José Luis Riechmann

Highlights and prospects of potyvirus molecular biology

The potyvirus group [named after its type member, potato virus Y (PVY)] is the largest of the 34 plant virus groups and families currently recognized (Ward & Shukla, 1991). It contains at least 180 definitive and possible members (or 30% of all known plant viruses) which cause significant losses in agricultural, pasture, horticultural and ornamental crops (Ward & Shukla, 1991). These viruses are unique in the diversity of inclusion bodies that are formed during the infection cycle (see Lesemann, 1988). A feature shared by all potyviruses is the induction of characteristic pinwheel or scroll-shaped inclusion bodies in the cytoplasm of the infected cells (Edwardson, 1974). These cylindrical inclusion (CI) bodies are formed by a virus-encoded protein and can be considered as the most important phenotypic criterion for assigning viruses to the poty-virus group (Milne, 1988; Shukla et al., 1989; Ward & Shukla, 1991).

Orchestration of Floral Initiation by APETALA1

Flower Power The transcription factor APETALA1 (AP1) controls the transition from vegetative growth to flower production in the plant Arabidopsis. A handful of factors that control AP1 have been identified, as well as some targets that AP1 controls. Kaufmann et al. (p. 85) now apply genome-wide microarray analysis to identify over a thousand genes whose transcription is regulated by AP1. By proximity to AP1 binding sites, over two thousand genes are implicated as putative AP1 targets. Analysis of this network of interactions indicates that AP1 functions first to repress vegetative identity, then to help establish floral primordia, and finally to shape the differentiation of floral parts. The master transcription factor APETALA1 dynamically regulates a complex genetic network to guide flower development. The MADS-domain transcription factor APETALA1 (AP1) is a key regulator of Arabidopsis flower development. To understand the molecular mechanisms underlying AP1 function, we identified its target genes during floral initiation using a combination of gene expression profiling and genome-wide binding studies. Many of its targets encode transcriptional regulators, including known floral repressors. The latter genes are down-regulated by AP1, suggesting that it initiates floral development by abrogating the inhibitory effects of these genes. Although AP1 acts predominantly as a transcriptional repressor during the earliest stages of flower development, at more advanced stages it also activates regulatory genes required for floral organ formation, indicating a dynamic mode of action. Our results further imply that AP1 orchestrates floral initiation by integrating growth, patterning, and hormonal pathways.

Redundancy and specialization among plant microRNAs: role of the MIR164 family in developmental robustness

In plants, members of microRNA (miRNA) families are often predicted to target the same or overlapping sets of genes. It has thus been hypothesized that these miRNAs may act in a functionally redundant manner. This hypothesis is tested here by studying the effects of elimination of all three members of the MIR164 family from Arabidopsis. It was found that a loss of miR164 activity leads to a severe disruption of shoot development, in contrast to the effect of mutation in any single MIR164 gene. This indicates that these miRNAs are indeed functionally redundant. Differences in the expression patterns of the individual MIR164 genes imply, however, that redundancy among them is not complete, and that these miRNAs show functional specialization. Furthermore, the results of molecular and genetic analyses of miR164-mediated target regulation indicate that miR164 miRNAs function to control the transcript levels, as well as the expression patterns, of their targets, suggesting that they might contribute to developmental robustness. For two of the miR164 targets, namely CUP-SHAPED COTYLEDON1 (CUC1) and CUC2, we provide evidence for their involvement in the regulation of growth and show that their derepression in miR164 loss-of-function mutants is likely to account for most of the mutant phenotype.

Mapping the Core of the Arabidopsis Circadian Clock Defines the Network Structure of the Oscillator

Tic TOC1 Plant Clock The molecular clocks intertwined with cellular physiology regulate daily cycles. In plants, these circadian rhythms affect processes as diverse as carbon metabolism and leaf orientation. Huang et al. (p. 75, published online 8 March) have now analyzed the interactions driven by a key element of the circadian clock in plants, TOC1 (“TIMING OF CAB EXPRESSION 1”). TOC1 helps to coordinate responses of the morning and evening cycles, functioning to repress activity of other clock components. Morning and evening components of circadian rhythms are coordinated and stabilized by a repressor. In many organisms, the circadian clock is composed of functionally coupled morning and evening oscillators. In Arabidopsis, oscillator coupling relies on a core loop in which the evening oscillator component TIMING OF CAB EXPRESSION 1 (TOC1) was proposed to activate a subset of morning-expressed oscillator genes. Here, we show that TOC1 does not function as an activator but rather as a general repressor of oscillator gene expression. Repression occurs through TOC1 rhythmic association to the promoters of the oscillator genes. Hormone-dependent induction of TOC1 and analysis of RNA interference plants show that TOC1 prevents the activation of morning-expressed genes at night. Our study overturns the prevailing model of the Arabidopsis circadian clock, showing that the morning and evening oscillator loops are connected through the repressing activity of TOC1.

The homeotic protein AGAMOUS controls microsporogenesis by regulation of SPOROCYTELESS.

The Arabidopsis homeotic gene AGAMOUS (AG) is necessary for the specification of reproductive organs (stamens and carpels) during the early steps of flower development. AG encodes a transcription factor of the MADS-box family that is expressed in stamen and carpel primordia. At later stages of development, AG is expressed in distinct regions of the reproductive organs. This suggests that AG might function during the maturation of stamens and carpels, as well as in their early development. However, the developmental processes that AG might control during organogenesis and the genes that are regulated by this factor are largely unknown. Here we show that microsporogenesis, the process leading to pollen formation, is induced by AG through activation of the SPOROCYTELESS gene (SPL, also known as NOZZLE,NZZ), a regulator of sporogenesis. Furthermore, we demonstrate that SPL can induce microsporogenesis in the absence of AG function, suggesting that AG controls a specific process during organogenesis by activating another regulator that performs a subset of its functions.

Genome-Wide Analysis of Spatial Gene Expression in Arabidopsis Flowers

We have compared the gene expression profiles of inflorescences of the floral homeotic mutants apetala1, apetala2, apetala3, pistillata, and agamous with that of wild-type plants using a flower-specific cDNA microarray and a whole genome oligonucleotide array. By combining the data sets from the individual mutant/wild type comparisons, we were able to identify a large number of genes that are, within flowers, predicted to be specifically or at least predominantly expressed in one type of floral organ. We have analyzed the expression patterns of several of these genes by in situ hybridization and found that they match the predictions that were made based on the microarray experiments. Moreover, genes with known floral organ–specific expression patterns were correctly assigned by our analysis. The vast majority of the identified transcripts are found in stamens or carpels, whereas few genes are predicted to be expressed specifically or predominantly in sepals or petals. These findings indicate that spatially limited expression of a large number of genes is part of flower development and that its extent differs significantly between the reproductive organs and the organs of the perianth.

The Transcription Factor WIN1/SHN1 Regulates Cutin Biosynthesis in Arabidopsis thaliana

The composition and permeability of the cuticle has a large influence on its ability to protect the plant against various forms of biotic and abiotic stress. WAX INDUCER1 (WIN1) and related transcription factors have recently been shown to trigger wax production, enhance drought tolerance, and modulate cuticular permeability when overexpressed in Arabidopsis thaliana. We found that WIN1 influences the composition of cutin, a polyester that forms the backbone of the cuticle. WIN1 overexpression induces compositional changes and an overall increase in cutin production in vegetative and reproductive organs, while its downregulation has the opposite effect. Changes in cutin composition are preceded by the rapid and coordinated induction of several genes known or likely to be involved in cutin biosynthesis. This transcriptional response is followed after a delay by the induction of genes associated with wax biosynthesis, suggesting that the regulation of cutin and wax production by WIN1 is a two-step process. We demonstrate that at least one of the cutin pathway genes, which encodes long-chain acyl-CoA synthetase LACS2, is likely to be directly targeted by WIN1. Overall, our results suggest that WIN1 modulates cuticle permeability in Arabidopsis by regulating genes encoding cutin pathway enzymes.

Genome-wide analysis of gene expression during early Arabidopsis flower development.

Detailed information about stage-specific changes in gene expression is crucial for the understanding of the gene regulatory networks underlying development. Here, we describe the global gene expression dynamics during early flower development, a key process in the life cycle of a plant, during which floral patterning and the specification of floral organs is established. We used a novel floral induction system in Arabidopsis, which allows the isolation of a large number of synchronized floral buds, in conjunction with whole-genome microarray analysis to identify genes with differential expression at distinct stages of flower development. We found that the onset of flower formation is characterized by a massive downregulation of genes in incipient floral primordia, which is followed by a predominance of gene activation during the differentiation of floral organs. Among the genes we identified as differentially expressed in the experiment, we detected a significant enrichment of closely related members of gene families. The expression profiles of these related genes were often highly correlated, indicating similar temporal expression patterns. Moreover, we found that the majority of these genes is specifically up-regulated during certain developmental stages. Because co-expressed members of gene families in Arabidopsis frequently act in a redundant manner, these results suggest a high degree of functional redundancy during early flower development, but also that its extent may vary in a stage-specific manner.

Gene networks controlling the initiation of flower development

The onset of flower formation is a key regulatory event during the life cycle of angiosperm plants, which marks the beginning of the reproductive phase of development. It has been shown that floral initiation is under tight genetic control, and deciphering the underlying molecular mechanisms has been a main area of interest in plant biology for the past two decades. Here, we provide an overview of the developmental and genetic processes that occur during floral initiation. We further review recent studies that have led to the genome-wide identification of target genes of key floral regulators and discuss how they have contributed to an in-depth understanding of the gene regulatory networks controlling early flower development. We focus especially on a master regulator of floral initiation in Arabidopsis thaliana APETALA1 (AP1), but also outline what is known about the AP1 network in other plant species and the evolutionary implications.

3'-terminal sequence of the plum pox virus PS and ŏ6 isolates: evidence for RNA recombination within the potyvirus group.

The sequence of the 3-terminal 1768 nucleotides of the PS and ŏ6 isolates of plum pox virus (PPV) has been determined and compared with that of the equivalent regions of other PPV isolates sequenced previously. The sequenced region is part of the PPV open reading frame encoding the last 186 amino acids of the NIb protein and the coat protein (CP, 330 amino acids), followed by a non-coding region of 220 nucleotides and a poly(A) tail. PPV-PS and PPV(-)ŏ6, just like PPV-El Amar, show rather high levels of nucleotide diversity in the sequence encoding the C-terminal region of the NIb protein (19.4 to 31%) and the N terminus of CP (22.8 to 41.1%) when compared with PPV-Rankovic, PPV-D and PPV-NAT, whereas the level of diversity in the rest of the CP sequence and the 3 non-coding region is low (8 to 10.8% and 5.5 to 7.7%, respectively). However, the first 429 sequenced nucleotides of PPV(-)ŏ6 are very similar to those of the PPV-Rankovic, PPV-D and PPV-NAT isolates, whereas the rest of the sequence clearly resembles PPV-PS. Thus, PPV(-)ŏ6 seems to be the result of a natural recombination event between two wild strains of PPV. To our knowledge this is the first evidence of homologous RNA recombination (a process which could play an important role in the evolution of RNA viruses) within the potyvirus group.