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Biomedical subjects

V F Irish

Publications and source records attributed to V F Irish.

At least 19 recordsLinked to original sources

Cell lineage, cell signaling and the control of plant morphogenesis.

It is clear that cell-cell signaling is critical for the development of both root and shoot structures. Recently, several of the key gene products required for intercellular signaling have been defined, and the developmental processes regulated by cell-cell interactions are beginning to be elucidated. Surprisingly, these results suggest that the mechanisms by which plant cells communicate with each other may be quite distinct from those used in animal systems.

Animals↗

The Arabidopsis floral homeotic gene APETALA3 differentially regulates intercellular signaling required for petal and stamen development.

Cell-cell signaling is crucial for the coordination of cell division and differentiation during plant organogenesis. We have developed a novel mosaic analysis method for Arabidopsis, based on the maize Ac/Ds transposable element system, to assess the requirements of individual genes in intercellular signaling. Using this strategy, we have shown that the floral homeotic APETALA3 (AP3) gene has distinct roles in regulating intercellular signaling in different tissues. In petals, AP3 acts primarily in a cell-autonomous fashion to regulate cell type differentiation, but its function is also required in a non-cell-autonomous fashion to regulate organ shape. In contrast, AP3-regulated intercellular interactions are required for conferring both cell type identity and organ shape and size in the stamens. Using antibodies raised against AP3, we have shown that the AP3 protein does not traffic between cells. These observations imply that AP3 acts by differentially regulating the production of intercellular signals in a whorl-specific manner.

Arabidopsis↗

Variations on a theme: flower development and evolution.

A recent study, comparing the maize SILKY1 gene to its well-characterized homolog APETALA3 from Arabidopsis, has provided some of the first evidence pointing to conservation of homeotic gene function between monocots and dicots.

Arabidopsis↗

Regulation of cell proliferation patterns by homeotic genes during Arabidopsis floral development.

The shoot apical meristem of Arabidopsis thaliana consists of three cell layers that proliferate to give rise to the aerial organs of the plant. By labeling cells in each layer using an Ac-based transposable element system, we mapped their contributions to the floral organs, as well as determined the degree of plasticity in this developmental process. We found that each cell layer proliferates to give rise to predictable derivatives: the L1 contributes to the epidermis, the stigma, part of the transmitting tract and the integument of the ovules, while the L2 and L3 contribute, to different degrees, to the mesophyll and other internal tissues. In order to test the roles of the floral homeotic genes in regulating these patterns of cell proliferation, we carried out similar clonal analyses in apetala3-3 and agamous-1 mutant plants. Our results suggest that cell division patterns are regulated differently at different stages of floral development. In early floral stages, the pattern of cell divisions is dependent on position in the floral meristem, and not on future organ identity. Later, during organogenesis, the layer contributions to the organs are controlled by the homeotic genes. We also show that AGAMOUS is required to maintain the layered structure of the meristem prior to organ initiation, as well as having a non-autonomous role in the regulation of the layer contributions to the petals.

Arabidopsis↗

Evolution of genetic mechanisms controlling petal development.

Molecular genetic studies in Arabidopsis thaliana and other higher-eudicot flowering plants have led to the development of the 'ABC' model of the determination of organ identity in flowers, in which three classes of gene, A, B and C, are thought to work together to determine organ identity. According to this model, the B-class genes APETALA3 (AP3) and PISTILLATA (PI) act to specify petal and stamen identity. Here we test whether the roles of these genes are conserved throughout the angiosperms by analysing the expression of AP3 and PI orthologues in the lower eudicot subclass Ranunculidae. We show that, although expression of these orthologues in the stamens is conserved, the expression patterns in the petals differ from those found in the higher eudicots. The differences between these expression patterns suggest that the function of AP3 and PI homologues as B-class organ-identity genes is not rigidly conserved among all angiosperms. These observations have important implications for understanding the evolution of both angiosperm petals and the genetic mechanisms that control the identities of floral organs.

Arabidopsis↗

Petal and stamen development.

Analyses of petal and stamen development are beginning to illuminate the molecular genetic processes that are required to elaborate these organ types. Floral homeotic genes are required to specify certain organ identities, and these functions also are required throughout organogenesis. These genes, either directly or indirectly, presumably control a wide array of tissue- and cell-type-specific differentiation processes. At least part of this repertoire seems to include the regulation of cell proliferation, coupling the specification of organ identity with changes in growth dynamics in different regions of the developing flower. Furthermore, cells have an enormous amount of developmental plasticity, which means that they have to be able to integrate multiple sources of information as they terminally differentiate. Some of the identified inputs include the position of the cell in the developing organ, the status of gene expression and epigenetic information, and environmental signals. How this information is disseminated between cells is largely unknown. Not only do individual cells need to respond to this information, but fields of cells must coordinate their differentiation to form a functionally complex structure. The challenge that is before us is to understand how this plasticity of response is regulated to give a reproducible and species-specific pattern of differentiated tissues.

Arabidopsis↗

CYP78A5 encodes a cytochrome P450 that marks the shoot apical meristem boundary in Arabidopsis.

The normal development of shoot structures depends on controlling the growth, proliferation and differentiation of cells derived from the shoot apical meristem. We have identified the CYP78A5 gene encoding a putative cytochrome P450 monooxygenase that is the first member of the CYP78 family from Arabidopsis. This gene is strongly expressed in the peripheral regions of the vegetative and reproductive shoot apical meristems, defining a boundary between the central meristematic zone and the developing organ primordia. In addition, CYP78A5 shows a dynamic pattern of expression during floral development. Overexpression of CYP78A5 affects multiple cell types, causing twisting and kinking of the stem and defects in floral development. To define the relationship of CYP78A5 to genes controlling meristem function, we examined CYP78A5 expression in plants mutant for SHOOT MERISTEMLESS, ZWILLE and ARGONAUTE, and have found that CYP78A5 expression is altered in these mutant backgrounds. We propose that CYP78A5 has a role in regulating directional growth in the peripheral region of the shoot apical meristem in response to cues established by genes regulating meristem function.

Amino Acid Sequence↗

Molecular evolution of genes controlling petal and stamen development: duplication and divergence within the APETALA3 and PISTILLATA MADS-box gene lineages.

The specification of floral organ identity in the higher dicots depends on the function of a limited set of homeotic genes, many of them members of the MADS-box gene family. Two such genes, APETALA3 (AP3) and PISTILLATA (PI), are required for petal and stamen identity in Arabidopsis; their orthologs in Antirrhinum exhibit similar functions. To understand how changes in these genes may have influenced the morphological evolution of petals and stamens, we have cloned twenty-six homologs of the AP3 and PI genes from two higher eudicot and eleven lower eudicot and magnolid dicot species. The sequences of these genes reveal the presence of characteristic PI- and AP3-specific motifs. While the PI-specific motif is found in all of the PI genes characterized to date, the lower eudicot and magnolid dicot AP3 homologs contain distinctly different motifs from those seen in the higher eudicots. An analysis of all the available AP3 and PI sequences uncovers multiple duplication events within each of the two gene lineages. A major duplication event in the AP3 lineage coincides with the base of the higher eudicot radiation and may reflect the evolution of a petal-specific AP3 function in the higher eudicot lineage.

Amino Acid Sequence↗

Discrete spatial and temporal cis-acting elements regulate transcription of the Arabidopsis floral homeotic gene APETALA3.

The APETALA3 floral homeotic gene is required for petal and stamen development in Arabidopsis. APETALA3 transcripts are first detected in a meristematic region that will give rise to the petal and stamen primordia, and expression is maintained in this region during subsequent development of these organs. To dissect how the APETALA3 gene is expressed in this spatially and temporally restricted domain, various APETALA3 promoter fragments were fused to the uidA reporter gene encoding beta-glucuronidase and assayed for the resulting patterns of expression in transgenic Arabidopsis plants. Based on these promoter analyses, we defined cis-acting elements required for distinct phases of APETALA3 expression, as well as for petal-specific and stamen-specific expression. By crossing the petal-specific construct into different mutant backgrounds, we have shown that several floral genes, including APETALA3, PISTILLATA, UNUSUAL FLORAL ORGANS, and APETALA1, encode trans-acting factors required for second-whorl-specific APETALA3 expression. We have also shown that the products of the APETALA1, APETALA3, PISTILLATA and AGAMOUS genes bind to several conserved sequence motifs within the APETALA3 promoter. We present a model whereby spatially and temporally restricted APETALA3 transcription is controlled via interactions between proteins binding to different domains of the APETALA3 promoter.

Arabidopsis↗

Floral homeotic gene expression defines developmental arrest stages in Brassica oleracea L. vars. botrytis and italica.

Brassica oleracea L. vars, italica (broccoli) and botrytis (cauliflower) both undergo developmental arrests which result in heading phenotypes. We characterized these arrested tissues at the morphological and molecular levels, and defined the developmental changes that ensure after arrest has been broken. We found that the order of floral organ initiation and the positions of resulting floral organ primordia in this species in some respects from that of Arabidopsis, which is a member of the same family, Brassicaceae. We also cloned homologs of the Arabidopsis floral homeotic genes APETALA1 (AP1) and APETALA3 (AP3) from B. oleracea and characterized their expression patterns. We found that the AP1 homolog was expressed in some of the meristems of arrest-stage cauliflower, providing evidence that this tissue is florally determined. In broccoli, both the AP1 and AP3 homologs were expressed. However, the spatial pattern of expression of the broccoli AP1 homolog differed from that of Arabidopsis. In addition we identified a homolog of the CAULIFLOWER (CAL) gene, BoiCAL, from broccoli. The predicted amino acid sequence indicated that the BoiCAL gene product does not contain the mutation thought to be responsible for the early arrest exhibited in cauliflower (Kempin et al. 1995), but contains other changes that might play a role in the broccoli heading phenotype.

Amino Acid Sequence↗

Nuclear localization of the Arabidopsis APETALA3 and PISTILLATA homeotic gene products depends on their simultaneous expression.

The Arabidopsis APETALA3 (AP3) and PISTILLATA (PI) proteins are thought to act as transcription factors and are required for specifying floral organ identities. To define the nuclear localization signals within these proteins, we generated translational fusions of the coding regions of AP3 and PI to the bacterial uidA gene that encodes beta-glucuronidase (GUS). Transient transformation assays of either the AP3-GUS or PI-GUS fusion protein alone resulted in cytoplasmic localization of GUS activity. However, coexpression of AP3-GUS with PI, or PI-GUS with AP3, resulted in nuclear localization of GUS activity. Stable transformation with these fusion proteins in Arabidopsis showed similar results. The nuclear colocalization signals in AP3 and PI were mapped to the amino-terminal regions of each protein. These observations suggest that the interaction of the AP3 and PI gene products results in the formation of a protein complex that generates or exposes a colocalization signal required to translocate the resulting complex into the nucleus. The colocalization phenomenon that we have described represents a novel mechanism to coordinate the functions of transcription factors within the nucleus.

Arabidopsis↗

Cellular interactions mediated by the homeotic PISTILLATA gene determine cell fate in the Arabidopsis flower.

Flowers develop from the coordinated division and differentiation of cells derived from the shoot apical meristem. By inducing chromosomal deletions in individual shoot apical meristem cells, we have generated Arabidopsis plants that are genetically mosaic for the homeotic PISTILLATA gene. Flowers bearing wild-type PISTILLATA epidermal tissue and mutant pistillata internal tissues are phenotypically normal. Based on this non-cell-autonomy, we suggest that PISTILLATA controls the production of a substance involved in cell-cell communication between the outer and inner tissue layers of the flower. These mosaic flowers were also used to assess the relative contributions of meristematic cells to the developing floral organs. These observations indicate that meristematic cells have discrete but somewhat variable contributions to the Arabidopsis flower. We have used these results to construct a fate map of the Arabidopsis floral primordium.

Arabidopsis↗

Conservation of floral homeotic gene function between Arabidopsis and antirrhinum.

Several homeotic genes controlling floral development have been isolated in both Antirrhinum and Arabidopsis. Based on the similarities in sequence and in the phenotypes elicited by mutations in some of these genes, it has been proposed that the regulatory hierarchy controlling floral development is comparable in these two species. We have performed a direct experimental test of this hypothesis by introducing a chimeric Antirrhinum Deficiens (DefA)/Arabidopsis APETALA3 (AP3) gene, under the control of the Arabidopsis AP3 promoter, into Arabidopsis. We demonstrated that this transgene is sufficient to partially complement severe mutations at the AP3 locus. In combination with a weak ap3 mutation, this transgene is capable of completely rescuing the mutant phenotype to a fully functional wild-type flower. These observations indicate that despite differences in DNA sequence and expression, DefA coding sequences can compensate for the loss of AP3 gene function. We discuss the implications of these results for the evolution of homeotic gene function in flowering plants.

Arabidopsis↗

Genetic ablation of petal and stamen primordia to elucidate cell interactions during floral development.

Two models have been proposed to explain the coordinated development of the four whorls of floral organs. The spatial model predicts that positional information defines the four whorls simultaneously, and that individual organs develop independently of surrounding tissues. The sequential model suggests that inductive events between the outer and inner whorl primordia are required for appropriate organogenesis. To test these models we have genetically ablated second and third whorl floral organ primordia to determine if organ identity, number or position are perturbed in the first or fourth whorls. We used diphtheria toxin to specifically ablate floral cells early in development in Nicotiana tabacum and in Arabidopsis thaliana. Second and third whorl expression of the diphtheria toxin A chain coding sequence (DTA) was conferred by the Arabidopsis APETALA3 (AP3) promoter. Both Nicotiana and Arabidopsis flowers that express the AP3-DTA construct lack petals and stamens; it appears that the second and third whorl cells expressing this construct arrest early in floral development. These results show that first and fourth whorl development is normal and can proceed without information from adjacent second and third whorl primordia. We propose that positional information specifies the establishment of all four whorls of organs prior to the expression of AP3 in the floral meristem.

Amino Acid Sequence↗

Cell lineage in plant development.

Lineage analyses in several plant species demonstrate that meristematic cells proliferate in a predictable manner to form the differentiated tissues of the mature shoot system. These studies also demonstrate, however, that the fates of meristematic cells are not absolutely dependent on their lineage. This variability indicates that interactions between cells must play a role in morphogenesis.

Cell Differentiation↗

Cell lineage in plant development.

Lineage analyses in several plant species demonstrate that meristematic cells proliferate in a predictable manner to form the differentiated tissues of the mature shoot system. These studies also demonstrate, however, that the fates of meristematic cells are not absolutely dependent on their lineage. This variability indicates that interactions between cells must play a role in morphogenesis.

Cell Differentiation↗