Short-interval traffic lines: versatile tools for genetic analysis in Arabidopsis thaliana.

Traffic lines are transgenic stocks of Arabidopsis thaliana that contain a pair of linked seed-specific eGFP and DsRed markers. These stocks were originally developed for the purpose of studying recombination, but can also be used to follow the inheritance of unmarked chromosomes placed in trans to the marked chromosome. They are particularly useful for this latter purpose if the distance between markers is short, making double recombination within this interval relatively rare. We generated 163 traffic lines that cover the Arabidopsis genome in overlapping intervals of approximately 1.2 Mb (6.9 cM). These stocks make it possible to predict the genotype of a plant based on its seed fluorescence (or lack thereof) and facilitate many experiments in genetic analysis that are difficult, tedious, or expensive to perform using current techniques. Here, we show how these lines enable a phenotypic analysis of alleles with weak or variable phenotypes, genetic mapping of novel mutations, introducing transgenes into a lethal or sterile genetic background, and separating closely linked mutations.

How was apical growth regulated in the ancestral land plant? Insights from the development of non-seed plants.

Land plant life cycles are separated into distinct haploid gametophyte and diploid sporophyte stages. Indeterminate apical growth evolved independently in bryophyte (moss, liverwort, and hornwort) and fern gametophytes, and tracheophyte (vascular plant) sporophytes. The extent to which apical growth in tracheophytes co-opted conserved gametophytic gene networks, or exploited ancestral sporophytic networks, is a long-standing question in plant evolution. The recent phylogenetic confirmation of bryophytes and tracheophytes as sister groups has led to a reassessment of the nature of the ancestral land plant. Here, we review developmental genetic studies of apical regulators and speculate on their likely evolutionary history.

VAL genes regulate vegetative phase change via miR156-dependent and independent mechanisms.

How organisms control when to transition between different stages of development is a key question in biology. In plants, epigenetic silencing by Polycomb repressive complex 1 (PRC1) and PRC2 plays a crucial role in promoting developmental transitions, including from juvenile-to-adult phases of vegetative growth. PRC1/2 are known to repress the master regulator of vegetative phase change, miR156, leading to the transition to adult growth, but how this process is regulated temporally is unknown. Here we investigate whether transcription factors in the VIVIPAROUS/ABI3-LIKE (VAL) gene family provide the temporal signal for the epigenetic repression of miR156. Exploiting a novel val1 allele, we found that VAL1 and VAL2 redundantly regulate vegetative phase change by controlling the overall level, rather than temporal dynamics, of miR156 expression. Furthermore, we discovered that VAL1 and VAL2 also act independently of miR156 to control this important developmental transition. In combination, our results highlight the complexity of temporal regulation in plants.

Lonely at the top? Regulation of shoot apical meristem activity by intrinsic and extrinsic factors.

All the above-ground organs of a plant are derived from stem cells that reside in shoot apical meristems (SAM). Over the past 25 years, the genetic pathways that control the proliferation of stem cells within the SAM, and the differentiation of their progenitors into lateral organs, have been described in great detail. However, longstanding questions regarding the importance of communication between cells within the SAM and lateral organs have, until recently, remained unanswered. In this review, we describe recent investigations into the extent, nature and significance of signaling both to and from the SAM.

ALTERED MERISTEM PROGRAM1 regulates leaf identity independently of miR156-mediated translational repression.

In Arabidopsis, loss of the carboxypeptidase ALTERED MERISTEM PROGRAM1 (AMP1) produces an increase in the rate of leaf initiation, an enlarged shoot apical meristem and an increase in the number of juvenile leaves. This phenotype is also observed in plants with reduced levels of miR156-targeted SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) transcription factors, suggesting that AMP1 might promote SPL activity. However, we found that the amp1 mutant phenotype is only partially corrected by elevated SPL gene expression, and that amp1 has no significant effect on SPL transcript levels, or on the level or the activity of miR156. Although AMP1 has been reported to promote miRNA-mediated translational repression, amp1 did not prevent the translational repression of the miR156 target SPL9 or the miR159 target MYB33. These results suggest that AMP1 regulates vegetative phase change downstream of, or in parallel to, the miR156/SPL pathway, and that it is not universally required for miRNA-mediated translational repression.

Role for the shoot apical meristem in the specification of juvenile leaf identity in Arabidopsis.

The extent to which the shoot apical meristem (SAM) controls developmental decisions, rather than interpreting them, is a longstanding issue in plant development. Previous work suggests that vegetative phase change is regulated by signals intrinsic and extrinsic to the SAM, but the relative importance of these signals for this process is unknown. We investigated this question by examining the effect of meristem-deficient mutations on vegetative phase change and on the expression of key regulators of this process, miR156 and its targets, SPL transcription factors. We found that the precocious phenotypes of meristem-deficient mutants are a consequence of reduced miR156 accumulation. Tissue-specific manipulation of miR156 levels revealed that the SAM functions as an essential pool of miR156 early in shoot development, but that its effect on leaf identity declines with age. We also found that SPL genes control meristem size by repressing WUSCHEL expression via a novel genetic pathway.

Candidate regulators of Early Leaf Development in Maize Perturb Hormone Signalling and Secondary Cell Wall Formation When Constitutively Expressed in Rice.

All grass leaves are strap-shaped with a series of parallel veins running from base to tip, but the distance between each pair of veins, and the cell-types that develop between them, differs depending on whether the plant performs C3 or C4 photosynthesis. As part of a multinational effort to introduce C4 traits into rice to boost crop yield, candidate regulators of C4 leaf anatomy were previously identified through an analysis of maize leaf transcriptomes. Here we tested the potential of 60 of those candidate genes to alter leaf anatomy in rice. In each case, transgenic rice lines were generated in which the maize gene was constitutively expressed. Lines grouped into three phenotypic classes: (1) indistinguishable from wild-type; (2) aberrant shoot and/or root growth indicating possible perturbations to hormone homeostasis; and (3) altered secondary cell wall formation. One of the genes in class 3 defines a novel monocot-specific family. None of the genes were individually sufficient to induce C4-like vein patterning or cell-type differentiation in rice. A better understanding of gene function in C4 plants is now needed to inform more sophisticated engineering attempts to alter leaf anatomy in C3 plants.

The role of small RNAs in vegetative shoot development.

Shoot development consists of the production of lateral organs in predictable spatial and temporal patterns at the shoot apex. To properly integrate such programs of growth across different cell and tissue types, plants require highly complex and robust genetic networks. Over the last twenty years, the roles of small, non-coding RNAs (sRNAs) in these networks have become increasingly apparent, not least in vegetative shoot growth. In this review, we describe recent progress in understanding the contribution of sRNAs to the regulation of vegetative shoot growth, and outline persisting experimental limitations in the field.

Cracking the Kranz enigma with systems biology.

Leaves with Kranz anatomy exhibit a highly characteristic arrangement of closely spaced veins surrounded by concentric wreaths of bundle sheath and mesophyll cells. This anatomical framework is vital for effective C4 photosynthesis in nearly all known land plant lineages and has evolved independently on over 60 occasions. Over the last 3 years, technological advances, particularly in high-throughput DNA sequencing, have allowed the development of Kranz anatomy to be interrogated at unprecedented depth. This review highlights the recent advances in our understanding that have been facilitated by systems biology approaches, and proposes a testable model for the regulation of Kranz development.

Genome-wide transcript analysis of early maize leaf development reveals gene cohorts associated with the differentiation of C4 Kranz anatomy.

Photosynthesis underpins the viability of most ecosystems, with C4 plants that exhibit 'Kranz' anatomy being the most efficient primary producers. Kranz anatomy is characterized by closely spaced veins that are encircled by two morphologically distinct photosynthetic cell types. Although Kranz anatomy evolved multiple times, the underlying genetic mechanisms remain largely elusive, with only the maize scarecrow gene so far implicated in Kranz patterning. To provide a broader insight into the regulation of Kranz differentiation, we performed a genome-wide comparative analysis of developmental trajectories in Kranz (foliar leaf blade) and non-Kranz (husk leaf sheath) leaves of the C4 plant maize. Using profile classification of gene expression in early leaf primordia, we identified cohorts of genes associated with procambium initiation and vascular patterning. In addition, we used supervised classification criteria inferred from anatomical and developmental analyses of five developmental stages to identify candidate regulators of cell-type specification. Our analysis supports the suggestion that Kranz anatomy is patterned, at least in part, by a SCARECROW/SHORTROOT regulatory network, and suggests likely components of that network. Furthermore, the data imply a role for additional pathways in the development of Kranz leaves.