Skewing in Arabidopsis roots involves disparate environmental signaling pathways

Background: Skewing root patterns provide key insights into root growth strategies and mechanism that produce root architectures Roots exhibit skewing and waving when grown on a tilted, impenetrable surface, and while the genetics guiding these morphologies have been examined, the underlying molecular mechanisms of skewing and waving remain unclear. In this study, transcriptome data were derived from two Arabidopsis ecotypes, WS and Col-0, under three tilted growth conditions in order to identify candidate genes involved in skewing. WS is a skewing ecotype. Col-0 is a non-skewing ecotype. Results: This work identifies a number of genes that are likely involved in skewing, using growth conditions that differentially affect skewing and waving. Comparing the gene expression profiles of WS and Col-0 in different tilted growth conditions identified 11 candidate genes as potentially involved in the control of skewing. These 11 genes are involved in several different cellular processes, including sugar transport, salt signaling, cell wall organization, and hormone signaling. Conclusions: Many of the 11 identified genes are involved in signaling and perception, rather than the physical restructuring of roots, leading to the conclusion that root skewing is enabled through diverse environmental signaling pathways. These findings revealed further insights into the molecular mechanisms behind root skewing. This work investigated the transcriptional differences between skewing and non-skewing roots. Comparisons within WS revealed gees that that responded to the angle of growth (Agp) during the process of skewing. These genes were cross referenced with transcripts differing between the WS and Col-0 genotypes to refine the list of genes that are most probably be involved in root skewing. More of the highly probable skew gene candidates (HPSGC) are associated with environmental sensing (e.g. salt, sugar, hormones, darkness) than with physical growth differences (e.g. cell wall remodeling, cell division, cell elongation). Thus, the root behavior of skewing appears to be primarily driven by pathways that respond to disparate signals from the root local environment. Future studies could investigate the HPSGC to find the specific pathways and molecular mechanisms contributing to root skewing. Arabidopsis thaliana plants (wild type cultivars Col-0 and WS) were grown on media plates made from 0.5x MS liquid media, autoclaved with 0.5% Phytagel and poured in square-gridded plates (Fisherbrand, Fisher Scientific, Pittsburgh, PA). Seeds were wet sterilized in 1.7 mL Eppendorf microfuge tubes (Eppendorf, Hamburg, Germany) using a 5-minute 70% ethanol wash, followed by a 5 minute 50% v/v sodium hypochlorate solution wash (8.3%; Clorox, Oakland, CA), followed by 6 washes with sterile ddH2O. Seeds were planted on plates and moved to 4°C for two days, followed by three days of vertical growth (Agp 90°) in 19°C +/- 2°C, and 24-hour fluorescent light at approximately 80 μmol m-2 s-1 PAR. Plates were photographed, moved to their respective experimental condition (Agp 45°, 90°, or 135°), and photographed again on day 8. Plants were harvested and fixed in RNAlater (Ambion, Grand Island, New York, USA).