The ability to produce morphologically and developmentally normal embryos and, indeed, whole plants from undifferentiated somatic cells in culture, through the process of somatic embryogenesis, resides uniquely within the plant kingdom. Since the initial description of somatic embryo production from carrot callus cells more than 35 years ago (Steward et al., 1958), this unique developmental potential has been recognized both as an important pathway for the regeneration of plants from cell culture systems and as a potential model for studying early regulatory and morphogenetic events in plant embryogenesis. The last 5 to 10 years have witnessed an explosion in the number of species that can now be regenerated from cell culture into whole plants through somatic embryogenesis. The literature contains hundreds of references describing the specific manipulations required to effect somatic embryo development from a variety of agronomically and horticulturally important plants. Although this is obviously an extremely important application of the process of somatic embryogenesis, it is not the focus of this review. Rather, this review will focus on the use of somatic embryogenesis as a model system for understanding the regulation of gene expression required for the earliest developmental events in the life of a higher plant: the development of the fertilized zygote into a mature embryo. The events of fertilization and subsequent embryo development normally occur deep within maternal tissues. The early embryo is minute and is surrounded by both endosperm and maternal cells. Although the morphological description of embryo development has been extensively recorded through microscopy, molecular and biochemical analyses of early embryogenesis have been hampered significantly by this physical inaccessibility. As a consequence, we know very little about the genes that are necessary for early embryogenesis in higher plants and even less about their regulation. This is beginning to be remedied by recent intensive efforts to genetically identify genes required for early embryogenesis in model systems such as Arabidopsis and maize (see West and Harada, 1993, this issue); many interesting mutants have been identified that may provide entry points into molecular analyses of major morphogenetic events in embryogenesis. These analyses would be greatly enhanced by the availability of cell, tissue, and developmental stage-specific markers of important events in the differentiation of cells and the establishment of the major tissue systems of the plant, which occur early in embryogenesis. In addition, once genes have been identified that are essential for embryogenesis, the subsequent analysis of their regulation would be greatly facilitated by the availability of an appropriate in vitro model system that is not limited in tissue quantity or accessibility. The somatic embryo system represents just such a model system. This review will summarize the process of somatic embryogenesis and will address the strengths and limitations of somatic embryos as potential models for studying early events in plant embryo development.

https://www.jstor.org/stable/info/3869792

Somatic Embryogenesis: A Model for Early Development in Higher Plants.

https://www.cell.com/trends/plant-science/pdf/S1360-1385(04)00221-3.pdf

Performing the paradoxical: how plant peroxidases modify the cell wall

that are expressed before the onset of nitrogen fixation and are involved in infection and nodule development. The products of the late nodulin genes are involved in the interaction with the endosymbiont and in the metabolic specialization of the nodule (Nap and Bisseling, 1990). In the first part of this review, we describe the early steps of the interaction between rhizobia and legumes that result in the formation of a nitrogen-fixing nodule. We focus on the role of specific lipooligosaccharides secreted by rhizobia in the induction of these early steps. In the second part, we describe nodule functioning and compare actinorhizal and legume nodules.

/pdf/symbiotic-nitrogen-fixation-1by9qh2egl.pdf

Symbiotic Nitrogen Fixation.

SUMMARY
Many legumes respond to Rhizobium inoculation by developing unique structures known as nodules on their roots. The development of a legume nodule in which rhizobia convert atmospheric N2 into ammonia is a finely tuned process. Gene expression from both partners of the symbiosis must be temporally and spatially coordinated. Exactly how this coordination takes place is an area of intense study. Nodule morphogenesis appears to be elicited by at least two distinct signals: one from Rhizobium, a product of the nod genes (Nod factor), and a second signal, which is generated within plant tissues after treatment with Nod factor. The identity of the second signal is unknown but changes in the balance of endogenous plant hormones or the sensitivity of plant tissues to these hormones are likely to be involved. These hormonal changes may be triggered by endogenous flavonoids produced by the root in response to inoculation with Rhizobium. There is some controversy as to whether the legume nodule is an organ sui generis or a highly derived lateral root. A resolution of this question may become more critical as attempts to induce nodules on non-legume hosts, such as rice or maize, increase in number and scope.

/pdf/developmental-biology-of-legume-nodulation-3ol366huf6.pdf

Developmental biology of legume nodulation

Arabinogalactan-proteins (AGPs) are a family of extensively glycosylated hydroxyproline-rich glycoproteins that are thought to have important roles in various aspects of plant growth and development. After a brief introduction to AGPs highlighting the problems associated with defining and classifying this diverse family of glycoproteins, AGP structure is described in terms of the protein component (including data from molecular cloning), carbohydrate component, processing of AGPs (including recent data on glycosylphosphatidylinositol membrane anchors) and overall molecular shape. Next, the expression of AGPs is examined at several different levels, from the whole plant to the cellular levels, using a variety of experimental techniques and tools. Finally, AGP function is considered. Although the existing functional evidence is not incontrovertible, it does clearly point to roles for AGPs in vegetative, reproductive, and cellular growth and development as well as programmed cell death and social control. In addition and most likely inextricably linked to their functions, AGPs are presumably involved in molecular interactions and cellular signaling at the cell surface. Some likely scenarios are discussed in this context. AGPs also have functions of real or potential commercial value, most notably as emulsifiers in the food industry and as potential immunological regulators for human health. Several important questions remain to be answered with respect to AGPs. Clearly, elucidating the unequivocal functions of particular AGPs and relating these functions to their respective structures and modes of action remain as major challenges in the years ahead.

Arabinogalactan-proteins: structure, expression and function.

A set of cDNA clones have been characterized that represent early nodulin mRNAs from pea root nodules. By RNA transfer blot analyses, the different early nodulin mRNAs were found to vary in time course of appearance during the development of the indeterminate pea root nodule. In situ hybridization studies demonstrated that the transcripts were located in different zones, representing subsequent steps in development of the central tissue of the root nodule. ENOD12 transcripts were present in every cell of the invasion zone, whereas ENOD5, ENOD3, and ENOD14 transcripts were restricted to the infected cells in successive but partially overlapping zones of the central tissue. We conclude that the corresponding nodulin genes are expressed at subsequent developmental stages. The amino acid sequence derived from the nucleotide sequences of the cDNAs, in combination with the localization data, showed that ENOD5 is an arabinogalactan-like protein involved in the infection process, whereas ENOD3 and ENOD14 have a cysteine cluster suggesting that these are metal-binding proteins. Furthermore, we showed that there is a clear difference in the way Rhizobium induced the infection-related early nodulin genes ENOD5 and ENOD12. A factor acting over a long distance induced the ENOD12 gene, whereas a factor acting over a short distance activated the ENOD5 gene.

Sequential induction of Nodulin gene expression in the developing Pea Nodule

Somatic embryogenesis of carrot (Daucus carota L.) is inhibited by the glycosylation inhibitor tunicamycin. This inhibition is reversible by the addition of correctly glycosylated glycoproteins which have been secreted into the culture medium. To identify the proteins responsible for complementation, glycoproteins present in the medium of embryo cultures were purified and tested for their activity in the tunicamycin inhibition/ complementation assay. A 38-kDa glycoprotein was purified that could restore embryogenesis to more than 50% of that in untreated controls. This 38-kDa glycoprotein was identified as a heme-containing peroxidase on the basis of its A405/A280 ratio (Reinheit Zahl or RZ) and enzyme activity. The 38-kDa peroxidase consisted of four different cationic isoenzymes of which only one or possibly two appeared active in the complementation assay. The cationic peroxidase isoenzymes from the carrot medium could be effectively replaced by cationic horseradish peroxidases which depended on their catalytic properties for their ability to restore tunicamycin-inhibited somatic embryogenesis.

Tunicamycin-inhibited carrot somatic embryogenesis can be restored by secreted cationic peroxidase isoenzymes

EP1, an extracellular protein from carrot (Daucus carota) cell suspensions, has been partially characterized by means of an antiserum and a cDNA clone. In both embryo and suspension cultures different molecular mass EP1 proteins were detected, some of which (31, 32, 52, and 54 kilodaltons) were bound to the cell wall and released into the medium, whereas others (49, 60, and 62 kilodaltons) were more firmly bound to the cell wall and could be extracted with a salt solution. Immunoprecipitation of in vitro translation products revealed a single primary translation product of 45 kilodaltons, suggesting that EP1 heterogeneity is due to differential posttranslational modification. In seedlings organ-specific modification of EP1 proteins was observed, a phenomenon which did not persist in suspension cultures initiated from different seedling organs. In culture EP1 proteins were only found to be associated with vacuolated, nonembryogenic cells, and on these cells they were localized in loosely attached, pectin-containing cell wall material. Purified 52/54 kilodaltons EP1 proteins did not alleviate the inhibitory effect of the glycosylation inhibitor tunicamycin on somatic embryogenesis.

Heterogeneity and cell type-specific localization of a cell wall glycoprotein from carrot suspension cells.

Non-embryogenic carrot suspension cells secrete the EP1 glycoprotein. A cDNA clone encoding EP1 was isolated and sequenced. The EP1 sequence revealed a region of homology with Brassica S-locus glycoprotein genes, an Arabidopsis S-like gene and putative S-like receptor protein kinases from maize and Arabidopsis. EP1 gene expression, analysed by in situ mRNA localization, was detected in cells located at the surface of the seedling: in the epidermis of the root, the hypocotyl and the cotyledons, in the root cap, and in a crescent of cells in the apical dome of the shoot. In developing seeds, expression was most pronounced in both the inner and outer integument epidermis.

https://onlinelibrary.wiley.com/doi/pdf/10.1046/j.1365-313X.1993.04050855.x

The carrot secreted glycoprotein gene EP1 is expressed in the epidermis and has sequence homology to Brassica S-locus glycoproteins.

A 47 kDa glycoprotein, termed EP4, was purified from carrot cell suspension culture medium. An antiserum raised against EP4 also recognized a protein of 45 kDa that was ionically bound to the cell wall. EP4 was detected in culture media from both embryogenic and non-embryogenic cell lines and was found to be secreted by a specific subset of non-embryogenic cells. Secretion of the 47 kDa glycoprotein by embryogenic cells was not evident. The 45 kDa cell wall-bound EP4 protein was specific for non-embryogenic cells and was shown by immunolocalization to occur in the walls of clustered cells, with the highest levels in the walls separating adjacent cells. In seedlings, EP4 proteins were mainly found in roots. EP4 cDNA was cloned by screening a cDNA library with an oligonucleotide derived from an EP4 peptide sequence. The EP4 cDNA sequence was found to be 55% homologous to ENOD8, an early nodulin gene from alfalfa.

F.A. van Engelen

Papers

Sequential induction of Nodulin gene expression in the developing Pea Nodule

Tunicamycin-inhibited carrot somatic embryogenesis can be restored by secreted cationic peroxidase isoenzymes

Heterogeneity and cell type-specific localization of a cell wall glycoprotein from carrot suspension cells.

The carrot secreted glycoprotein gene EP1 is expressed in the epidermis and has sequence homology to Brassica S-locus glycoproteins.

Purification, immunological characterization and cdna cloning of a 47-kda glycoprotein secreted by carrot suspension cells.