation 2) by using plant hypernodulation mutants of the model legume Medicago truncatula and 3) by nodulating M. truncatula with different strains of rhizobia Plants were grown in the glasshouse for four weeks post-germination and treated with four nitrate concentrations, 0, 0.1, 2 and 10 mM, in the absence or presence of rhizobia. Shoot to root (S: R) biomass ratio, root length and lateral root density, nodulation, N2 fixation and N content were assessed. Generally, uninoculated M. truncatula (barrel medic), M. sativa (alfalfa) and Trifolium subterraneum (subclover) responded to increasing nitrate availability by increasing S: R biomass ratio, total root length and lateral root density, similar to non-legumes. Hypernodulation genes were found to partially regulate the S: R biomass allocation in response to external N in the absence of rhizobia, suggesting more general roles of these genes in legumes in C: N allocation than just for nodule number control. The presence of nodules in M. truncatula altered both lateral root density and root length in response to nitrate availability. Nodule density showed an inverse relationship with lateral root density and root length, which suggest that plants regulate C resource allocation within the root system to accommodate the extra C sink to nodules. Rhizobial strains that efficiently fixed N2 for its host, maintained an inverse relationship between nodule density and lateral root density and root length. However, hypernodulation mutants, lss (Like sunn Supernodulator), sunn-1 (Super Numeric Nodules), sunn-4 and rdn-1 (Root Determined Nodulation) were defective in regulating lateral root density and length when nodulated. The S: R biomass allocation response to nitrate availability in inoculated legumes was dependent on the regulation by hypernodulation genes and the N2 fixing efficiency of rhizobial strains. The hypernodulation mutants were unable to increase shoot dry biomass even at high nitrate concentrations, likely due to the excessive number of nodules. The S: R biomass allocation response to nitrate remained unchanged in plants inoculated with highly efficient N2-fixing rhizobia.