Isolating Genetic Regulators of Root System Architecture

Lateral root development is influenced by the environment in which a plant is grown. For example, wildtype Columbia plants produce root systems of vastly different sizes when grown on media containing different amounts of osmolytes simulating different levels of water availability (Deak and Malamy, 2005). Our studies have shown that osmotica have little effect on the intiation of lateral root primordia but strongly repress the development of these primordia into lateral roots.

In order to identify genes responsible for osmotic regulation of lateral root development, we conducted a forward genetic screen. This screen identified many recessive mutations which confer a more highly branched root system to plants. We have further characterized two of these mutants, named lateral root development1 and 2 (lrd1 & lrd2). Both mutants show increased levels of lateral root initiation and primordia development into lateral roots. Interestingly, these mutations separate intrinsic plant developmental pathways from pathways that regulate environmental response. lrd1 plants have more highly branched root systems under all environmental conditions. However, lrd1 plants remain responsive to environmental cues, and repress branching to a similar extent as do wildtype plants in response to osmotica. This shows that LRD1 is an intrinsic regulator of root system growth and does not participate in the plant's response to its environment. lrd2 plants also show increased branching under all of the conditions we have tested. However, these plants do not repress branching in an environmentally responsive way. Therefore, we conclude that LRD2 is a critical component of both the mechanism that represses lateral root formation in response to the environment and of the mechanism that constrains lateral root formation during normal growth and development.

Ongoing work in the lab is aimed at the identification of the LRD genes. Mapping experiments have localized the lesions to chromosome 4 (lrd1) and chromosome 1 (lrd2). Transgenic complementation experiments are in progress to confirm the identity of each gene.

NRT2.1: A Regulator of Environmental Response

Developmental plasticity is believed to be a crucial mechinism by which plants cope with the constraints of sessile life. Thoughout its adult life, a plant is continuously undergoing a process of sensing its environment, integrating the information it recieves into developmental decisions, and enacting these decisions. Through this process, a plant can regulate the timing, placement, and extent of growth of its new organs - attributes that define the plant's architecture. It is predicted that this architecture in turn delineates the physiological requirements and potentials of the plant. Thus this proccess of sensing, integration, and enactment allows the plant to optimize its morphology, and perhaps its physiology, to its environment.

This process can be illustrated in the regulation of the Arabisopsis root system architecture in response to nitrate (NO3). On our high sucrose media, wildtype (Col) seedlings grown under nitrate plentiful conditions develop a root system architecture characterized by a single primary root and a large number of lateral roots. In contrast, wildtype seedlings grown under limiting NO3 conditions exhibit a repression of primary root growth and significantly sparser lateral root formations. Examining the roots under a microscope, we find that this repression of lateral root formation is occuring at or before the initial cell divisions that indicate lateral root initiation. Using this assay as a model for developmental plasticity, we have taken a forward genetics approach to indentify regulators of this pathway.

lateral root initiation 1 (lin1) is one of the mutants isolated for its lack of repression of lateral root formation under limited NO3 conditions. While lin1 seedlings initiate more lateral roots than wildtype under limiting NO3 conditions, no significant difference is seen under NO3 plentiful conditions. This indicates that the LIN1 regulator participates specifically in nitrate response and is not an intrinsic growth regulator. We have shown that lin1 carries a missense mutation in the NRT2.1 gene. NRT2.1 encodes a putative high-affinity nitrate transporter that functions at low external nitrate concentrations. Direct measurement of nitrate uptake and nitrate content in the lin1 mutant seedlings established that both are indeed reduced. Because repression of lateral root initiation in WT plants can be relieved by increased concentrations of external nitrate, it is surprising to find that repression is also relieved by a defect in a component of the high-affinity nitrate uptake system. Furthermore, lateral root initiation is increased in lin1 relative to WT even when seedlings are grown on nitrate-free media, demonstrating that the mutant phenotype is nitrate-independent. These results indicate that NRT2.1 is a repressor of lateral root initiation and that this role is independent of the protein's role in nitrate uptake. We propose that Arabidopsis NRT2.1 acts either as a nitrate sensor or signal transducer to coordinate the development of the root system with nutritional cues.