02854nas a2200373 4500008004100000022001400041245012300055210006900178260001600247300001100263490000800274520174600282653001602028653002502044653001802069653002302087653002802110653001902138653001602157653001702173100002502190700001802215700001802233700001902251700001602270700002102286700002102307700001802328700002202346700002502368700001802393700002002411856004902431 2015 eng d a1477-912900aThe cyclophilin A DIAGEOTROPICA gene affects auxin transport in both root and shoot to control lateral root formation.0 acyclophilin A DIAGEOTROPICA gene affects auxin transport in both c2015 Feb 15 a712-210 v1423 a

Cyclophilin A is a conserved peptidyl-prolyl cis-trans isomerase (PPIase) best known as the cellular receptor of the immunosuppressant cyclosporine A. Despite significant effort, evidence of developmental functions of cyclophilin A in non-plant systems has remained obscure. Mutations in a tomato (Solanum lycopersicum) cyclophilin A ortholog, DIAGEOTROPICA (DGT), have been shown to abolish the organogenesis of lateral roots; however, a mechanistic explanation of the phenotype is lacking. Here, we show that the dgt mutant lacks auxin maxima relevant to priming and specification of lateral root founder cells. DGT is expressed in shoot and root, and localizes to both the nucleus and cytoplasm during lateral root organogenesis. Mutation of ENTIRE/IAA9, a member of the auxin-responsive Aux/IAA protein family of transcriptional repressors, partially restores the inability of dgt to initiate lateral root primordia but not the primordia outgrowth. By comparison, grafting of a wild-type scion restores the process of lateral root formation, consistent with participation of a mobile signal. Antibodies do not detect movement of the DGT protein into the dgt rootstock; however, experiments with radiolabeled auxin and an auxin-specific microelectrode demonstrate abnormal auxin fluxes. Functional studies of DGT in heterologous yeast and tobacco-leaf auxin-transport systems demonstrate that DGT negatively regulates PIN-FORMED (PIN) auxin efflux transporters by affecting their plasma membrane localization. Studies in tomato support complex effects of the dgt mutation on PIN expression level, expression domain and plasma membrane localization. Our data demonstrate that DGT regulates auxin transport in lateral root formation.

10aArabidopsis10aBiological Transport10aCyclophilin A10aIndoleacetic Acids10aLycopersicon esculentum10aPlant Proteins10aPlant Roots10aPlant Shoots1 aIvanchenko, Maria, G1 aZhu, Jinsheng1 aWang, Bangjun1 aMedvecká, Eva1 aDu, Yunlong1 aAzzarello, Elisa1 aMancuso, Stefano1 aMegraw, Molly1 aFilichkin, Sergei1 aDubrovsky, Joseph, G1 aFriml, Jiří1 aGeisler, Markus uhttps://megraw.cgrb.oregonstate.edu/node/31602178nas a2200277 4500008004100000022001400041245011900055210006900174260000900243300000800252490000700260520133000267653001601597653001701613653001801630653001901648653001601667653003001683653002701713653001301740653003401753100002101787700002501808700001801833856004901851 2015 eng d a1471-216400aNanoCAGE-XL and CapFilter: an approach to genome wide identification of high confidence transcription start sites.0 aNanoCAGEXL and CapFilter an approach to genome wide identificati c2015 a5970 v163 a

BACKGROUND: Identifying the transcription start sites (TSS) of genes is essential for characterizing promoter regions. Several protocols have been developed to capture the 5' end of transcripts via Cap Analysis of Gene Expression (CAGE) or linker-ligation strategies such as Paired-End Analysis of Transcription Start Sites (PEAT), but often require large amounts of tissue. More recently, nanoCAGE was developed for sequencing on the Illumina GAIIx to overcome these difficulties.

RESULTS: Here we present the first publicly available adaptation of nanoCAGE for sequencing on recent ultra-high throughput platforms such as Illumina HiSeq-2000, and CapFilter, a computational pipeline that greatly increases confidence in TSS identification. We report excellent gene coverage, reproducibility, and precision in transcription start site discovery for samples from Arabidopsis thaliana roots.

CONCLUSION: nanoCAGE-XL together with CapFilter allows for genome wide identification of high confidence transcription start sites in large eukaryotic genomes.

[Link to Protocol, Additional Data, and Supplementary Materials]

[Link to CapFilter Software]

10aArabidopsis10aGenes, Plant10aGenome, Plant10aNanotechnology10aPlant Roots10aPromoter Regions, Genetic10aSequence Analysis, DNA10aSoftware10aTranscription Initiation Site1 aCumbie, Jason, S1 aIvanchenko, Maria, G1 aMegraw, Molly uhttps://megraw.cgrb.oregonstate.edu/node/311