Meristem function in plants requires both the maintenance of stem cells and the specification of founder cells from which lateral organs arise. Lateral organs are patterned along proximodistal, dorsoventral and mediolateral axes. Here we show that the Arabidopsis mutant asymmetric leaves1 (as1) disrupts this process. AS1 encodes a myb domain protein, closely related to PHANTASTICA in Antirrhinum and ROUGH SHEATH2 in maize, both of which negatively regulate knotted-class homeobox genes. AS1 negatively regulates the homeobox genes KNAT1 and KNAT2 and is, in turn, negatively regulated by the meristematic homeobox gene SHOOT MERISTEMLESS. This genetic pathway defines a mechanism for differentiating between stem cells and organ founder cells within the shoot apical meristem and demonstrates that genes expressed in organ primordia interact with meristematic genes to regulate shoot morphogenesis.

Leaves are determinate organs that arise from the flanks of the shoot apical meristem as polar structures with distinct adaxial (dorsal) and abaxial (ventral) sides. Opposing regulatory interactions between genes specifying adaxial or abaxial fates function to maintain dorsoventral polarity. One component of this regulatory network is the Myb-domain transcription factor gene ASYMMETRIC LEAVES1 (AS1). The contribution of AS1 to leaf polarity varies across different plant species; however, in Arabidopsis, as1 mutants have only mild defects in leaf polarity, suggesting that alternate pathways exist for leaf patterning. Here, we describe three genes, PIGGYBACK1 (PGY1), PGY2 and PGY3, which alter leaf patterning in the absence of AS1. All three pgy mutants develop dramatic ectopic lamina outgrowths on the adaxial side of the leaf in an as1 mutant background. This leaf-patterning defect is enhanced by mutations in the adaxial HD-ZIPIII gene REVOLUTA (REV), and is suppressed by mutations in abaxial KANADI genes. Thus, PGY genes influence leaf development via genetic interactions with the HD-ZIPIII-KANADI pathway. PGY1, PGY2 and PGY3 encode cytoplasmic large subunit ribosomal proteins, L10a, L9 and L5, respectively. Our results suggest a role for translation in leaf dorsoventral patterning and indicate that ribosomes are regulators of key patterning events in plant development. Development 135, 1315Development 135, -1324Development 135, (2008 DEVELOPMENT 1316 from the Arabidopsis Biological Resource Centre (ABRC). kan1-2 and kan2-1 were obtained from John Bowman. All genetic interactions were in a Ler background. Plants were grown either in soil or on Murashige and Skoog media at 22°C with a day length of 16 hours. KEY WORDS: Ribosomal protein, Leaf polarity, ASYMMETRIC LEAVES1, PIGGYBACK, Arabidopsis Geneticspgy genes were cloned using Ler ϫ Columbia F2 mapping populations. For complementation a 2.1 kb genomic fragment encompassing At2g27530, a 5 kb genomic fragment encompassing At1g33140 and a 3.5 kb genomic fragment encompassing At3g25520 were cloned into the binary vector pMDC123 (Curtis and Grossniklaus, 2003) and transformed into pgy1-1/pgy1-1 as1/+, pgy2-1/pgy2-1 as1/+ and pgy3-1/pgy3-1 as1/+ plants, respectively, using standard agrobacterium-mediated transformation (Clough and Bent, 1998). For each complementation construct, basta resistant plants with an as1 phenotype were confirmed as as1 pgy homozygotes.as1-1 rev-6 was analysed in the F3 generation of the cross as1-1 ϫ rev-6. In the F2 generation of this cross as1-1 rev-6 segregated at 1:15. pgy1-1 rev-6 were obtained from the F3 generation of the cross pgy1-1 ϫ rev-6. Progeny from pgy1-1 rev-6/+ individuals segregated 1:3 pgy1-1 rev-6 mutants. as1-1 pgy1-1 rev-6 triple mutants were analysed in the F4 generation of the cross as1-1 pgy1-1 ϫ as1-1 rev-6, after selfing as1-1 pgy1-1 rev-6/+ F3 plants. Segregation of as1-1 pgy1-1 rev-6 in this F4 generation was 1:3. as1-1 kan1-2 and pgy1-1 kan1-2 were obtained from the F3 generation of the respective crosses...

SummaryThe interphase nucleus exists as a highly dynamic system, the physical properties of which have functional importance in gene regulation. Not only can gene expression be influenced by the local sequence context, but also by the architecture of the nucleus in three-dimensions (3D), and by the interactions between these levels via chromatin modifications. A challenging task is to resolve the complex interplay between sequence-and genome structure-based control mechanisms. Here, we created a collection of 277 Arabidopsis lines that allow the visual tracking of individual loci in living plants while comparing gene expression potential at these locations, via an identical reporter cassette. Our studies revealed regional gene silencing near a heterochromatin island, via DNA methylation, that is correlated with mobility constraint and nucleolar association. We also found an example of nucleolar association that does not correlate with gene suppression, suggesting that distinct mechanisms exist that can mediate interactions between chromatin and the nucleolus. These studies demonstrate the utility of this novel resource in unifying structural and functional studies towards a more comprehensive model of how global chromatin organization may coordinate gene expression over large scales.

To identify genes involved in Arabidopsis thaliana petal and stamen organogenesis, we used a gene trap approach to examine the patterns of reporter expression at each stage of flower development of 1765 gene trap lines. In 80 lines, the reporter gene showed petal-and/or stamen-specific expression or lack of expression, or expression in distinct patterns within the petals and/or the stamens, including distinct suborgan domains of expression, such as tissue-specific lines marking epidermis and vasculature, as well as lines demarcating the proximodistal or abaxial/adaxial axes of the organs. Interestingly, reporter gene expression was typically restricted along the proximodistal axis of petals and stamens, indicating the importance of this developmental axis in patterning of gene expression domains in these organs. We identified novel domains of gene expression along the axis marking the midregion of the petals and apical and basal parts of the anthers. Most of the genes tagged in these 80 lines were identified, and their possible functions in petal and/or stamen differentiation are discussed. We also scored the floral phenotypes of the 1765 gene trap lines and recovered two mutants affecting previously uncharacterized genes. In addition to revealing common domains of gene expression, the gene trap lines reported here provide both useful markers and valuable starting points for reverse genetic analyses of the differentiation pathways in petal and stamen development.

Secreted and membrane-spanning proteins play fundamental roles in plant development but pose challenges for genetic identification and characterization. We describe a “secretion trap” screen for gene trap insertions in genes encoding proteins routed through the secretory pathway. The gene trap transposon encodes a β-glucuronidase reporter enzyme that is inhibited by N-linked glycosylation specific to the secretory pathway. Treatment of seedlings with tunicamycin inhibits glycosylation, resulting in increased activity of secreted β-glucuronidase fusions that result from gene trap integration downstream of exons encoding signal peptides. In the 2,059 gene trap lines that we screened, 32 secretion trap expression patterns were identified in a wide variety of tissues including embryos, meristems, and the developing vasculature. Genes disrupted by the secretion traps encode putative extracellular signaling proteins, membrane transport proteins, and novel secreted proteins of unknown function missed by conventional mutagenesis and gene prediction. Secretion traps provide a unique reagent for gene expression studies and can guide the genetic combination of loss of function alleles in related genes.