Rebecca S. Lamb

The bHLH transcription factor SPATULA regulates root growth by controlling the size of the root meristem

BackgroundThe Arabidopsis thaliana gene SPATULA (SPT), encoding a bHLH transcription factor, was originally identified for its role in pistil development. SPT is necessary for the growth and development of all carpel margin tissues including the style, stigma, septum and transmitting tract. Since then, it has been shown to have pleiotropic roles during development, including restricting the meristematic region of the leaf primordia and cotyledon expansion. Although SPT is expressed in roots, its role in this organ has not been investigated.ResultsAn analysis of embryo and root development showed that loss of SPT function causes an increase in quiescent center size in both the embryonic and postembryonic stem cell niches. In addition, root meristem size is larger due to increased division, which leads to a longer primary root. spt mutants exhibit other pleiotropic developmental phenotypes, including more flowers, shorter internodes and an extended flowering period. Genetic and molecular analysis suggests that SPT regulates cell proliferation in parallel to gibberellic acid as well as affecting auxin accumulation or transport.ConclusionsOur data suggest that SPT functions in growth control throughout sporophytic growth of Arabidopsis, but is not necessary for cell fate decisions except during carpel development. SPT functions independently of gibberellic acid during root development, but may play a role in regulating auxin transport or accumulation. Our data suggests that SPT plays a role in control of root growth, similar to its roles in above ground tissues.

AGRIS and AtRegNet. A Platform to Link cis-Regulatory Elements and Transcription Factors into Regulatory Networks

Gene regulatory pathways converge at the level of transcription, where interactions among regulatory genes and between regulators and target genes result in the establishment of spatiotemporal patterns of gene expression. The growing identification of direct target genes for key transcription factors (TFs) through traditional and high-throughput experimental approaches has facilitated the elucidation of regulatory networks at the genome level. To integrate this information into a Web-based knowledgebase, we have developed the Arabidopsis Gene Regulatory Information Server (AGRIS). AGRIS, which contains all Arabidopsis (Arabidopsis thaliana) promoter sequences, TFs, and their target genes and functions, provides the scientific community with a platform to establish regulatory networks. AGRIS currently houses three linked databases: AtcisDB (Arabidopsis thaliana cis-regulatory database), AtTFDB (Arabidopsis thaliana transcription factor database), and AtRegNet (Arabidopsis thaliana regulatory network). AtTFDB contains 1,690 Arabidopsis TFs and their sequences (protein and DNA) grouped into 50 (October 2005) families with information on available mutants in the corresponding genes. AtcisDB consists of 25,806 (September 2005) promoter sequences of annotated Arabidopsis genes with a description of putative cis-regulatory elements. AtRegNet links, in direct interactions, several hundred genes with the TFs that control their expression. The current release of AtRegNet contains a total of 187 (September 2005) direct targets for 66 TFs. AGRIS can be accessed at http://Arabidopsis.med.ohio-state.edu.

Participation of the Arabidopsis bHLH Factor GL3 in Trichome Initiation Regulatory Events

The development of trichomes (leaf hairs) from pluripotent epidermal cells in Arabidopsis (Arabidopsis thaliana) provides a powerful system to investigate the regulatory motifs involved in plant cell differentiation. We show here that trichome initiation is triggered within 4 h of the induction of the GLABRA3 (GL3) basic helix-loop-helix transcription factor. Within this developmental window, GL3 binds to the promoters of at least three genes previously implicated in the development and patterning of trichomes (GL2, CAPRICE, and ENHANCER OF TRIPTYCHON AND CAPRICE1) and activates their transcription. The in vivo binding of GL3 to the promoters of these genes requires the presence of the R2R3-MYB factor GL1, supporting a model in which a GL3-GL1 complex is part of the trichome initiation enhanceosome. In contrast, GL3 is recruited to its own promoter in a GL1-independent manner, and this results in decreased GL3 expression, suggesting the presence of a GL3 negative autoregulatory loop. In support of genetic analyses indicating that ENHANCER OF GL3 (EGL3) is partially redundant with GL3, we show that EGL3 shares some direct targets with GL3. However, our results suggest that GL3 and EGL3 work independently of each other. Taken together, our results provide a regulatory framework to understand early events of epidermal cell differentiation.

Evolutionary history of the poly(ADP-ribose) polymerase gene family in eukaryotes.

BackgroundThe Poly(ADP-ribose)polymerase (PARP) superfamily was originally identified as enzymes that catalyze the attachment of ADP-ribose subunits to target proteins using NAD+ as a substrate. The family is characterized by the catalytic site, termed the PARP signature. While these proteins can be found in a range of eukaryotes, they have been best studied in mammals. In these organisms, PARPs have key functions in DNA repair, genome integrity and epigenetic regulation. More recently it has been found that proteins within the PARP superfamily have altered catalytic sites, and have mono(ADP-ribose) transferase (mART) activity or are enzymatically inactive. These findings suggest that the PARP signature has a broader range of functions that initially predicted. In this study, we investigate the evolutionary history of PARP genes across the eukaryotes.ResultsWe identified in silico 236 PARP proteins from 77 species across five of the six eukaryotic supergroups. We performed extensive phylogenetic analyses of the identified PARPs. They are found in all eukaryotic supergroups for which sequence is available, but some individual lineages within supergroups have independently lost these genes. The PARP superfamily can be subdivided into six clades. Two of these clades were likely found in the last common eukaryotic ancestor. In addition, we have identified PARPs in organisms in which they have not previously been described.ConclusionsThree main conclusions can be drawn from our study. First, the broad distribution and pattern of representation of PARP genes indicates that the ancestor of all extant eukaryotes encoded proteins of this type. Second, the ancestral PARP proteins had different functions and activities. One of these proteins was similar to human PARP1 and likely functioned in DNA damage response. The second of the ancestral PARPs had already evolved differences in its catalytic domain that suggest that these proteins may not have possessed poly(ADP-ribosyl)ation activity. Third, the diversity of the PARP superfamily is larger than previously documented, suggesting as more eukaryotic genomes become available, this gene family will grow in both number and type.

The Paralogous Genes RADICAL-INDUCED CELL DEATH1 and SIMILAR TO RCD ONE1 Have Partially Redundant Functions during Arabidopsis Development

RADICAL-INDUCED CELL DEATH1 (RCD1) and SIMILAR TO RCD ONE1 (SRO1) are the only two proteins encoded in the Arabidopsis (Arabidopsis thaliana) genome containing both a putative poly(ADP-ribose) polymerase catalytic domain and a WWE protein-protein interaction domain, although similar proteins have been found in other eukaryotes. Poly(ADP-ribose) polymerases mediate the attachment of ADP-ribose units from donor NAD+ molecules to target proteins and have been implicated in a number of processes, including DNA repair, apoptosis, transcription, and chromatin remodeling. We have isolated mutants in both RCD1 and SRO1, rcd1-3 and sro1-1, respectively. rcd1-3 plants display phenotypic defects as reported for previously isolated alleles, most notably reduced stature. In addition, rcd1-3 mutants display a number of additional developmental defects in root architecture and maintenance of reproductive development. While single mutant sro1-1 plants are relatively normal, loss of a single dose of SRO1 in the rcd1-3 background increases the severity of several developmental defects, implying that these genes do share some functions. However, rcd1-3 and sro1-1 mutants behave differently in several developmental events and abiotic stress responses, suggesting that they also have distinct functions. Remarkably, rcd1-3; sro1-1 double mutants display severe defects in embryogenesis and postembryonic development. This study shows that RCD1 and SRO1 are at least partially redundant and that they are essential genes for plant development.

The poetry of reproduction: the role of "LEAFY" in "Arabidopsis thaliana" flower formation

For successful reproduction, angiosperms must form fertile flowers at the appropriate positions and at the appropriate times. The reproductive transition is especially important for monocarpic plants that only flower once. In the model annual plant Arabidopsis thaliana, this transition is controlled through regulation of a group of genes termed floral meristem identity genes, of which LEAFY (LFY) is arguably the most important. LFY orthologs are found throughout land plants and are essential for angiosperm reproduction. These genes have also been implicated in reproductive development in gymnosperms. LFY encodes a plant-specific transcription factor that can act as either an activator or repressor depending on context, including what co-factors it is interacting with. It controls multiple aspects of floral morphogenesis, including phyllotaxis, organ number, organ identity and determinacy. Much progress has been made in elucidating the molecular mechanisms through which LFY and its orthologs contribute to a precise switch to flowering. We discuss the current state of knowledge in Arabidopsis, with an emphasis on known target genes and co-factors of LFY.

Functions of the poly(ADP-ribose) polymerase superfamily in plants

Poly(ADP-ribosyl)ation is the covalent attachment of ADP-ribose subunits from NAD+ to target proteins and was first described in plants in the 1970s. This post-translational modification is mediated by poly(ADP-ribose) polymerases (PARPs) and removed by poly(ADP-ribose) glycohydrolases (PARGs). PARPs have important functions in many biological processes including DNA repair, epigenetic regulation and transcription. However, these roles are not always associated with enzymatic activity. The PARP superfamily has been well studied in animals, but remains under-investigated in plants. Although plants lack the variety of PARP superfamily members found in mammals, they do encode three different types of PARP superfamily proteins, including a group of PARP-like proteins, the SRO family, that are plant specific. In plants, members of the PARP family and/or poly(ADP-ribosyl)ation have been linked to DNA repair, mitosis, innate immunity and stress responses. In addition, members of the SRO family have been shown to be necessary for normal sporophytic development. In this review, we summarize the current state of plant research into poly(ADP-ribosyl)ation and the PARP superfamily in plants.

RCD1 and SRO1 are necessary to maintain meristematic fate in Arabidopsis thaliana

The RADICAL-INDUCED CELL DEATH1 and SIMILAR TO RCD ONE1 genes of Arabidopsis thaliana encode members of the poly(ADP-ribose) polymerase (PARP) superfamily and have pleiotropic functions in development and abiotic stress response. In order to begin to understand the developmental and molecular bases of the defects seen in rcd1-3; sro1-1 plants, this study used the root as a model. Double mutant roots are short and display abnormally organized root apical meristems. However, acquisition of most cell fates within the root is not significantly disrupted. The identity of the quiescent centre is compromised, the zone of cell division is smaller than in wild-type roots and abnormal divisions are common, suggesting that RCD1 and SRO1 are necessary to maintain cells in a division-competent state and to regulate division plane placement. In addition, differentiation of several cell types is disrupted in rcd1-3; sro1-1 roots and shoots, demonstrating that RCD1 and SRO1 are also necessary for proper cell differentiation. Based on the data shown in this article and previous work, we hypothesize that RCD1 and SRO1 are involved in redox control and, in their absence, an altered redox balance leads to abnormal development.

The R2R3 MYB Transcription Factors FOUR LIPS and MYB88 Regulate Female Reproductive Development

Gamete formation is an important step in the life cycle of sexually reproducing organisms. In flowering plants, haploid spores are formed after the meiotic division of spore mother cells. These spores develop into male and female gametophytes containing gametes after undergoing mitotic divisions. In the female, the megaspore mother cell undergoes meiosis forming four megaspores, of which one is functional and three degenerate. The megaspore then undergoes three mitotic cycles thus generating an embryo sac with eight nuclei. The embryo sac undergoes cellularization to form the mature seven-celled female gametophyte. Entry into and progression through meiosis is essential for megasporogenesis and subsequent megagametogenesis, but control of this process is not well understood. FOUR LIPS (FLP) and its paralogue MYB88, encoding R2R3 MYB transcription factors, have been extensively studied for their role in limiting the terminal division in stomatal development by direct regulation of the expression of cell cycle genes. Here it is demonstrated that FLP and MYB88 also regulate female reproduction. Both FLP and MYB88 are expressed during ovule development and their loss significantly increases the number of ovules produced by the placenta. Despite the presence of excess ovules, single and double mutants exhibit reduced seed set due to reduced female fertility. The sterility results at least in part from defective meiotic entry and progression. Therefore, FLP and MYB88 are important regulators of entry into megasporogenesis, and probably act via the regulation of cell cycle genes.

A conserved domain in the N‐terminus is important for LEAFY dimerization and function in Arabidopsis thaliana

The floral meristem identity gene LEAFY (LFY) of Arabidopsis thaliana is essential for the formation of fertile flowers and has roles in the control of several aspects of floral development, which include phyllotaxy and organ number and identity. This gene encodes a land plant-specific transcription factor and regulates expression of a number of genes that include other floral meristem identity genes and floral homeotic genes. Although the LFY DNA-binding domain has a structure that resembles that of helix-turn-helix proteins, LFY and its orthologs represent a novel family of transcription factors that are characterized by a conserved N-terminus domain of unknown function and a C-terminus DNA-binding domain. Many transcription factors act as dimers. These dimers are essential for the biological activity of the proteins. We demonstrate that LFY forms homodimers or oligomers in solution. This association is mediated through the N-terminus conserved region of the LFY protein. Although mutant LFY proteins that cannot dimerize in solution can bind DNA, the binding is weaker than that of wild type LFY protein. LFY-LFY interactions mediated by the N-terminus domain are essential for the biological activity of this protein, as mutations that abolish the ability to self-associate cannot complement an lfy null allele. Our data indicate: (i) that LFY, and probably its orthologs in other plants, must act in complexes that contain at least two LFY molecules; and (ii) that the N-terminus is essential for stabilization of LFY complexes. This situation is integral to the ability of LFY to regulate gene expression.