Isolation and properties of minB, a complex genetic locus involved in correct placement of the division site in Escherichia coli.

Mutation of Escherichia coli minicell locus (minB) results in aberrant placement of the division septum. In this paper we report the isolation and characterization of the minB locus. Replacement of the chromosomal minB+ allele by cloned minB sequences containing transposon insertions resulted in the minicell phenotype, indicating that minB+ function is required to maintain the normal division pattern. Paradoxically, overexpression of the locus also resulted in the minicell phenotype. The locus codes for several peptides whose expression is coordinately affected by transposon mutations that also eliminate minB+ function. A subset of the minB peptides is sufficient to prevent minicell formation in minB1 mutants or to induce minicell formation when overproduced in wild-type strains, implicating these peptides in the normal process of localization of the division site. The results indicate that minB is a complex locus whose expression must be maintained within certain limits to maintain the normal pattern of localization of the division septum.     

Maternal Thalidomide Treatment and Cell Division in Cultured Rabbit Embryos: Automated Image Analysis of Autoradiographs

To determine whether thalidomide dysmorphogenesis was preceded by a changed pattern of cell division, embryos from thalidomide treated rabbits were cultured for four hours in a medium containing ³H-thymidine. Automated image analysis of serial autoradiographs of the embryos revealed no focus of increased or decreased cell division in the neural tube or limb bud. Thalidomide pretreatment caused a general decrease in cell division at 10 days post coitum. A similar effect was observed only in those 10 3/4 day embryos whose status in culture was low. It was concluded that unlike methotrexate, the dysmorphogenic effect of thalidomide could not be attributed to a generally suppressed mitotic rate; rather, this teratogen preprogrammes dysmorphogenesis before the onset of the limb bud outgrowth (10 days) with the result that teratogenesis is manifest only during limb development after 11 days.     

Cell lineage,axis formation,and the origin of germ layers in the amphipod crustacean Orchestia cavimana

Embryos of the amphipod crustacean Orchestia cavimana are examined during cleavage, gastrulation, and segmentation by using in vivo labelling. Single blastomeres of the 8- and 16-cell stages were labelled with DiI to trace cell lineages. Early cleavage follows a distinct pattern and the a/p and d/v body axes are already determined at the 4- and 8-cell stages, respectively. In these stages, the germinal rudiment and the naupliar mesoderm can be traced back to a single blastomere each. In addition, the ectoderm and the postnaupliar mesoderm are separated into right and left components. At the16-cell stage, naupliar ectoderm is divided from the postnaupliar ectoderm, and extraembryonic lineages are separated from postnaupliar mesoderm and endoderm. From our investigation, it is evident that the cleavage pattern and cell lineage of Orchestia cavimana are not of the spiral type. Furthermore, the results of the labelling show many differences to cleavage patterns and cell lineages in other crustaceans, in particular, other Malacostraca. The cleavage and cell lineage patterns of the amphipod Orchestia are certainly derived within Malacostraca, whose ancestral cleavage mode was most likely of the superficial type. On the other hand, Orchestia exhibits a stereotyped cell division pattern during formation and differentiation of the germ band that is typical for malacostracans. Hence, a derived (apomorphic) early cleavage pattern is the ontogenetic basis for an evolutionarily older cell division pattern of advanced developmental stages. O. cavimana offers the possibility to trace the lineages and the fates of cells from early developmental stages up to the formation of segmental structures, including neurogenesis at a level of resolution that is not matched by any other arthropod system.     

植物多肽激素研究概况

目前发现的植物多肽多达9 种。基于配基- 受体的胞间互作模式, 目前公认的植物多肽激素包括4种: 系统素(Systemin) 、植物硫肽激素(Phytosulfokine) 、SCR􊄯SP11 和CLV3 , 分别参与了植食性昆虫防御反应、细胞增殖、自交不亲和的识别, 以及茎分生组织干细胞分裂与分化平衡的维持。本文对四种植物多肽激素基因家族的研究进展做了较为详尽的综述, 并结合本试验室的研究进展做了展望。     

Dynamics of Escherichia coli chromosome segregation during multifork replication

Slowly growing Escherichia coli cells have a simple cell cycle, with replication and progressive segregation of the chromosome completed before cell division. In rapidly growing cells, initiation of replication occurs before the previous replication rounds are complete. At cell division, the chromosomes contain multiple replication forks and must be segregated while this complex pattern of replication is still ongoing. Here, we show that replication and segregation continue in step, starting at the origin and progressing to the replication terminus. Thus, early-replicated markers on the multiple-branched chromosomes continue to separate soon after replication to form separate protonucleoids, even though they are not segregated into different daughter cells until later generations. The segregation pattern follows the pattern of chromosome replication and does not follow the cell division cycle. No extensive cohesion of sister DNA regions was seen at any growth rate. We conclude that segregation is driven by the progression of the replication forks.     

From endocytosis to tumors through asymmetric cell division of stem cells

Recent studies in vertebrate and invertebrate model organisms uncover the importance of endocytosis for biased signaling during asymmetric cell division. In stem cells, perturbing polarity and asymmetric division affect their selfrenewal causing exponential proliferation, thereby giving rise to cancer. An emerging pattern is that endocytosis controls asymmetric cell division, which underlies stem cell selfrenewal and defective selfrenewal is on the basis of tumorigenesis caused by cancer stem cells.     

Mutations in the FASS gene uncouple pattern formation and morphogenesis in Arabidopsis development.

The pattern of cell division is very regular in Arabidopsis embryogenesis, enabling seedling structures to be traced back to groups of cells in the early embryo. Recessive mutations in the FASS gene alter the pattern of cell division from the zygote, without interfering with embryonic pattern formation: although no primordia of seedling structures can be recognised by morphological criteria at the early-heart stage, all elements of the body pattern are differentiated in the seedling. fass seedlings are strongly compressed in the apical-basal axis and enlarged circumferentially, notably in the hypocotyl. Depending on the width of the hypocotyl, fass seedlings may have up to three supernumerary cotyledons. fass mutants can develop into tiny adult plants with all parts, including floral organs, strongly compressed in their longitudinal axis. At the cellular level, fass mutations affect cell elongation and orientation of cell walls but do not interfere with cell polarity as evidenced by the unequal division of the zygote. The results suggest that the FASS gene is required for morphogenesis, i.e., oriented cell divisions and position-dependent cell shape changes generating body shape, but not for cell polarity which seems essential for pattern formation.     

A Mechanistic Model of the Regulation of Division Timing by the Circadian Clock in Cyanobacteria

The cyanobacterium Synechococcus elongatus possesses a circadian clock in the form of a group of proteins whose concentrations and phosphorylation states oscillate with daily periodicity under constant conditions. The circadian clock regulates the cell cycle such that the timing of the cell divisions is biased toward certain times during the circadian period, but the mechanism underlying this phenomenon remains unclear. Here, we propose a mechanism in which a protein limiting for division accumulates at a rate proportional to the cell volume growth and is modulated by the clock. This “modulated rate” model, in which the clock signal is integrated over time to affect division timing, differs fundamentally from the previously proposed “gating” concept, in which the clock is assumed to suppress divisions during a specific time window. We found that although both models can capture the single-cell statistics of division timing in S. elongatus, only the modulated rate model robustly places divisions away from darkness during changes in the environment. Moreover, within the framework of the modulated rate model, existing experiments on S. elongatus are consistent with the simple mechanism that division timing is regulated by the accumulation of a division limiting protein in a phase with genes whose activity peaks at dusk.     

Cell division events are essential for embryo patterning and morphogenesis: studies on dominant-negative cdc2aAt mutants of arabidopsis

During plant development, cell division events are coordinately regulated, leading to specific growth patterns. Experimental evidence indicates that the morphogenetic controls that act at the vegetative plant growth stage are flexible and tolerate distortions in patterns and frequencies of cell division. To address questions concerning the relationship between cell division and embryo formation, a novel experimental approach was used. The frequencies of cell division were reduced exclusively during embryo development of Arabidopsis by the expression of a dominant cdc2a mutant. The five independent transgenic lines with the highest levels of the mutant cdc2a affected embryo formation. In the C13 line, seeds failed to germinate. The C1, C5 and C12 lines displayed a range of distortions on the apical-basal embryo pattern. In the C3 line, the shoot apical meristem of the seedlings produced leaves defective in growth and with an incorrect phyllotactic pattern. The results demonstrate that rates of cell division do not dictate cellular differentiation of embryos. Nevertheless, whereas cell divisions are uncoupled from vegetative development, they are instrumental in elaborating embryo structures and modulating embryo and seedling morphogenesis.     

Division and differentiation during normal and liguleless-1 maize leaf development

The maize leaf is composed of a blade and a sheath, which are separated at the ligular region by a ligule and an auricle. Mutants homozygous for the recessive liguleless-1 (lg1) allele exhibit loss of normal ligule and auricle. The cellular events associated with development of these structures in both normal and liguleless plants are investigated with respect to the timing of cell division and differentiation. A new method is used to assess orientation of anticlinal division planes during development and to determine a division index based on recent epidermal cross-wall deposition. A normal leaf follows three stages of development: first is a preligule stage, in which the primordium is undifferentiated and dividing throughout its length. This stage ends when a row of cells in the preligule region divides more rapidly in both transverse and longitudinal anticlinal planes. During the second stage, ligule and auricle form, blade grows more rapidly than sheath, divisions in the blade become exclusively transverse in orientation, and differentiation begins. The third stage is marked by rapid increase in sheath length. The leaf does not have a distinct basal meristem. Instead, cell divisions are gradually restricted to the base of the leaf with localized sites of increased division at the preligule region. Divisions are not localized to the base of the sheath until near the end of development. The liguleless-1 homozygote shows no alteration in this overall pattern of growth, but does show distinct alteration in the anticlinal division pattern in the preligule region. Two abnormal patterns are observed: either the increase in division rate at the preligule site is absent or it exhibits loss of all longitudinal divisions so that only transverse (or cell-file producing) divisions are present. This pattern is particularly apparent in developing adult leaves on older lg1 plants, in which sporadic ligule vestiges form. From these and results previously published (Becraft et al. (1990) Devl Biol. 14), we conclude that the information carried by the Lg1+ gene product acts earlier in development than formation of the ligule proper. We hypothesize that Lg1+ may be effective at the stage when the blade-sheath boundary is first determined.     

Clonal variation and aging of diploid fibroblasts : Cinematographic studies of cell pedigrees

Time-lapse cinematographic (TLC) analysis of clones of human diploid fibroblasts indicate heterogeneity in clonal division behaviour. Variations are noted in interdivision time, clone size and generations per clone. Correlation coefficients for interdivision times of sister pairs are high in young clones and generally low in aged clones. A consistent division pattern at all population doubling levels is one of low average interdivision time for early and late generations of a clone and high average interdivision time for the middle range of generations of a clone. The clonal division patterns observed experimentally have been duplicated in computer simulated pedigrees. The computer model is based on an oscillating system which allows for flux of regulator substances. The critical concentrations of regulator substances determine the clonal division pattern for a given progenitor cell.     

Selection of the midcell division site in Bacillus subtilis through MinD-dependent polar localization and activation of MinC

Bacterial cell division commences with the assembly of the tubulin-like protein, FtsZ, at midcell to form a ring. Division site selection in rod-shaped bacteria is mediated by MinC and MinD, which form a division inhibitor. Bacillus subtilis DivIVA protein ensures that MinCD specifically inhibits division close to the cell poles, while allowing division at midcell. We have examined the localization of MinC protein and show that it is targeted to midcell and retained at the mature cell poles. This localization is reminiscent of the pattern previously described for MinD. Localization of MinC requires both early (FtsZ) and late (PbpB) division proteins, and it is completely dependent on MinD. The effects of a divIVA mutation on localization of MinC now suggest that the main role of DivIVA is to retain MinCD at the cell poles after division, rather than recruitment to nascent division sites. By overexpressing minC or minD, we show that both proteins are required to block division, but that only MinD needs to be in excess of wild-type levels. The results suggest a mechanism whereby MinD is required both to pilot MinC to the cell poles and to constitute a functional division inhibitor.     

Dnm1 forms spirals that are structurally tailored to fit mitochondria

Dynamin-related proteins (DRPs) are large self-assembling GTPases whose common function is to regulate membrane dynamics in a variety of cellular processes. Dnm1, which is a yeast DRP (Drp1/Dlp1 in humans), is required for mitochondrial division, but its mechanism is unknown. We provide evidence that Dnm1 likely functions through self-assembly to drive the membrane constriction event that is associated with mitochondrial division. Two regulatory features of Dnm1 self-assembly were also identified. Dnm1 self-assembly proceeded through a rate-limiting nucleation step, and nucleotide hydrolysis by assembled Dnm1 structures was highly cooperative with respect to GTP. Dnm1 formed extended spirals, which possessed diameters greater than those of dynamin-1 spirals but whose sizes, remarkably, were equal to those of mitochondrial constriction sites in vivo. These data suggest that Dnm1 has evolved to form structures that fit the dimensions of mitochondria.     

Regulation of DNA synthesis in bacteria: Analysis of the Bates/Kleckner licensing/initiation-mass model for cell cycle control

Bates and Kleckner have recently proposed that bacterial cell division is a licensing agent for a subsequent initiation of DNA replication. They also propose that initiation mass for DNA replication is not constant. These two proposals do not take into account older data showing that initiation of DNA replication can occur prior to the division event. This critical analysis is derived from measurements of DNA replication during the division cycle in cells growing at different, and more rapid, growth rates. Furthermore, mutants impaired in division can initiate DNA synthesis. The data presented by Bates and Kleckner do not support the proposal that initiation mass is variable, and the proposed pattern of DNA replication during the division cycle of the K12 cells analysed is not consistent with prior data on the pattern of DNA replication during the division cycle.     

Potential use of toxic thermolabile proteins to study protein quality control systems

SulA is an Escherichia coli division inhibitor with a short half-life whose accumulation results in filamentation. Here, we show that SulA is thermally unstable and forms aggregates at elevated temperatures. This property enables the selection of isolates with mutated protein quality control systems.     

The simulation model of growth and cell divisions for the root apex with an apical cell in application to Azolla pinnata

In contrast to seed plants, the roots of most ferns have a single apical cell which is the ultimate source of all cells in the root. The apical cell has a tetrahedral shape and divides asymmetrically. The root cap derives from the distal division face, while merophytes derived from three proximal division faces contribute to the root proper. The merophytes are produced sequentially forming three sectors along a helix around the root axis. During development, they divide and differentiate in a predictable pattern. Such growth causes cell pattern of the root apex to be remarkably regular and self-perpetuating. The nature of this regularity remains unknown. This paper shows the 2D simulation model for growth of the root apex with the apical cell in application to Azolla pinnata. The field of growth rates of the organ, prescribed by the model, is of a tensor type (symplastic growth) and cells divide taking principal growth directions into account. The simulations show how the cell pattern in a longitudinal section of the apex develops in time. The virtual root apex grows realistically and its cell pattern is similar to that observed in anatomical sections. The simulations indicate that the cell pattern regularity results from cell divisions which are oriented with respect to principal growth directions. Such divisions are essential for maintenance of peri-anticlinal arrangement of cell walls and coordinated growth of merophytes during the development. The highly specific division program that takes place in merophytes prior to differentiation seems to be regulated at the cellular level.     

Comparison of the size of paternal and maternal homologous chromosomes during the first 2 cleavage divisions in mouse embryos

The mouse metaphase chromosomes of the 1st and 2nd cleavage divisions were prepared without colchicine and stained with trypsin-Giemsa. Both the homologues had the same pattern of differential staining (position and number of bands and interbands) in all pairs of chromosomes. The measurements of homologues of the 1st, 2nd, 3rd, 4th and 5th pairs of autosomes have shown that at the first cleavage division metaphase the paternal chromosomes are 1.2 times, on the average longer than the maternal ones, whereas at the second division metaphase no reliable differences in the length of homologues were found. In mice, thus, the heterocyclic pattern of the paternal and maternal sets of chromosomes manifested itself during the 1st cleavage division only and disappeared fully beginning from the 2nd division. This appears to be due to the early functional activity of chromosomes, i.e. to the fact that already in the 2-cell embryos both the maternal and paternal genes are expressed.     

Osmolality-dependent relocation of penicillin-binding protein PBP2 to the division site in Caulobacter crescentus

The synthesis of the peptidoglycan cell wall is carefully regulated in time and space. In nature, this essential process occurs in cells that live in fluctuating environments. Here we show that the spatial distributions of specific cell wall proteins in Caulobacter crescentus are sensitive to small external osmotic upshifts. The penicillin-binding protein PBP2, which is commonly branded as an essential cell elongation-specific transpeptidase, switches its localization from a dispersed, patchy pattern to an accumulation at the FtsZ ring location in response to osmotic upshifts as low as 40 mosmol/kg. This osmolality-dependent relocation to the division apparatus is initiated within less than a minute, while restoration to the patchy localization pattern is dependent on cell growth and takes 1 to 2 generations. Cell wall morphogenetic protein RodA and penicillin-binding protein PBP1a also change their spatial distribution by accumulating at the division site in response to external osmotic upshifts. Consistent with its ecological distribution, C. crescentus displays a narrow range of osmotolerance, with an upper limit of 225 mosmol/kg in minimal medium. Collectively, our findings reveal an unsuspected level of environmental regulation of cell wall protein behavior that is likely linked to an ecological adaptation.