Temporal rhythm of petal programmed cell death in Ipomoea purpurea

• Flowers are the main sexual reproductive organs in plants. The shapes, colours and scents of corolla of plant flowers are involved in attracting insect pollinators and increasing reproductive success. The process of corolla senescence was investigated in Ipomoea purpurea (Convolvulaceae) in this study.
• In the research methods of plant anatomy, cytology, cell chemistry and molecular biology were used.
• The results showed that at the flowering stage cells already began to show distortion, chromatin condensation, mitochondrial membrane degradation and tonoplast dissolution and rupture. At this stage genomic DNA underwent massive but gradual random degradation. However, judging from the shape and structure, aging characteristics did not appear until the early flower senescence stage. The senescence process was slow, and it was completed at the late stage of flower senescence with a withering corolla.
• We may safely arrive at the conclusion that corolla senescence of I. purpurea was mediated by programmed cell death (PCD) that occurred at the flowering stage. The corolla senescence exhibited an obvious temporal rhythm, which demonstrated a high degree of coordination with pollination and fertilization.

     

Senescence mechanisms

Senescence in plants is usually viewed as an internally programmed degeneration leading to death. It is a developmental process that occurs in many different tissues and serves different purposes. Generally, apoptosis refers to programmed death of small numbers of animal cells, and it shows some special features at the cell level. Some senescing plant cells show some symptoms typical of apoptosis, while others do not. This review will focus primarily on leaf senescence with ultimate aim of explaining whole plant senescence (i.e., monocarpic senescence). Traditionally, the ideas on senescence mechanisms fall into two major groupings, nutrient deficiencies (e.g., starvation) and genetic programming (i.e., senescence-promoting and senescence-inhibiting genes). Considerable evidence indicates that nutrient deficiencies are not central senescence program components, while increasing evidence supports genetic programming. Because chlorophyll (Chl) and chloroplast (CP) breakdown are so prominent, leaf senescence is generally measured in terms of Chl loss. Although CP breakdown may not be the proximate cause of leaf cell death, it certainly is important as a source of nutrients for use elsewhere, e.g., for developing reproductive structures in monocarpic plants, and this loss limits assimilatory capacity. The CP is dismantled in an orderly sequence. Individual protein complexes seem to be taken out all at once, not one subunit at a time. Removal of any component, e.g., Chl, seems to destabilize the whole complex. It is of special interest that senescing CPs secrete Chl-containing globules indicating that some CP components are broken down outside the CP. Senescence appears to be imposed on the CP by the nucleus, and all the known senescence-altering genes except one, cytG in soybean, are nuclear. Only the d₁d₂ mutation(s) in soybean prevents a broad range of leaf senescence processes. Exactly, what causes cell death is unclear; however, the selective thiol protease inhibitor, E-64, does delay death, and this suggests that proteases play a key role.     

Autophagy activity contributes to programmed cell death in Caenorhabditis elegans

The physiological relationship between autophagy and programmed cell death during C. elegans development is poorly understood. In C. elegans, 131 somatic cells and a large number of germline cells undergo programmed cell death. Autophagy genes function in the removal of somatic cell corpses during embryogenesis. Here we demonstrated that autophagy activity participates in germ-cell death induced by genotoxic stress. Upon γ ray treatment, fewer germline cells execute the death program in autophagy mutants. Autophagy also contributes to physiological germ-cell death and post-embryonic cell death in ventral cord neurons when ced-3 caspase activity is partially compromised. Our study reveals that autophagy activity contributes to programmed cell death during C. elegans development.     

Gene Pathways That Delay Caenorhabditis elegans Reproductive Senescence

Reproductive senescence is a hallmark of aging. The molecular mechanisms regulating reproductive senescence and its association with the aging of somatic cells remain poorly understood. From a full genome RNA interference (RNAi) screen, we identified 32 Caenorhabditis elegans gene inactivations that delay reproductive senescence and extend reproductive lifespan. We found that many of these gene inactivations interact with insulin/IGF-1 and/or TGF-β endocrine signaling pathways to regulate reproductive senescence, except nhx-2 and sgk-1 that modulate sodium reabsorption. Of these 32 gene inactivations, we also found that 19 increase reproductive lifespan through their effects on oocyte activities, 8 of them coordinate oocyte and sperm functions to extend reproductive lifespan, and 5 of them can induce sperm humoral response to promote reproductive longevity. Furthermore, we examined the effects of these reproductive aging regulators on somatic aging. We found that 5 of these gene inactivations prolong organismal lifespan, and 20 of them increase healthy life expectancy of an organism without altering total life span. These studies provide a systemic view on the genetic regulation of reproductive senescence and its intersection with organism longevity. The majority of these newly identified genes are conserved, and may provide new insights into age-associated reproductive senescence during human aging.     

Developmental changes in the levels of SDS-extractable polypeptides during plastid morphogenesis

Summary A quantitative estimation of the levels of plastidic SDS-extractable polypeptides as separated by polyacrylamide gel electrophoresis is described to demonstrate the practicality of such an approach. Using an internal standard of cytochrome c and expressing all polypeptide levels as cytochrome c relative stain equivalents, the levels of most polypeptides from developing Avena plastids change relative to the period of greening especially over the period 12–20 h. Some changes in certain polypeptides can be shown to be due to plastid senescence rather than plastid development. There is also a distinct difference in the pattern of polypeptides when plastids are isolated from different laminar regions. An incubation study using etioplasts showed of the original 8 polypeptides, six were retained, two were lost, another two were formed during incubation but eleven polypeptides found in the in situ study never appeared.SDS: Sodium dodecyl sulphate.     

Stage-specific hypermutability of the regA locus of Volvox, a gene regulating the germ-soma dichotomy

Mutation at the regA locus confers on somatic cells of Volvox (which otherwise undergo programmed death) ability to redifferentiate as reproductive cells. Stable mutations at the regA locus, but not at other loci, were induced at high frequency when embryos at one particular stage were exposed to either UV irradiation, novobiocin, nalidixic acid, bleomycin, 4-hydroxyaminoquinoline-1-oxide, 5-bromodeoxyuridine, or 5-fluorouracil. All treatments led to some mutations that were not expressed until the second generation after treatment. The sensitive period was after somatic and reproductive cells of the next generation had been set apart, but before they had undergone cytodifferentiation. Hypermutability occurs in presumptive reproductive cells (in which regA is normally not expressed) somewhat before regA normally acts in somatic cells. We postulate that hypermutability of regA in the reproductive cells at this time reflects a change of state that the locus undergoes as it is inactivated.     

MORPHOGENETIC BEHAVIOR OF TOBACCO TISSUE CULTURES AND IMPLICATION OF PLANT SENESCENCE

Decreases in the growth and organ-forming capacities characterized continuously cultured tobacco (Nicotiana tabacum ‘Wisconsin 38‘) callus. The root-initiation ability was completely lost in 1 1/2-year-old cultures. The rate of shoot formation decreased to a low plateau in cultures that reached 1 1/2-3 years of age since the explanting. An inverse relationship between callus growth and in vitro clonal age was also observed. Studies with callus clones started from individually isolated pith cells showed that the growth and organ-forming potentials of somatic cells varied, signifying that cell alterations had occurred in vivo. Both totipotent and non-totipotent cell lines were obtained. Subculturing the single-cell lines through several passages disclosed that the morphogenetically depressed state was irreversible and instability was characteristic of the totipotent lines. In the latter, a change toward the morphogenetically repressed level was observed. These findings are discussed in relation to the phenomenon of senescence. It is suggested that an accumulation of somatic mutations, i.e., genetic alterations resulting in reductions in the morphogenetic potential of cells, is a basis underlying senescence.     

Ontogenesis secretion and senescence of Tocoyena bullata (Vell.) Mart. (Rubiacaeae) colleters

Colleters are secretory structure present on many families including Rubiaceae. Particular characteristics have been described about colleters secretory cells, however senescence process are still under debate. Tocoyena bullata (Vell.) Mart. (Rubiaceae) shoot apex were collected at Jardim Botânico do Rio de Janeiro, RJ/Brazil. Stipules were separated and fragments were fixed in 2.5% glutaraldehyde and 4.0% formaldehyde in 0.05 m sodium cacodylate buffer, pH 7.2, post fixed in 1.0% osmium tetroxide in the same buffer, dehydrated in acetone, critical‐point‐drying, sputtered coated and observed. For light microscopy fragments were fixed and dehydrated, infiltrated with historesin and stained with 1% toluidine blue. For transmission electron microscopy, the samples were infiltrated with Epoxi resin. Colleters are present on stipule adaxial surface. On the beginning of development, these structures are recognized as small projections. Later on, colleters differentiated and secrete by cuticle rupture. The colleters senescence occurs in a concomitant and indissoluble way of programmed cell death. Ultrastructural analyses during the process strongly suggest the senescence is based on a non‐autolitic programmed cell death. T. bullata colleters, present at stipule abaxial surface are cylindrical secretory structures. Colleters secretory cells originated as stipule projections; differentiate; secrete and senesce by programmed cell death. The secretion and the cell dead occurs in a concomitantly and indissoluble way.     

EVOLUTION OF DEVELOPMENTAL PROGRAMS IN VOLVOX (CHLOROPHYTA)1

The volvocine green algal genus Volvox includes ～20 species with diverse sizes (in terms of both diameter and cell number), morphologies, and developmental programs. Two suites of characters are shared among distantly related lineages within Volvox. The traits characteristic of all species of Volvox—large (>500) numbers of small somatic cells, much smaller numbers of reproductive cells, and oogamy in sexual reproduction—have three or possibly four separate origins. In addition, some species have evolved a suite of developmental characters that differs from the ancestral developmental program. Most multicellular volvocine algae, including some species of Volvox, share an unusual pattern of cell division known as palintomy or multiple fission. Asexual reproductive cells (gonidia) grow up to many times their initial size and then divide several times in rapid succession, with little or no growth between divisions. Three separate Volvox lineages have evolved a reduced form of palintomy in which reproductive cells are small and grow between cell divisions. In each case, these changes are accompanied by a reduction in the rate of cell division and by a requirement of light for cell division to occur. Thus, two suites of characters—those characteristic of all Volvox species and those related to reduced palintomy—have each evolved convergently or in parallel in lineages that diverged at least 175 million years ago (mya).     

Incorporation of tritiated thymidine in germinal and somatic cells of the onion fly, Hylemya antiqua (Diptera: Anthomyiidae), as determined by autoradiography

Incorporation of³H-thymidine both in germinal and somatic cell types in young male adults of the onion fly,Hylemya antiqua (Meigen) reveals among other things the temporal pattern of spermatogenesis. Labeling of cells in the female reproductive organs is also described. Nuclei of fat cells, midgut epithelium, accessory glands and muscles, and occasionally hemocytes also show labeling. In oenocytes and the Malpighian tubules tritiated thymidine is incorporated in the cytoplasm.     

Ultrastructure and histochemistry of the male reproductive system of the genus Callinectes Stimpson, 1860 (Brachyura: Portunidae)

We described the ultrastructure and histochemistry of the reproductive system of five Callinectes species, and evaluate the seasonal variation in weight of the reproductive system and hepatopancreas by comparing annual changes of somatic indices. The somatic indices changed little throughout the year. In Callinectes, spermatogenesis occurs inside the lobular testes and, within each lobule, the cells are at the same developmental stage. Spermatogenesis and spermiogenesis follow the same development pattern in all Callinectes studied. Mature spermatozoa are released into the seminiferous ducts through the collecting ducts. Cells of the vas deferens are secretory as evidenced by rough endoplasmic reticulum, Golgi complex, and secretory vesicles that produce the seminal fluid. The anterior vas deferens shows two portions: proximal and distal. In proximal portion (AVDp), spermatozoa are clustered and embedded in an electron-dense, basophilic glycoproteinaceous secretion Type I. In the distal portion (AVDd), the spermatophore wall is formed by incorporation of a less electron-dense glycoproteinaceous secretion Type II. The secretion Type I change to an acid polysaccharide-rich matrix that separates the spermatophores from each other. The median vas deferens (MVD) stores the spermatophores and produces the granular glycoproteinaceous seminal fluid. The posterior vas deferens (PVD) has few spermatophores. Its epithelium has many mitochondria and the PVD seminal fluid changes into a liquid and homogeneous glycoprotein. Many outpocketings in the PVD and MVD help to increase the fluid production. Overall, the reproductive pattern of Callinectes is similar to other species that produce sperm plugs. The secretions of AVD, MVD, and PVD are responsible for the polymerization that forms the solid, waxy plug in the seminal receptacle. The traits identified here are common to all Portunidae species studied so far.     

Germ-cell cluster formation in the telotrophic meroistic ovary of <Emphasis Type="Italic">Tribolium castaneum</Emphasis> (Coleoptera,Polyphaga, Tenebrionidae) and its implication on insect phylogeny

Tribolium castaneum has telotrophic meroistic ovarioles of the Polyphaga type. During larval stages, germ cells multiply in a first mitotic cycle forming many small, irregularly branched germ-cell clusters which colonize between the anterior and posterior somatic tissues in each ovariole. Because germ-cell multiplication is accompanied by cluster splitting, we assume a very low number of germ cells per ovariole at the beginning of ovariole development. In the late larval and early pupal stages, we found programmed cell death of germ-cell clusters that are located in anterior and middle regions of the ovarioles. Only those clusters survive that rest on posterior somatic tissue. The germ cells that are in direct contact with posterior somatic cells transform into morphologically distinct pro-oocytes. Intercellular bridges interconnecting pro-oocytes are located posteriorly and are filled with fusomes that regularly fuse to form polyfusomes. Intercellular bridges connecting pro-oocytes to pro-nurse cells are always positioned anteriorly and contain small fusomal plugs. During pupal stages, a second wave of metasynchronous mitoses is initiated by the pro-oocytes, leading to linear subclusters with few bifurcations. We assume that the pro-oocytes together with posterior somatic cells build the center of determination and differentiation of germ cells throughout the larval, pupal, and adult stages. The early developmental pattern of germ-cell multiplication is highly similar to the events known from the telotrophic ovary of the Sialis type. We conclude that among the common ancestors of Neuropterida and Coleoptera, a telotrophic meroistic ovary of the Sialis type evolved, which still exists in Sialidae, Raphidioptera, and a myxophagan Coleoptera family, the Hydroscaphidae. Consequently, the telotrophic ovary of the Polyphaga type evolved from the Sialis type. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Sex differences in senescence: the role of intra-sexual competition in early adulthood

Males and females frequently differ in their rates of ageing, but the origins of these differences are poorly understood. Sex differences in senescence have been hypothesized to arise, because investment in intra-sexual reproductive competition entails costs to somatic maintenance, leaving the sex that experiences stronger reproductive competition showing higher rates of senescence. However, evidence that sex differences in senescence are attributable to downstream effects of the intensity of intra-sexual reproductive competition experienced during the lifetime remains elusive. Here, we show using a 35 year study of wild European badgers (Meles meles), that (i) males show higher body mass senescence rates than females and (ii) this sex difference is largely attributable to sex-specific downstream effects of the intensity of intra-sexual competition experienced during early adulthood. Our findings provide rare support for the view that somatic maintenance costs arising from intra-sexual competition can cause both individual variation and sex differences in senescence.     

The site of RanGTP generation can act as an organizational cue for mitotic microtubules

Background information. RanGTP, which is generated on chromosomes during mitosis, is required for microtubule spindle assembly. Due to its restricted spatial generation within the cell it has been suggested that RanGTP acts as a spatial cue to organize site‐specific spindle assembly within the cell. However, the absence of a detectable sharp gradient of RanGTP in somatic cells has led to suggestions that it may only act as a spatial cue in large cells and that it may operate as a general activator of the mitotic cytosol in somatic cells. Results. We report that ectopic generation of RanGTP at the plasma membrane stimulates the formation of organized arrays of microtubules at the plasma membrane. Conclusions. These results suggest that the site of RanGTP generation in a mitotic somatic cell can generate critical spatial information that specifies where microtubules grow towards and where microtubules are organized. As RanGTP is normally generated on chromosomes, these results suggest that RanGTP may play an important role in specifying that spindle assembly occurs around chromosomes.     

Detection of the Nicotiana rustica chloroplast genome coding for the large subunit of Fraction I protein in a somatic hybrid in which only the N. tabacum chloroplast genome appeared to have been expressed

Nine plants were produced from anthers of a somatic hybrid which had been obtained by fusion of Nicotiana tabacum L. and N. rustica L. protoplants. As determined by electrofocusing, the Fraction I protein of the original somatic hybrid had largesubunit polypeptides exclusively of the N. tabacum type. Two of the plants regenerated from anthers contained Fraction-I-protein large subunits exclusively of the N. rustica type. Since each plant was regenerated from a single cell, the somatic hybrid must have had cells containing both the N. tabacum and N. rustica chloroplast genome although the latter was not expressed. Possibilities to account for this non-expression of a chloroplast genome in the somatic hybrid are discussed.     

Reproductive overload in <Emphasis Type="Italic">D. melanogaster</Emphasis> females—Its connection with longevity

The conception of individual reproduction is that fruit flies are genetically designed to oviposit at the highest possible rate. This rate is maintained right up to the moment when the organism begins to age, and the senescence is characterized by the reproduction rate falling exponentially. The population can be divided into z, s, m, and l-phenotypes depending on their resource allocation. The main part of the population consists of m-phenotype flies with a balanced resource allocation. The flies with z-phenotype are infecund; s-flies do not reach senility; and l-flies, in which the allocation is biased toward somatic organism maintenance, outlast the period of oviposition and die only after they have completely used up their reproductive potential. Individual reproductive patterns and reproductive phenotypes are analyzed for two D. melanogaster populations consisting of 493 and 474 flies. It is shown that there is a mortality curve for each phenotype, and that a part of the population dies out “prematurely” due to reproductive overload.     

Hydra,a powerful model for aging studies

Cnidarian Hydra polyps escape senescence, most likely due to the robust activity of their three stem cell populations. These stem cells continuously self-renew in the body column and differentiate at the extremities following a tightly coordinated spatial pattern. Paul Brien showed in 1953 that in one particular species, Hydra oligactis, cold-dependent sexual differentiation leads to rapid aging and death. Here, we review the features of this inducible aging phenotype. These cellular alterations, detected several weeks after aging was induced, are characterized by a decreasing density of somatic interstitial cell derivatives, a disorganization of the apical nervous system, and a disorganization of myofibers of the epithelial cells. Consequently, tissue replacement required to maintain homeostasis, feeding behavior, and contractility of the animal are dramatically affected. Interestingly, this aging phenotype is not observed in all H. oligactis strains, thus providing a powerful experimental model for investigations of the genetic control of aging. Given the presence in the cnidarian genome of a large number of human orthologs that have been lost in ecdysozoans, such approaches might help uncover novel regulators of aging in vertebrates.     

Protein synthesis in Volvox carteri f. nagariensis

Incorporation of radioactive amino acids was used to examine protein synthesis during the asexual reproductive cycle of the HK10 (female) strain of Volvox carteri f. nagariensis. It was observed that arginine is incorporated into TCA-precipitable material 500 to 1000 times as rapidly as other amino acids. Arginine is taken up by the organisms by a saturable transport system and concentrated against a steep concentration gradient. Intracellular arginine is incorporated with high efficiency: after 1 hr, more than 80% of the intracellular radioactivity is in a TCA-precipitable form. N-terminal analysis ruled out the possibility that this tracer arginine was being incorporated via a ribosome-independent arginyl tRNA-protein transferase and supported the alternative explanation that it was being incorporated via conventional protein synthesis. Proteins labeled with arginine were found to decay with the same time course as those labeled with another amino acid.A method for isolation of somatic and reproductive cells free of cross-contamination is reported. Using this method to isolate cells of prelabeled Volvox, it was observed that arginine is preferentially incorporated into reproductive cells. Reproductive cells exhibited four times as high an arginine-to-lysine incorporation ratio as somatic cells. Over half the total incorporated arginine that was recovered was in reproductive cells. Thus arginine incorporation should serve well as a probe to investigate developmentally significant protein synthesis in the reproductive cells of this species.