Indiana University-Purdue University Indianapolis (IUPUI) / The root system architecture of higher plants plays an essential role in the uptake of water and nutrients as well as the production of hormones. These root systems are highly branched with the formation of post-embryonic organs such as lateral roots. The initiation and development of lateral roots has been well defined. WAG1 and WAG2 are protein-serine/threonine kinases from Arabidopsis that are closely related to PINOID and suppress root waving. The wag1;wag2 double mutants exhibit a strong root waving phenotype on vertical hard agar plates only seen in wild-type roots when the seedlings are grown on inclined plates. Here an additional root phenotype in the wag1;wag2 mutant is reported. The wag1;wag2 double mutant displays both an increased total number and density of emerged lateral roots (approximately 1.5-fold). An increased LRP density of 1.5-fold over wild-type is observed. To ascertain the role of WAG1 and WAG2 in lateral root development we examined promoter activity in the WAG1::GUS and WAG2::GUS lines. The WAG1 promoter showed no detectable activity at any stage of development. The WAG2 promoter was active in stage IV onward, however there was no detectable activity in the cell types associated with initiation events. The lateral root density and spatial patterning in wild-type, when grown on inclined hard agar plates, was similar to wag1;wag2 on vertical plates. Seedlings of both genotypes were treated with hormones such as auxin and MeJA, and inhibitors. Auxin response in wag1;wag2 was normal with a similar number of LR as the wild-type after treatment. Treatment with MeJA resulted in a similar induction of LRP in both genotypes, however the percent lateral root emergence in wag1;wag2 was reduced while Col-0 was increased compared to controls. Treatment with the calcium blocker tetracaine resulted in wag1;wag2 displaying a wild-type level of LR but had no significant effect on wild-type. Genetic analysis of the wag1;wag2 LR pathway revealed that WAG1 and WAG2 are acting in the same pathway as AUX1, AXR1and PGM1. pgm1-1 was not previously reported to have a LR defect but showed decreased LR formation here, while pgm1;wag1;wag2 had a similar LR density to wag1;wag2. TIR7 and ARG1 were both deduced to operate in separate pathways from WAG1 and WAG2. The data presented here shows that the wag1;wag2 double mutant has an increased number of LR compared to Col-0. This defect appears to be caused by increased pre-initiation events and seems to be tied to the root waving phenotype. However, the treatment with MeJA revealed a possible role for WAG1 or WAG2 in LRP development, potentially under stress conditions. Calcium also seems to play a significant role in the wag1;wag2 LR phenotype, possibly independent of the root waving phenotype.