Epigenetic aspects of sexual and asexual seed development.

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Fruit development is actively restricted in the absence of fertilization in Arabidopsis

Flowering plants usually require fertilization to form fruit and seed and to initiate floral organ abscission in structures that do not contribute to the fruit. An Arabidopsis mutant that initiates seedless fruit without fertilization (fwf) or parthenocarpy was isolated and characterized to understand the factors regulating the transition between the mature flower and the initiation of seed and fruit development. The fwf mutant is fertile and has normal plant growth and stature. It sets fertile seed following self-pollination and fertilization needs to be prevented to observe parthenocarpy. The initiation of parthenocarpic siliques (fruit) was found to be dependent upon carpel valve identity conferred by FRUITFULL but was independent of the perception of gibberellic acid, shown to stimulate parthenocarpy in Arabidopsis following exogenous application. The recessive nature of fwf is consistent with the involvement of FWF in processes that inhibit fruit growth and differentiation in the absence of fertilization. The enhanced cell division and expansion in the silique mesocarp layer, and increased lateral vascular bundle development imply FWF has roles also in modulating silique growth post-fertilization. Parthenocarpy was inhibited by the presence of other floral organs suggesting that both functional FWF activity and inter-organ communication act in concert to prevent fruit initiation in the absence of fertilization

Developmentally regulated HEART STOPPER, a mitochondrially targeted L18 ribosomal protein gene, is required for cell division, differentiation, and seed development in Arabidopsis.

Evidence is presented for the role of a mitochondrial ribosomal (mitoribosomal) L18 protein in cell division, differentiation, and seed development after the characterization of a recessive mutant, heart stopper (hes). The hes mutant produced uncellularized endosperm and embryos arrested at the late globular stage. The mutant embryos differentiated partially on rescue medium with some forming callus. HES (At1g08845) encodes a mitochondrially targeted member of a highly diverged L18 ribosomal protein family. The substitution of a conserved amino residue in the hes mutant potentially perturbs mitoribosomal function via altered binding of 5S rRNA and/or influences the stability of the 50S ribosomal subunit, affecting mRNA binding and translation. Consistent with this, marker genes for mitochondrial dysfunction were up-regulated in the mutant. The slow growth of the endosperm and embryo indicates a defect in cell cycle progression, which is evidenced by the down-regulation of cell cycle genes. The down-regulation of other genes such as EMBRYO DEFECTIVE genes links the mitochondria to the regulation of many aspects of seed development. HES expression is developmentally regulated, being preferentially expressed in tissues with active cell division and differentiation, including developing embryos and the root tips. The divergence of the L18 family, the tissue type restricted expression of HES, and the failure of other L18 members to complement the hes phenotype suggest that the L18 proteins are involved in modulating development. This is likely via heterogeneous mitoribosomes containing different L18 members, which may result in differential mitochondrial functions in response to different physiological situations during development.Hongyu Zhang, Ming Luo, Robert C. Day, Mark J. Talbot, Aneta Ivanova, Anthony R. Ashton, Abed M. Chaudhury, Richard C. Macknight, Maria Hrmova, and Anna M. Koltuno

Somatic small RNA pathways promote the mitotic events of megagametogenesis during female reproductive development in arabidopsis

Female gamete development in Arabidopsis ovules comprises two phases. During megasporogenesis, a somatic ovule cell differentiates into a megaspore mother cell and undergoes meiosis to produce four haploid megaspores, three of which degrade. The surviving functional megaspore participates in megagametogenesis, undergoing syncytial mitosis and cellular differentiation to produce a multicellular female gametophyte containing the egg and central cell, progenitors of the embryo and endosperm of the seed. The transition between megasporogenesis and megagametogenesis is poorly characterised, partly owing to the inaccessibility of reproductive cells within the ovule. Here, laser capture microdissection was used to identify genes expressed in and/or around developing megaspores during the transition to megagametogenesis. ARGONAUTE5 (AGO5), a putative effector of small RNA (sRNA) silencing pathways, was found to be expressed around reproductive cells during megasporogenesis, and a novel semi-dominant ago5-4 insertion allele showed defects in the initiation of megagametogenesis. Expression of a viral RNAi suppressor, P1/Hc-Pro, driven by the WUSCHEL and AGO5 promoters in somatic cells flanking the megaspores resulted in a similar phenotype. This indicates that sRNA-dependent pathways acting in somatic ovule tissues promote the initiation of megagametogenesis in the functional megaspore. Notably, these pathways are independent of AGO9, which functions in somatic epidermal ovule cells to inhibit the formation of multiple megaspore-like cells. Therefore, one somatic sRNA pathway involving AGO9 restricts reproductive development to the functional megaspore and a second pathway, inhibited by ago5-4 and P1/Hc-Pro, promotes megagametogenesis.Matthew R. Tucker, Takashi Okada, Yingkao Hu, Andrew Scholefield, Jennifer M. Taylor and Anna M. G. Koltuno

Transcriptomes of the Anther Sporophyte: Availability and Uses

An anther includes sporophytic tissues of three outer cell layers and an innermost layer, the tapetum, which encloses a locule where the gametophytic microspores mature to become pollen. The sporophytic tissues also comprise some vascular cells and specialized cells of the stomium aligning the long anther axis for anther dehiscence. Studies of the anther sporophytic cells, especially the tapetum, have recently expanded from the use of microscopy to molecular biology and transcriptomes. The available sequencing technologies, plus the use of laser microdissection and in silico subtraction, have produced high-quality anther sporophyte transcriptomes of rice, Arabidopsis and maize. These transcriptomes have been used for research discoveries and have potential for future discoveries in diverse areas, including developmental gene activity networking and changes in enzyme and metabolic domains, prediction of protein functions by quantity, secretion, antisense transcript regulation, small RNAs and promoters for generating male sterility. We anticipate that these studies with rice and other transcriptomes will expand to encompass other plants, whose genomes will be sequenced soon, with ever-advancing sequencing technologies. In comprehensive gene activity profiling of the anther sporophyte, studies involving transcriptomes will spearhead investigation of the downstream gene activity with proteomics and metabolomics

Generation of an integrated Hieracium genomic and transcriptomic resource enables exploration of small RNA pathways during apomixis initiation

Background: Application of apomixis, or asexual seed formation, in crop breeding would allow rapid fixation of complex traits, economizing improved crop delivery. Identification of apomixis genes is confounded by the polyploid nature, high genome complexity and lack of genomic sequence integration with reproductive tissue transcriptomes in most apomicts. Results: A genomic and transcriptomic resource was developed for Hieracium subgenus Pilosella (Asteraceae) which incorporates characterized sexual, apomictic and mutant apomict plants exhibiting reversion to sexual reproduction. Apomicts develop additional female gametogenic cells that suppress the sexual pathway in ovules. Disrupting small RNA pathways in sexual Arabidopsis also induces extra female gametogenic cells; therefore, the resource was used to examine if changes in small RNA pathways correlate with apomixis initiation. An initial characterization of small RNA pathway genes within Hieracium was undertaken, and ovary-expressed ARGONAUTE genes were identified and cloned. Comparisons of whole ovary transcriptomes from mutant apomicts, relative to the parental apomict, revealed that differentially expressed genes were enriched for processes involved in small RNA biogenesis and chromatin silencing. Small RNA profiles within mutant ovaries did not reveal large-scale alterations in composition or length distributions; however, a small number of differentially expressed, putative small RNA targets were identified. Conclusions: The established Hieracium resource represents a substantial contribution towards the investigation of early sexual and apomictic female gamete development, and the generation of new candidate genes and markers. Observed changes in small RNA targets and biogenesis pathways within sexual and apomictic ovaries will underlie future functional research into apomixis initiation in Hieracium. research into apomixis initiation in Hieracium.David S. Rabiger, Jennifer M. Taylor, Andrew Spriggs, Melanie L. Hand, Steven T. Henderson, Susan D. Johnson, Karsten Oelkers, Maria Hrmova, Keisuke Saito, Go Suzuki, Yasuhiko Mukai, Bernard J. Carroll, and Anna M. G. Koltuno