Regeneration of transgenic shape Vitis vinifera L. Sultana plants: genotypic and phenotypic analysis

Different approaches to producing transgenic grapevines based on regeneration via embryogenesis were investigated. Embryogenic callus was initiated from anther tissue of Vitis vinifera cv. Sultana and three embryogenic culture types (embryogenic callus, tissue type I; proliferating embryos, tissue type II; and a suspension) were established. The three culture types were incolucaled with Agrobacterium tumefaciens harbouring a binary vector which contained a uidA reporter gene and either a hpt or nptII selectable marker gene or the cultures were bombarded with microprojectiles carrying a uidA/nptII binary vector. Transgenic plants were produced only from Agrobacterium transformation experiments. Transformed embryos were selected with kanamycin or hygromycin antibiotics and recovered with the highest efficiency from inoculated type I cultures. Southern analysis of genomic DNA extracted from ten transgenic plants showed that the number of T-DNA insertions in the genome ranged from 1 to at least 4. Evidence for methylation of the T-DNA at cytosine and adenine residues in transgenic plants was found by Southern analysis of DNA digested with two isoschizomer pairs of restriction endonucleases. No evidence for genotype alterations or somatic meiosis was found when DNA from 80 somatic embryos and seven plants regenerated from embryogenic culture were analysed at six sequence-tagged sites which are heterozygous in cv. Sultana. Expression of the uidA gene in in vitro grown leaves of transgenic plants was most often high and uniform but GUS staining was occasionally observed to be low and/or patchy. Transgenic plants and all plants regenerated from embryogenic culture produced red veined, lobed leaves which are uncharacteristic of the accepted ampelographic phenotype of Sultana. It is suggested that this phenotype may represent a juvenile growth stage.     

Genetic modification of the waxy character in barley after an injection of wild-type exogenous DNA. Analysis of the second seed generation

Preparations of the DNA extracted from the endospermal milk and leaves of normal distichous barley plants were injected into the grains of hexastichous barley at a milk-ripeness stage and carrying waxy mutation (i.e. having defective synthesis of amylose). Restoration of normal starch synthesis in plant cells was considered as an indication of the changes produced by the exogenous DNA. In normal barley, starch contains amylose and amylopectin and is stained blue-to-black in an iodine solution, whereas in the waxy mutant, starch has no amylose and is stained red-to-brown. To exclude any relationship of dominance the analysis was carried out at a haploid level, that is on the pollen harvested from the injected plants. This approach had the additional advantage of allowing us to work with individual cells instead of whole plants or their organs. Upon injection of the preparations of wild-type DNA a certain fraction of plants showed changes toward wild type. The seeds obtained from the altered plants were sown and the progeny examined. It was established that in the second generation most plants returned to the recepient type. A detailed analysis of the progeny of plant No. 506/19, in the first generation of which almost all pollen (over 99.5%) was of wild type, showed that in these plants the alterations were considerably more stable, only one fifth or one sixth of plants returning to the original recipient type. In half of the progeny of plant No. 506/19 the starch was of the donor type, and in 30% of the plants, as in the first generations, the bulk of the polen (99.98%) was stained black by iodine. In the second generation, segregation of plants for starch staining differed from the Mendelian one, which is probably indicative of non-chromosomal localization of the genetic determinants brought along with the injected DNA. Concurrent with the alterations of the starch structure, alterations of the type of spike (hexastichous barley plants modified distichous ones) were observed in those plants whose pollen became predominantly or wholly normal. Also, delayed manifestation of the synthesis of normal starch was noted. Certain plants which did not show any changes in the first generation yielded wild-type pollen grains in the second generation.     

Cadmium tolerance and phytochelatin content of Arabidopsis seedlings over-expressing the phytochelatin synthase gene AtPCS1

Previous studies demonstrated that expression of the Arabidopsis phytochelatin (PC) biosynthetic gene AtPCS1 in Nicotiana tabacum plants increases the Cd tolerance in the presence of exogenous glutathione (GSH). In this paper, the Cd tolerance of Arabidopsis plants over-expressing AtPCS1 (AtPCSox lines) has been analysed and the differences between Arabidopsis and tobacco are shown. Based on the analysis of seedling fresh weight, primary root length, and alterations in root anatomy, evidence is provided that, at relatively low Cd concentrations, the Cd tolerance of AtPCSox lines is lower than the wild type, while AtPCS1 over-expressing tobacco is more tolerant to Cd than the wild type. At higher Cd concentrations, Arabidopsis AtPCSox seedlings are more tolerant to Cd than the wild type, while tobacco AtPCS1 seedlings are as sensitive as the wild type. Exogenous GSH, in contrast to what was observed in tobacco, did not increase the Cd tolerance of AtPCSox lines. The PC content in wild-type Arabidopsis at low Cd concentrations is more than three times higher than in tobacco and substantial differences were also found in the PC chain lengths. These data indicate that the differences in Cd tolerance and in its dependence on exogenous GSH between Arabidopsis and tobacco are due to species-specific differences in the endogenous content of PCs and GSH and may be in the relative abundance of PCs of different length.     

Actin cytoskeleton in Arabidopsis thaliana under blue and red light

BACKGROUND INFORMATION: Actin cytoskeleton is the basis of chloroplast-orientation movements. These movements are activated by blue light in the leaves of terrestrial angiosperms. Red light has been shown to affect the spatial reorganization of F-actin in water plants, where chloroplast movements are closely connected with cytoplasmic streaming. The aim of the present study was to determine whether blue light, which triggers characteristic responses of chloroplasts, i.e. avoidance and accumulation, also influences F-actin organization in the mesophyll cells of Arabidopsis thaliana. Actin filaments in fixed mesophyll tissue were labelled with Alexa Fluor 488-conjugated phalloidin. The configuration of actin filaments, expressed as a form factor (4 pi x area/perimeter(2)), was determined for all actin formations which were measured in fluorescence confocal images. RESULTS: In the present study, we compare form-factor distributions and the median form factors for strong and weak, blue- and red-irradiated tissues. Spatial organization of the F-actin network did not undergo any changes which could be attributed specifically to blue light. Actin patterns were similar in blue-irradiated wild-type plants and phot2 (phototropin 2) mutants which lack the avoidance response of chloroplasts. However, significant differences in the shape and distribution of F-actin formations were observed between mesophyll cells of phot2 mutants irradiated with strong and weak red light. These differences were absent in wild-type leaves. CONCLUSIONS: Actin does not appear to be the main target for the blue-light chloroplast-orientation signal. The modes of actin involvement in chloroplast translocations are different in water and terrestrial angiosperms. The results suggest that co-operation occurs between blue- and red-light photoreceptors in the control of the actin cytoskeleton architecture in Arabidopsis.     

Occurrence of a special type of "polypoid cushion" and valve-like formations in arteries]

Intraarterial structures, known as 'polypoid cushions' and 'valve-like' projections at arterial branching sites, were observed in serial sections of human cleft lip and nasal mucous membrane biopsies. A particular type of 'polypoid cushions', which additionally contains a small artery in its center, is distinguished. It is suggested that both the particular type of 'polypoid cushions' and these 'valve-like' projections originate from the invagination of the entire vessel wall at arterial branching sites according to the location of the dissection or damage. The differences between these 'valve-like' formations and genuine arterial valves are mentioned.     

The Miocene floras of Iceland and their significance for late Cainozoic North Atlantic biogeography

A large number of plant macrofossils from several Middle to Upper Miocene localities from Iceland have been studied. The fossil material includes four ferns and fern allies, seven conifers, and about 40 species of flowering plants. Betula islandica and Salix gruberi are described as new species. Coniferous twigs previously ascribed to the genus Sequoia are shown to belong to Cryptomeria based on macro‐morphological and epidermal features. Fossil plants from Iceland are compared with coeval fossil taxa from Europe and North America and with living plants. The main finding is that the Miocene flora of Iceland belongs to a widespread Neogene northern hemispheric floral type including plants whose representatives are restricted to East Asia, North America and to western Eurasia at the present time. Previously inferred conspicuous similarities to North American modern equivalents appear to be misleading. The type of vegetation in four plant‐bearing sedimentary formations from the late Mid Miocene to Late Miocene, the 12 Ma Brjánslækur‐Seljá Formation, the 10 Ma Tröllatunga‐Gautshamar Formation, the 9–8 Ma Skarðsströnd‐Mókollsdalur Formation, and the 7–6 Ma Hreðavatn‐Stafholt Formation, corresponds to a humid temperate broadleaved (deciduous)–coniferous mixed forest dominated by Betulaceae, Fagaceae and Acer. Changes in species composition in the sedimentary formations reflect a shift from warm temperate (Cfa climate) to cool temperate (Cfb climate) conditions from the late Mid Miocene to the latest Miocene. This shift was connected to repeated phases of extinction and colonization. Specifically, one set of thermophilic taxa including Magnolia, Liriodendron, Sassafras and Comptonia went extinct between 12 and 10 Ma, and appears to have been replaced by another set of thermophilic taxa in the 10 Ma formation (Juglandaceae aff. Pterocarya/Cyclocarya, Rhododendron ponticum type). The 9–8 and 7–6 Ma formations are characterized by taxa that migrated to Iceland from Europe, such as Fagus gussonii, Betula cristata and Pterocarya fraxinifolia type. Although there is convincing evidence that plants colonized Iceland both from North America and Europe until 12 Ma, migration in the younger formations (9–8, 7–6 Ma) is suggested to have occurred mainly from Europe. © 2005 The Linnean Society of London, Botanical Journal of the Linnean Society, 2005, 149 , 369–417.     

Occurrence of Temperature-Sensitive Phenotypic Plasticity in Chlorophyll-Deficient Mutants of Arabidopsis thaliana

A collection of 75 putative mutants with alterations in leaf pigmentation was visually selected from Arabidopsis thaliana plants (M₂ generation) grown at 26°C from seeds treated with the mutagen ethylmethanesulfonate. Fifty-eight of the plants were found to have chlorophyll contents decreased by at least 10% from the parental Columbia ecotype. These plants were screened for chlorophyll content and the ratio of chlorophyll b/a after growth at 20 or 26°C. Relative to the parental type, a significant number of individuals in which the chlorophyll-deficient phenotype was exacerbated at one of the growth temperatures were identified. We conclude that temperature-sensitive phenotypic plasticity for chlorophyll content is relatively common in mutant populations of higher plants.     

Plant functional types – a strategic perspective

Abstract. This paper discusses the use of Plant Functional Types to describe vegetation types – from plant communities to formations – in order to understand the ecological constraints of plants and plant communities on a higher level of abstraction. The terminology around plant functional types, traits and strategies is in need of further clarification and unification. The use of the term plant functional type is recommended. We are in need of a total environmental framework in which, hierarchically or reticulately, plant functional types can be compared and interpreted.     

Specific DNA Alterations Associated with the Environmental Induction of Heritable Changes in Flax

Several flax varieties have been shown to undergo environmentally induced heritable changes resulting in stable lines termed genotrophs. The most notable of these is the variety Stormont Cirrus, also termed "plastic" or Pl. A number of morphological, biochemical and genetic differences are associated with environmental induction of heritable changes in flax. We have studied 5S rDNA alterations as a model system for understanding environmental induction of heritable changes in flax. This paper reports the isolation of a flax 5S rRNA gene variant which identifies genotroph specific restriction fragment length polymorphisms (RFLPs) in flax. Restriction fragment patterns for several enzymes were observed in both large and small genotrophs which consistently differed from the progenitor, Stormont Cirrus. Identical RFLP profiles for all restriction endonucleases tested were observed in four small genotrophs produced from separate environmental induction experiments. Comparison between Stormont Cirrus and these small genotrophs showed at least six differing bands in addition to several high molecular weight polymorphisms. Genetic data indicate that the polymorphisms were all produced from a repetitive 5S rRNA gene cluster at a single chromosomal locus. Similar, but not identical, polymorphisms are also detected in other flax varieties and Linum species suggesting that the induced variation is related to that which occurs naturally. The results are evidence that a specific set of DNA alterations occur in association with the induction of heritable changes in flax. This is the first genetic marker which is altered to an identical state in one type of genotroph. The results are discussed with respect to mechanisms for environmentally induced heritable change in plants.     

ULTRASTRUCTURAL AND PHYSIOLOGICAL DIFFERENCES IN SOYBEANS WITH GENETICALLY ALTERED LEVELS OF PHOTOSYNTHETIC PIGMENTS

Soybeans displaying incomplete dominance for leaf pigmentation possess chloroplasts with characteristic shape and organization of photosynthetic membranes. The chloroplasts of light green plants lack grana typical of dark green or a field type (Beeson) but normally possess doubled thylakoids. Achlorophyllous lethal yellow plants have thylakoids reduced to single spiralled membranes. The yellow plants lack a waxy cuticle over the leaf surface which is characteristic of all other soybeans examined, and they lack catalase activity in microbodies. Photosynthetic rates in the light green plants are at least as high as in the fully pigmented ones and photorespiration levels are not significantly different. Thus, in light green plants greater efficiency of enzymatic processes in photosynthesis apparently offsets the inhibitory activities associated with photorespiration. Single allele alterations from dark green to light green and light green to lethal yellow appear to alter a variety of structures and functions.     

Influence of short photoperiods on the ultrastructure of hamster Harderian glands

Summary A sexual dimorphism of the hamster Harderian gland at the ultrastructural level, has been reported. Castration of males and the administration of testosterone to females are known to alter the male type gland to the female type and vice versa. In this paper we present data that exposure to a short photoperiod (1L:23D) can induce similar structural alterations between the 4th and 10th week of exposure. Changes in the male included reduction in the number of tubular clusters and large vacuoles, while in the female the number of membrane formations decreased with an accompanying appearance of tubular clusters.     

Genome size variations within Dasypyrum villosum: correlations with chromosomal traits, environmental factors and plant phenotypic characteristics and behaviour in reproduction

 Feulgen/DNA cytophotometric determinations carried out in the root meristem of seedlings showed that substantial quantitative alterations in the nuclear genome are present between and within 15 natural populations of Dasypyrum villosum in Italy. When the most variant values are considered, there is a 17.6% difference between the mean genome size of the populations, and a 66.2% difference between the genome size of individual plants within a population. A highly significant, positive correlation was found to exist between the genome size of D. villosum plants and the altitude of their stations, and differences in DNA contents between individual plants were greater in populations from mountain sites. Karyological analyses showed all chromosome pairs to differ largely in size between plants with differing DNA contents. A highly significant, positive correlation was found to exist between genome size and both the length of the chromosome complement at metaphase and the length and arm ratio of pair VII. Significant correlations were also found between DNA content and certain phenotypic characteristics of the plants. The mean genome size of the populations was negatively correlated with the mean leaf length and width. In contrast, the genome size of individual plants was positively correlated with the weight of the seed from which they originated and their flowering interval. A large range of genome sizes was found in the half-sib progeny of a plant having a relatively large genome. In contrast, in the half-sib progeny of a plant having a small genome, the genome sizes of the individual plants were less divergent and similar to that of the mother plant. All siblings from crosses between plants with differing genome sizes had similar DNA contents, which were intermediate between those of the parental plants, even if closer to the DNA content of the parent plant having the smaller genome size. Size polymorphism within pairs was never observed in plants obtained from these crosses or in half-sibs whose genome size differed from that of the mother plant. The intraspecific alterations observed in the nuclear genome and their effects on plant development and phenotype are briefly discussed as evolutionary factors which allow D. villosum populations to withstand different environmental conditions as well as the variability of conditions in a given environment. Received: 6 October 1997 / Accepted: 28 October 1997     

Genetic change in the waxy trait of barley under the influence of wild type DNA. Analysis of the composition of starch and the electrophoretic spectrum of caryopsis hordein from altered plants and the retention of these changes to the fourth generation

Injection of DNA isolated from the wild type of barley into grains of recipient mutant plants (waxy mutants) at the milk stage of maturity leads to a change in starch synthesis; type of spikes and hordein composition. In the first generation of injected plants the wild type starch synthesis was observed in some separate plants (these observations were made at a haploid level in pollen cells). In the second generation of transformed plants along with the change in starch and hordein synthesis a modification of the type of spikes was also revealed. Recipient plants had six-rowded (hexastichous) spikes, and donor plants--two rowded (distichous) spikes. Disc-electrophoresis of hordeins of the wild type barley (Yuzhny var.), hordeins of the waxy mutant (defected in synthesis of normal starch) and barley plants transformed under the action of wild type exogenous DNA reveals differences in the protein spectrum between donor, recipient and transformants. In the second generation in many of the transformed plants starch synthesis reverted to the recipient mutant type. Simultaneously a reversion of hordein composition to the initial mutant type was observed, and the distichous pikes became hexastichous. Analysis of the components of starch revealed that donor plant that have amilose and amilopectin in starch, and the recipient plants that lack amilose, can be distinguished by the spectra of light absorption of starch. For characterizing these differences the plot of absoprtions at 490 versus that at 590 nm was used. The tangens of angles of these curves for the waxy mutant were equal to 1.05 +/- 0.07 and 1.81 +/- 0.04 for the wild type barley. All transformants have a 1.78 ratio and for revertants this value was 1.02.     

Dispersion of energies in worldwide crop formations

The findings reported here lend further support to a decade of research which suggests that over 95% of worldwide crop formations involve organized ion plasma vortices that deliver lower atmosphere energy components of sufficient magnitude to produce significant bending, expansion and the formation of unique expulsion cavities in plant stem pulvini, as well as significant changes in seedling development. Here we demonstrated that, in a number of formations, quantitative node alterations can be directly related to fundamental concepts of electromagnetic energy absorption through the atmosphere. Also, in a detailed field study of simulated crop formations, we show that over-fertilization or gravitropism cannot account for observed physiological alterations. In addition, we demonstrate that details of flattened, intertwined crop patterns can be described by the application of fluid dynamic principles governing the interaction of rectilinearly moving vortex pairs.     

Aquatic and terrestrial vegetation of the Prespa area

We studied the vegetation of the aquatic ecosystems ofLake Mikri Prespa. The lacustrine vegetationcomprises three distinct forms: floating plants,benthic hydrophytes and helophytes, which aredescribed and classified from the phytosociologicalpoint of view, as follows: (a) the vegetation of thefloating plants belong to the Lemnetea class and isrepresented by two plant communities; (b) thevegetation of the benthic hydrophytes, belongs to thePotametea class consisting of two differentcategories, namely the submersed formations and theemergent formations of the hydrophytes. Various plantcommunities were recognised in this type of vegetationand three among them are considered as the mostrepresentative; (c) the vegetation of helophytes, theprevailing life form in this wetland, belongs to thePhragmitetea class and is represented mainly by sevenwell organised plant communities. The respectivevegetation of two of the five more important wetlandsites is described.The terrestrial vegetation is composed of forestand meadow vegetation. The forest vegetation of theNational Park belongs to the class Querco-Fagetea andshows the following zonation: (a) in the vicinity ofthe lake, at the elevation of 860–1000 m, twoassociations have been found: the mixed deciduous andevergreen forests of Ostryo-Carpinion orientalis andthe evergreen forests of Ostryo-Carpinion adriaticum;(b) the deciduous oak forests surround the previouszone at the altitude of 900–1300 m with two principalassociations, namely the Quercetum frainetto and theQuercetum petraeae; (c) in the upper forest layerbetween 1200–1800 m asl, dominate beech forests of theassociation Fagion moesiacum and the less extensiveoccurrence of the mixed beech-fir stands (Ass.Abieti-Fagetum moesiacum).The zone above the tree limit is distinguished by itssubalpine character semi-shrub vegetation extendinghigher than the forest (1800–2000 m), whereas alpine meadowscover the vegetation at higheraltitudes. On the plains and in the forest clearingsexist herbaceous meadow formations of variablestructure, in parallel with the vegetation of specifichabitats, such as nitrophilous and ammophilousplants.     

滇西北县域生物多样性本底调查与评估

中国是世界上生物多样性最丰富的国家之一, 同时也是生物多样性受威胁最严重的国家之一。为了有效保护生物多样性, 2010年国务院批准实施了《中国生物多样性保护战略与行动计划(2010-2030年)》, 划定了32个陆地生物多样性保护优先区, 并设定了开展优先区生物多样性本底调查的战略目标、优先领域与优先行动。为此, 2010-2011年, 环境保护部联合中国科学院和高校的科研人员, 在滇西北开展了18个县的以县域为单元的生物多样性本底示范调查与研究。调查内容包括生态系统(植被类型)和物种两个层次。生态系统主要调查县域内植被类型的多样性, 完成了以群系为单位的植被类型编目; 物种层次主要调查县域内高等植物、脊椎动物、大型真菌的物种多样性组成、数量和用途等, 分析了特有物种和珍稀濒危物种数量等, 完成了县域物种编目。本文基于调查结果, 比较研究了不同县域间的生物多样性组成, 发现植被类型(108个群系)和物种(高等植物4,481种、脊椎动物625种、大型真菌222种)最丰富的县均为玉龙县。同时, 与历史记录对比研究发现, 滇西北的生物多样性分布数据十分欠缺, 严重影响了生物多样性保护的客观有效决策。生物多样性本底调查是生物多样性保护的一项基础工作, 本研究为中国未来开展大规模的生物多样性本底调查与评估提供了案例。     

散枝猪毛菜的种子多型性及其萌发行为

散枝猪毛菜(Salsola bracchita)主要分布于新疆准噶尔荒漠,具有很强的抗干旱和抗盐碱能力。该文对散枝猪毛菜的果实进行了观察,表明散枝猪毛菜具有4种类型的散布单位和果实,这4种果实在形状、大小、颜色和着生方式上均有显著差异。A型果实绿色、球形、大,着生方式为横生,宿存花被革质,背部有紫红色翅状附属物,直径为(2.161±0.138) mm,单粒重为(3.810±0.113) mg;B型果实绿色、大、扁平、长圆形,着生方式为直立,宿存花被革质,背部有翅状附属物,长为(2.062±0.188) mm,宽为(1.720±0.148) mm,单粒重为(2.665±0.113) mg;C型果实绿色、大、扁平、长圆形,着生方式为直立,宿存花被膜质,背部无翅状附属物,长为(2.239±0.277) mm,宽为(1.844±0.150) mm,单粒重为(2.723±0.559) mg;D型果实黄色、小、扁平、长卵形,着生方式为直立,宿存花被膜质,背部无翅状附属物,长为(1.678±0.163) mm,宽为(1.390±0.110) mm,单粒重为(0.928±0.025) mg。A型、B型和C型种子(果实)在5 ℃/15 ℃、5 ℃/25 ℃、15 ℃/25 ℃(暗12 h/光12 h)变温条件下萌发率>68%,且B型和C型种子比A型种子有较高的萌发率和萌发速率。D型种子在3种变温条件下萌发缓慢,最终萌发率始终维持在较低水平(<20%),划破果皮和种皮能够显著提高D型种子的萌发率和萌发指数,表明D型种子处于生理休眠状态。散枝猪毛菜的种子多型性是对荒漠异质环境的一种适应。     

Expression of the ribosome-inactivating protein JIP60 from barley in transgenic tobacco leads to an abnormal phenotype and alterations on the level of translation

In this paper we report the in-planta activity of the ribosome-inactivating protein JIP60, a 60-kDa jasmonate-induced protein from barley (Hordeum vulgare L.), in transgenic tobacco (Nicotiana tabacum L.) plants. All plants expressing the complete JIP60 cDNA under the control of the cauliflower mosaic virus (CaMV) 35S promoter exhibited conspicuous and similar phenotypic alterations, such as slower growth, shorter internodes, lanceolate leaves, reduced root development, and premature senescence of leaves. Microscopic inspection of developing leaves showed a loss of residual meristems and higher degree of vacuolation of mesophyll cells as compared to the wild type. When probed with an antiserum which was immunoreactive against both the N- and the C-terminal half of JIP60, a polypeptide with a molecular mass of about 30 kDa, most probably a processed JIP60 product, could be detected. Phenotypic alterations could be correlated with the differences in the detectable amount of the JIP60 mRNA and processed JIP60 protein. The protein biosynthesis of the transformants was characterized by an increased polysome/monosome ratio but a decreased in-vivo translation activity. These findings suggest that JIP60 perturbs the translation machinery in planta. An immunohistological analysis using the JIP60 antiserum indicated that the immunoreactive polypeptide(s) are located mainly in the nucleus of transgenic tobacco leaf cells and to a minor extent in the cytoplasm. Received: 31 July 1996 / Accepted: 18 February 1997     

Morphological changes in CHO and VERO cells treated with T-2 mycotoxin. Correlation with inhibition of protein synthesis

Exposure of Chinese hamster ovary and African green monkey kidney cells to T-2 mycotoxin resulted in several morphological changes which were related to inhibition of protein synthesis, the basic in vitro mechanism of action of the toxin. These changes, which occurred in both cell types, included disassociation of polysomes and mitochondrial cristae alterations. In addition, CHO cells displayed membrane bleb formations similar to those found in CHO cells after exposure to established inhibitors of protein synthesis, puromycin and anisomycin. Blebs could be either a result of protein synthesis inhibition or a non-specific early pathological response. Bleb formations were not observed in VERO cells under any experimental condition.