Rod J. Scott

Stamen Structure and Function

Stamens are the male reproductive organs of flowering plants. They consist of an anther, the site of pollen development, and in most species a stalk-like filament, which transmits water and nutrients to the anther and positions it to aid pollen dispersal. Within the anther, male sporogenous cells

The AUXIN RESPONSE FACTOR 2 gene of Arabidopsis links auxin signalling, cell division, and the size of seeds and other organs

Control of seed size involves complex interactions among the zygotic embryo and endosperm, the maternally derived seed coat, and the parent plant. Here we describe a mutant in Arabidopsis, megaintegumenta (mnt), in which seed size and weight are dramatically increased. One factor in this is extra cell division in the integuments surrounding mnt mutant ovules, leading to the formation of enlarged seed coats. Unusually for integument mutants, mnt does not impair female fertility. The mnt lesion also has pleiotropic effects on vegetative and floral development, causing extra cell division and expansion in many organs. mnt was identified as a mutant allele of AUXIN RESPONSE FACTOR 2 (ARF2), a member of a family of transcription factors that mediate gene expression in response to auxin. The mutant phenotype and gene expression studies described here provide evidence that MNT/ARF2 is a repressor of cell division and organ growth. The mutant phenotype also illustrates the importance of growth of the ovule before fertilization in determining final size of the seed.

A mouse model of Rubinstein-Taybi syndrome: Defective long-term memory is ameliorated by inhibitors of phosphodiesterase 4

Mice carrying a truncated form of cAMP-responsive element binding protein (CREB)-binding protein (CBP) show several developmental abnormalities similar to patients with Rubinstein-Taybi syndrome (RTS). RTS patients suffer from mental retardation, whereas long-term memory formation is defective in mutant CBP mice. A critical role for cAMP signaling during CREB-dependent long-term memory formation appears to be evolutionarily conserved. From this observation, we reasoned that drugs that modulate CREB function by enhancing cAMP signaling might yield an effective treatment for the memory defect(s) of CBP+/− mice. To this end, we designed a cell-based drug screen and discovered inhibitors of phosphodiesterase 4 (PDE4) to be particularly effective enhancers of CREB function. We extend previous behavioral observations by showing that CBP+/− mutants have impaired long-term memory but normal learning and short-term memory in an object recognition task. We demonstrate that the prototypical PDE4 inhibitor, rolipram, and a novel one (HT0712) abolish the long-term memory defect of CBP+/− mice. Importantly, the genetic lesion in CBP acts specifically to shift the dose sensitivity for HT0712 to enhance memory formation, which conveys molecular specificity on the drugs mechanism of action. Our results suggest that PDE4 inhibitors may be used to treat the cognitive dysfunction of RTS patients.

EXS, a Putative LRR Receptor Kinase, Regulates Male Germline Cell Number and Tapetal Identity and Promotes Seed Development in Arabidopsis

BACKGROUND Plant germlines arise late in development from archesporial initials in the L2 layer of the anther and ovule primordia. These cells generate a radially symmetrical array of tissues that, in the Arabidopsis anther, comprises a core of sporogenous cells (meiocytes) and the enveloping tapetum, middle cell, and endothecium layers. The putative transcription factor NZZ/SPL is required for the specification of archesporial cells, but nothing is known of how their number is regulated, or what controls cell fate in the lineages they generate. Here, we report detailed characterization of extra sporogenous cells (exs), a male sterile mutant that generates extra meiocytes but lacks tapetal and middle cell layers. RESULTS We identified the EXS locus by map-based cloning and found it to encode a putative LRR receptor kinase. In the anther, an increased number of L2 layer cells assume an archesporial fate and divide to generate a larger number of sporogenous cells. In seeds, the exs mutation results in smaller embryonic cells, delayed embryo development, and smaller mature embryos. Consistent with the observed phenotype, EXS is expressed in the inflorescence meristem, floral apices, anthers, and in developing seeds. CONCLUSIONS EXS regulates the number of cells that divide in the L2 layer of the anther, and thus the number of functional male archesporial initials. In the young seed, EXS affects cell size in the embryo and the rate at which it develops. The apparently contrasting roles of EXS in the anther and embryo suggest that signaling through the EXS receptor kinase is a feature of a number of regulatory pathways in Arabidopsis.

Comparative Analysis between Homoeologous Genome Segments of Brassica napus and Its Progenitor Species Reveals Extensive Sequence-Level Divergence

Homoeologous regions of Brassica genomes were analyzed at the sequence level. These represent segments of the Brassica A genome as found in Brassica rapa and Brassica napus and the corresponding segments of the Brassica C genome as found in Brassica oleracea and B. napus. Analysis of synonymous base substitution rates within modeled genes revealed a relatively broad range of times (0.12 to 1.37 million years ago) since the divergence of orthologous genome segments as represented in B. napus and the diploid species. Similar, and consistent, ranges were also identified for single nucleotide polymorphism and insertion-deletion variation. Genes conserved across the Brassica genomes and the homoeologous segments of the genome of Arabidopsis thaliana showed almost perfect collinearity. Numerous examples of apparent transduplication of gene fragments, as previously reported in B. oleracea, were observed in B. rapa and B. napus, indicating that this phenomenon is widespread in Brassica species. In the majority of the regions studied, the C genome segments were expanded in size relative to their A genome counterparts. The considerable variation that we observed, even between the different versions of the same Brassica genome, for gene fragments and annotated putative genes suggest that the concept of the pan-genome might be particularly appropriate when considering Brassica genomes.

The Basis of Natural and Artificial Postzygotic Hybridization Barriers in Arabidopsis Species

The success or failure of interspecific crosses is vital to evolution and to agriculture, but much remains to be learned about the nature of hybridization barriers. Several mechanisms have been proposed to explain postzygotic barriers, including negative interactions between diverged sequences, global genome rearrangements, and widespread epigenetic reprogramming. Another explanation is imbalance of paternally and maternally imprinted genes in the endosperm. Interspecific crosses between diploid Arabidopsis thaliana as the seed parent and tetraploid Arabidopsis arenosa as the pollen parent produced seeds that aborted with the same paternal excess endosperm phenotype seen in crosses between diploid and hexaploid A. thaliana. Doubling maternal ploidy restored seed viability and normal endosperm morphology. However, substituting a hypomethylated tetraploid A. thaliana seed parent reestablished the hybridization barrier by causing seed abortion and a lethal paternal excess phenotype. We conclude from these findings that the dominant cause of seed abortion in the diploid A. thaliana × tetraploid A. arenosa cross is parental genomic imbalance. Our results also demonstrate that manipulation of DNA methylation can be sufficient to erect hybridization barriers, offering a potential mechanism for speciation and a means of controlling gene flow between species.

MATERNALLY EXPRESSED PAB C-TERMINAL, a Novel Imprinted Gene in Arabidopsis, Encodes the Conserved C-Terminal Domain of Polyadenylate Binding Proteins

Parental imprinting is important for seed development, but few imprinted genes have been identified in plants. The four known imprinted genes in Arabidopsis thaliana encode transcriptional regulators. Here, we describe a novel imprinted gene, MATERNALLY EXPRESSED PAB C-TERMINAL (MPC), which encodes the C-terminal domain of poly(A) binding proteins (PABPs). PABPs play roles in mRNA stability and translation. MPC interacts with proteins that also interact with the C-terminal domain of typical PABPs, suggesting that MPC may regulate translation by modulating PABP activity. In the endosperm, MPC is expressed only from the maternal allele. Reduction of MPC expression affects seed development. In dna methyltransferase1 (met1) mutants, MPC is ectopically expressed, and the paternal allele is active in the endosperm. CGs in the 5′ flanking region and gene body of MPC lose methylation in a met1 background. Both regions are required to confer imprinted reporter expression, suggesting that the gene body contains imprinting control region elements. In Arabidopsis, DEMETER (DME) activates expression of maternal alleles. MPC expression is reduced in flowers and seeds in a dme-4 mutant but only after fertilization in dme-1. We conclude that other factors along with DME promote MPC expression and that DME has indirect effects on imprinted gene expression in endosperm.

The Maternally Expressed WRKY Transcription Factor TTG2 Controls Lethality in Interploidy Crosses of Arabidopsis

The molecular mechanisms underlying lethality of F1 hybrids between diverged parents are one target of speciation research. Crosses between diploid and tetraploid individuals of the same genotype can result in F1 lethality, and this dosage-sensitive incompatibility plays a role in polyploid speciation. We have identified variation in F1 lethality in interploidy crosses of Arabidopsis thaliana and determined the genetic architecture of the maternally expressed variation via QTL mapping. A single large-effect QTL, DR. STRANGELOVE 1 (DSL1), was identified as well as two QTL with epistatic relationships to DSL1. DSL1 affects the rate of postzygotic lethality via expression in the maternal sporophyte. Fine mapping placed DSL1 in an interval encoding the maternal effect transcription factor TTG2. Maternal parents carrying loss-of-function mutations in TTG2 suppressed the F1 lethality caused by paternal excess interploidy crosses. The frequency of cellularization in the endosperm was similarly affected by both natural variation and ttg2 loss-of-function mutants. The simple genetic basis of the natural variation and effects of single-gene mutations suggests that F1 lethality in polyploids could evolve rapidly. Furthermore, the role of the sporophytically active TTG2 gene in interploidy crosses indicates that the developmental programming of the mother regulates the viability of interploidy hybrid offspring.

Hypomethylation Promotes Autonomous Endosperm Development and Rescues Postfertilization Lethality in fie Mutants

In most flowering plants, fertilization is necessary for development of the central cell into endosperm, but in the fie-1 mutant of Arabidopsis, the central cell can proliferate autonomously. However, autonomous fie-1 endosperms do not develop completely: They have fewer nuclei than sexually produced endosperms, cellularization does not take place, and no clear distinction is seen between the different endosperm compartments. Here, we show that autonomous endosperm develop much further in hypomethylated than normally methylated fie-1 mutants, undergoing cellularization and regional specification to resemble endosperm in sexually produced wild-type seeds. Therefore, the combination of maternal hypomethylation and loss of FIE function enables formation of differentiated endosperm without fertilization. A maternal fie-1 mutation is also lethal to sexual seeds, even if the pollen donor is wild type. We report that sexual mutant fie-1 endosperms fail to cellularize and overproliferate, consistent with the hypothesis that embryo abortion may be due, at least in part, to a defect in endosperm development. Finally, we show that pollen from hypomethylated plants rescues fie-1 mutant seeds provided that it also donates a wild-type paternal FIE allele. These results are discussed in light of models for parent-of-origin effects on seed development.

Genomic imprinting in plants and mammals: how life history constrains convergence

In both flowering plants and mammals, DNA methylation is involved in silencing alleles of imprinted genes, but surprising differences in imprinting control are emerging between the two taxa which may be traced to differences in their life cycles. Imprinted gene expression in plants occurs in the endosperm, a separate fertilisation product which transmits nutrients to the embryo and does not contribute a genome to the next generation. Regulation of expression of the known imprinted genes in Arabidopsis involves a cascade of gene expression beginning in the gametophyte, a haploid life phase interposed between the meiotic products and the gametes, which evolved from free-living organisms that constitute the dominant life phase of lower plants. Although the gametophytes of flowering plants are highly reduced they still express large numbers of genes, perhaps reflecting their evolutionary legacy, and which may now be recruited for control of imprinting. Strikingly, the genes at the top of the expression cascade appear to be specifically activated by demethylation, rather than targeted for silencing. Unlike in mammals, there is no evidence for global resetting of methylation in plants, and although imprinting involves the activity of a maintenance methyltransferase, de novo methyltransferases do not appear to be required. Plants do not set aside a germline; instead the cells that undergo meiosis to produce gametophytes differentiate in the adult plant during flower development. Both the late differentiation of the lineage producing germ cells, and the extent of gene expression during the haploid phase, may be incompatible with global resetting of methylation. Resetting may be unnecessary in any case because the adult plant expresses imprinted loci either biallelically or not at all, suggesting there is no chromosomal memory of parent-of-origin in the lineage that produces the gametophytes. Thus several features of the plant life cycle may account for the different strategies used by plants and animals to regulate parent-specific gene expression.