代谢工程应用研究进展

代谢工程发展已有二十多年的时间,其利用重组DNA技术,调控细胞生理功能,在微生物、植物和动物细胞中得到了广泛的应用。综述了代谢工程在微生物、植物和动物细胞中应用研究的最新进展,并对其今后发展方向做出展望。     

Plant cells - young at heart?

Dolly has become a synonym for one of the greatest breakthroughs in animal reproductive biology: the regeneration of a whole mammal from a somatic cell nucleus. The equivalent experiments in plants - the regeneration of whole plants from single differentiated cells - are comparatively easy. Does this apparent difference in the developmental potential of animal and plant somatic cells reflect mechanistic differences in the regulation and maintenance of their respective cell differentiation?     

Plant and animal homeodomains use convergent mechanisms for intercellular transfer

Homeoproteins are defined by the structure of their DNA-binding domain, the homeodomain. Intercellular transfer of homeoprotein was observed ex vivo between animal cells and in vivo in higher plants. In the latter case, transfer is through intercytoplasmic channels that connect plant cells, but these do not exist in animals. Here, we show that the homeodomain of KNOTTED1, a maize homeoprotein, is transferred between animal cells and that a mutation in the homeodomain blocking the intercellular transfer of KNOTTED1 in plants also inhibits the transfer of the KNOTTED1 homeodomain in animal cells. This mutation decreases nuclear addressing, and its effect on nuclear import and intercellular transfer is reverted by the addition of an ectopic nuclear localization signal. We propose that, despite evolutionary distance and the differences in multicellular organization, similar mechanisms are at work for intercellular transfer of homeoprotein in plants and animals. Furthermore, our results suggest that, at least in animals, homeodomain secretion requires passage through the nucleus.     

Ultrastructure of autophagy in plant cells

Just as with yeasts and animal cells, plant cells show several types of autophagy. Microautophagy is the uptake of cellular constituents by the vacuolar membrane. Although microautophagy seems frequent in plants it is not yet fully proven to occur. Macroautophagy occurs farther away from the vacuole. In plants it is performed by autolysosomes, which are considerably different from the autophagosomes found in yeasts and animal cells, as in plants these organelles contain hydrolases from the onset of their formation. Another type of autophagy in plant cells (called mega-autophagy or mega-autolysis) is the massive degradation of the cell at the end of one type of programmed cell death (PCD). Furthermore, evidence has been found for autophagy during degradation of specific proteins, and during the internal degeneration of chloroplasts. This paper gives a brief overview of the present knowledge on the ultrastructure of autophagic processes in plants.     

Plant cells — young at heart?

Dolly has become a synonym for one of the greatest breakthroughs in animal reproductive biology: the regeneration of a whole mammal from a somatic cell nucleus. The equivalent experiments in plants — the regeneration of whole plants from single differentiated cells — are comparatively easy. Does this apparent difference in the developmental potential of animal and plant somatic cells reflect mechanistic differences in the regulation and maintenance of their respective cell differentiation?     

Splicing of plant pre-mRNAs in animal systems and vice versa

To investigate similarities and differences of pre-mRNA splicing between higher plants and animals, we tested whether pre-mRNAs of higher plants can be accurately spliced in animal systems, and whether an animal pre-mRNA can be spliced in plant cells. Pre-mRNAs of the maize bronze locus and oat phytochrome type 3 are accurately spliced with moderate efficiency in a human (HeLa cell) nuclear extract. The first intervening sequence (IVS1) of bean phaseolin pre-mRNA is not excised in the human nuclear extract, but is removed (although at low efficiency) in intact monkey cells. However, the IVS1 of human alpha-globin pre-mRNA is not removed in tobacco cells. Our data suggest that the mechanisms of pre-mRNA splicing are similar, but not identical, in plants and animals.     

A unifying new model of cytokinesis for the dividing plant and animal cells

Cytokinesis ensures proper partitioning of the nucleocytoplasmic contents into two daughter cells. It has generally been thought that cytokinesis is accomplished differently in animals and plants because of the differences in the preparatory phases, into the centrosomal or acentrosomal nature of the process, the presence or absence of rigid cell walls, and on the basis of 'outside-in' or 'inside-out' mechanism. However, this long-standing paradigm needs further reevaluation based on new findings. Recent advances reveal that plant cells, similarly to animal cells, possess astral microtubules that regulate the cell division plane. Furthermore, endocytosis has been found to be important for cytokinesis in animal and plant cells: vesicles containing endocytosed cargo provide material for the cell plate formation in plants and for closure of the midbody channel in animals. Thus, although the preparatory phases of the cell division process differ between plant and animal cells, the later phases show similarities. We unify these findings in a model that suggests a conserved mode of cytokinesis.     

Characteristics of the organization of cell proliferation in plants in relation to the problem of stem cells

Cellular patterns of continual cell proliferation are considered in plants and animals. In plants, the cells, analogous to the animal stem cells, can be formed many times during plant ontogenesis. Their functioning as stem cells is determined by their position in the growing organ. This is the reason why a plant can grow for a very long time. There are some common features of cellular patterns of proliferation in plants and animals providing the stability and optimal subordination between cell proliferation, differentiation and function.     

H+-adenosine triphosphatases from plasma membranes

New data are presented on the organization of H+-pumps in plasma membranes of cells of bacteria fungi, plants and animals. It is shown that H+-ATPase of bacteria differs in principle from H+-ATPases of plasma membranes of other organisms. The transport H+, K+-ATPase functioning in cells of mucous membrane of the animal stomach as an electroneutral H+-pump is similar by its properties to Na+, K+-ATPase of plasma membranes of animal cells. H+-ATPase of plasma membranes in cells of fungi and higher plants which functions as an electrogenic H+-pump differs essentially from H+-ATPases of F0 X F1-type. Distribution of H+-ATPases in cells of different organisms and their evolution are under discussion.     

Bcl-2 family members localize to tobacco chloroplasts and inhibit programmed cell death induced by chloroplast-targeted herbicides

In mammalian cells, apoptosis is often mediated via organelles. While apoptotic-like cell death occurs in plants, the mechanistic details are unresolved. Transgenic tobacco plants have been generated that harbour selected animal anti-apoptotic genes. Subcellular fractionation followed by western blot analysis indicated that chloroplasts serve as a location for these animal anti-apoptotic proteins in addition to the established mitochondrial location. To explore the functional significance of this observation, tobacco plants were treated with three chloroplast-directed herbicides. Wild-type plants died and exhibited features associated with apoptosis. Transgenic plants survived and did not show any apoptotic-like characteristics. Moreover, the herbicide-induced apoptotic-like cell death was light requiring. It was concluded that chloroplasts may be involved in mediating certain types of plant programmed cell death.     

Chemical-induced apoptotic cell death in tomato cells: involvement of caspase-like proteases

 A new system to study programmed cell death in plants is described. Tomato (Lycopersicon esculentum Mill.) suspension cells were induced to undergo programmed cell death by treatment with known inducers of apoptosis in mammalian cells. This chemical-induced cell death was accompanied by the characteristic features of apoptosis in animal cells, such as typical changes in nuclear morphology, the fragmentation of the nucleus and DNA fragmentation. In search of processes involved in plant apoptotic cell death, specific enzyme inhibitors were tested for cell-death-inhibiting activity. Our results showed that proteolysis plays a crucial role in apoptosis in plants. Furthermore, caspase-specific peptide inhibitors were found to be potent inhibitors of the chemical-induced cell death in tomato cells, indicating that, as in animal systems, caspase-like proteases are involved in the apoptotic cell death pathway in plants. Received: 5 August 1999 / Accepted: 14 March 2000     

Endocytosis in plants

Endocytosis in animal cells has been heavily documented. Both fluid-phase and receptor-mediated modes of uptake have been frequently studied, and the endocytic pathway is well defined. This contrasts markedly with the situation in plants where our knowledge of this process is still rudimentary. Partly responsible for this situation has been the view, widely held among plant physiologists, that because of turgor, endocytosis cannot occur in plant cells. As discussed below, the case against endocytosis is no longer water-tight.
Endocytosis is a fact in protoplasts. It can be demonstrated with electron-dense tracers as well as with membrane impermeant dye Lucifer Yellow CH. The latter has also been used with success on both suspension-cultured and tissue cells of higher plants, suggesting that fluid-phase endocytosis is also a feature of cells with walls. Through the application of fluorescently labelled elicitor molecules, which specifically bind to the cell surface of suspension-cultured cells, it has also been possible to provide convincing evidence for the operation of receptor-mediated endocytosis in plants. A number of studies on protoplasts and cells clearly indicate that endocytosis in plants can be mediated by coated pits in the plasma membrane. At least one of the organelles that lie on the endocytic pathway in plants has a structurally similar counterpart in animal cells: the multivesicular body. The first recipient of internalized molecules is the partially coated reticulum, although its relationship to the Golgi apparatus and Golgi function remain to be clarified. The final target for endocytosis in plants appears to be the vacuole.     

Borate Transport and Cell Growth and Proliferation: Not Only in Plants

Boron is an abundant mineral essential for the life cycle of plants and may play a role in animal development and growth. Very little is known about boron homeostasis in plant and animal cells and the physiological roles of boron in animals. The recent identification of boron transporters, BOR1 in plants and NaBC1 in mammals, and that NaBC1 functions as an electrogenic Na+-coupled borate transporter essential for cell growth and proliferation open the way to probe the roles of boron in cellular function and physiology.     

Embryo pathway and stem cells in higher plants

The Weisman's conception of germ track is considered in historical context focusing on fundamental differences among germ tracks in animals and plants. Differentiation of animal germ track cells occurrs once whereas in plant ontogenesis this process is multiply realizing (a concept of recurrent embriony). Fundamental differences in morphogenesis and embryogenesis of animals and plants as well as the differences in the properties of somatic and stem cells provide plants with special and additional modes of variability and evolution which are absent in animals.     

ABCA1-dependent but apoA-I-independent cholesterol efflux mediated by fatty acid-bile acid conjugates (FABACs)

PCD (programmed cell death) in plants presents important morphological and biochemical differences compared with apoptosis in animal cells. This raises the question of whether PCD arose independently or from a common ancestor in plants and animals. In the present study we describe a cell-free system, using wheat grain nucellar cells undergoing PCD, to analyse nucleus dismantling, the final stage of PCD. We have identified a Ca2+/Mg2+ nuclease and a serine protease localized to the nucleus of dying nucellar cells. Nuclear extracts from nucellar cells undergoing PCD triggered DNA fragmentation and other apoptotic morphology in nuclei from different plant tissues. Inhibition of the serine protease did not affect DNA laddering. Furthermore, we show that the nuclear extracts from plant cells triggered DNA fragmentation and apoptotic morphology in nuclei from human cells. The inhibition of the nucleolytic activity with Zn2+ or EDTA blocked the morphological changes of the nucleus. Moreover, nuclear extracts from apoptotic human cells triggered DNA fragmentation and apoptotic morphology in nuclei from plant cells. These results show that degradation of the nucleus is morphologically and biochemically similar in plant and animal cells. The implication of this finding on the origin of PCD in plants and animals is discussed.     

Glutathione metabolism and possible biological roles in higher plants

Synthesis of glutathione in plants seems to proceed in the same series of enzyme catalysed reactions observed in animal cells; the pathway of glutathio     

Nutritional Aspects of Magnesium Metabolism

The absolute necessity for magnesium in plant and animal nutrition is easily appreciated when one realizes that magnesium is found in high concentration in cells. It is necessary for photosynthesis in plants and for all reactions involving adenosine triphosphate in plant and animal cells. Although it is abundant in nature in general, deficiencies occur in both plants and animals. Human beings need about 5 mg of magnesium per kg of body weight per day. Infants and young children need twice as much. Children and women during pregnancy or lactation require significantly greater amounts than normal adults. Various diseases result in a deficiency of magnesium because of interruption of food intake or intestinal or renal wasting of the mineral.     

Plant endocytosis: it is clathrin after all

Endocytosis occurs in plants, but the involvement of clathrin-coated vesicles has been unclear; a new study provides strong evidence that, as in animal cells, clathrin-coated vesicles are a major means of internalisation by plant cells.     

Messenger DNA in higher plants

Evidence is presented that, as in animal and human cells, plant cells can release a newly-synthesized DNA which can freely circulate in the plants. This DNA enters cells and their nuclei where it may be integrated and be expressed so acting, apparently, as a messenger-DNA.     

Cytokinesis: an emerging unified theory for eukaryotes?

In animal and fungal cells, cytokinesis involves an actomyosin ring that forms and contracts at the division plane. Important new details have emerged concerning the composition, assembly, and dynamics of these contractile rings. In addition, recent advances suggest that targeted membrane addition is a central feature of cytokinesis in animal cells - as it is in fungi and plants - and the coordination of actomyosin ring function with targeted exocytosis at the cleavage plane is being explored. Important new information has also emerged about the spatial and temporal regulation of cytokinesis, especially in relation to the function of the spindle midzone in animal cells and the control of cytokinesis by GTPase systems.