Transparent soil for imaging the rhizosphere

Understanding of soil processes is essential for addressing the global issues of food security, disease transmission and climate change. However, techniques for observing soil biology are lacking. We present a heterogeneous, porous, transparent substrate for in situ 3D imaging of living plants and root-associated microorganisms using particles of the transparent polymer, Nafion, and a solution with matching optical properties. Minerals and fluorescent dyes were adsorbed onto the Nafion particles for nutrient supply and imaging of pore size and geometry. Plant growth in transparent soil was similar to that in soil. We imaged colonization of lettuce roots by the human bacterial pathogen Escherichia coli O157:H7 showing micro-colony development. Micro-colonies may contribute to bacterial survival in soil. Transparent soil has applications in root biology, crop genetics and soil microbiology

Modulation of Arabidopsis and monocot root architecture by CLAVATA3/EMBRYO SURROUNDING REGION 26 peptide

Plant roots are important for a wide range of processes, including nutrient and water uptake, anchoring and mechanical support, storage functions, and as the major interface with the soil environment. Several small signalling peptides and receptor kinases have been shown to affect primary root growth, but very little is known about their role in lateral root development. In this context, the CLE family, a group of small signalling peptides that has been shown to affect a wide range of developmental processes, were the focus of this study. Here, the expression pattern during lateral root initiation for several CLE family members is explored and to what extent CLE1, CLE4, CLE7, CLE26, and CLE27, which show specific expression patterns in the root, are involved in regulating root architecture in Arabidopsis thaliana is assessed. Using chemically synthesized peptide variants, it was found that CLE26 plays an important role in regulating A. thaliana root architecture and interacts with auxin signalling. In addition, through alanine scanning and in silico structural modelling, key residues in the CLE26 peptide sequence that affect its activity are pinpointed. Finally, some interesting similarities and differences regarding the role of CLE26 in regulating monocot root architecture are presented

Mutations in the Arabidopsis Peroxisomal ABC Transporter COMATOSE Allow Differentiation between Multiple Functions In Planta: Insights from an Allelic Series

COMATOSE (CTS), the Arabidopsis homologue of human Adrenoleukodystrophy protein (ALDP), is required for import of substrates for peroxisomal β-oxidation. A new allelic series and a homology model based on the bacterial ABC transporter, Sav1866, provide novel insights into structure-function relations of ABC subfamily D proteins. In contrast to ALDP, where the majority of mutations result in protein absence from the peroxisomal membrane, all CTS mutants produced stable protein. Mutation of conserved residues in the Walker A and B motifs in CTS nucleotide-binding domain (NBD) 1 resulted in a null phenotype but had little effect in NBD2, indicating that the NBDs are functionally distinct in vivo. Two alleles containing mutations in NBD1 outside the Walker motifs (E617K and C631Y) exhibited resistance to auxin precursors 2,4-dichlorophenoxybutyric acid (2,4-DB) and indole butyric acid (IBA) but were wild type in all other tests. The homology model predicted that the transmission interfaces are domain-swapped in CTS, and the differential effects of mutations in the conserved "EAA motif" of coupling helix 2 supported this prediction, consistent with distinct roles for each NBD. Our findings demonstrate that CTS functions can be separated by mutagenesis and the structural model provides a framework for interpretation of phenotypic data

A method to determine the displacement velocity field in the apical region of the Arabidopsis root

In angiosperms, growth of the root apex is determined by the quiescent centre. All tissues of the root proper and the root cap are derived from initial cells that surround this zone. The diversity of cell lineages originated from these initials suggests an interesting variation of the displacement velocity within the root apex. However, little is known about this variation, especially in the most apical region including the root cap. This paper shows a method of determination of velocity field for this region taking the Arabidopsis root apex as example. Assuming the symplastic growth without a rotation around the root axis, the method combines mathematical modelling and two types of empirical data: the published velocity profile along the root axis above the quiescent centre, and dimensions of cell packet originated from the initials of epidermis and lateral root cap. The velocities, calculated for points of the axial section, vary in length and direction. Their length increases with distance from the quiescent centre, in the root cap at least twice slower than in the root proper, if points at similar distance from the quiescent centre are compared. The vector orientation depends on the position of a calculation point, the widest range of angular changes, reaching almost 90, in the lateral root cap. It is demonstrated how the velocity field is related to both distribution of growth rates and growth-resulted deformation of the cell wall system. Also changes in the field due to cell pattern asymmetry and differences in slope of the velocity profile are modelled

Quantitative fluorescence microscopy: from art to science

A substantial number of elegant experimental approaches have been developed to image the distribution and dynamics of DNA, mRNA, proteins, organelles, metabolites, and ions in living plant cells. Although the human brain can rapidly assimilate visual information, particularly when presented as animations and movies, it is much more challenging to condense the phenomenal amount of data present in three-, four-, or even five-dimensional images into statistically useful measurements. This review explores a range of in vivo fluorescence imaging applications in plants, with particular emphasis on where quantitative techniques are beginning to emerge