Stages Of Cell Division Research Articles

Foliar Application of Salicylic Acid at Different Phenological Stages of Peach Fruit CV. 'Flordaking' Improves Harvest Quality and Reduces Chilling Injury during Low Temperature Storage.

Peaches are well-liked amongst the stone fruits in Pakistan. The peach industry faces significant losses, from harvesting to marketing. The objective of this study was to investigate the effectiveness of foliar sprays of salicylic acid (SA) on the fruit quality of peaches (cv. ‘Flordaking’) at the harvest and postharvest life or stages. Different concentrations of SA (control, 1, 2 and 3 mM) were sprayed on the plants at three growth stages of fruit, i.e., the cell division, cell enlargement and pit-hardening stages. In general, all the SA treatments improved the fruit quality at harvest and maintained higher levels of flesh firmness, titratable acidity and ascorbic acid during storage. However, fruit weight loss, soluble solid contents, membrane leakage, chilling injury, color development, disease and decay incidence and the climacteric peak of ethylene were lowered by SA treatment after six weeks of low-temperature storage. SA at a 3-mM concentration was proven to be the most effective in maintaining the quality for a longer period of time during low-temperature storage. Based on the results, it can be concluded that the application of SA at fruit development stages can improve the harvest quality and storability of ‘Flordaking’ peaches.

Molecular dynamics study of CDC25BR492L mutant causing the activity decrease of CDC25B

Cell division cycle 25B (CDC25B) was responsible for regulating the various stages of cell division in the cell cycle. R492L was one of the common types of CDC25B mutants. Researches showed that compared to CDC25BWT, CDC25BR492L mutant had a ∼100-fold reduction in the rate constant for forming phosphatase intermediate (k2). However, the molecular basis of how the CDC25BR492L mutant influenced the process of binding between CDC25B and CDK2/CyclinA was not yet known. Therefore, the optimizations of three-dimensional structure of the CDC25BWT-CDK2/CyclinA system and the CDC25BR492L-CDK2/CyclinA system were constructed by ZDOCK and RDOCK, and five methods were employed to verify the reasonability of the docking structure. Then the molecular dynamics simulations on the two systems were performed to explore the reason why CDC25BR492L mutant caused the weak interactions between CDC25BR492L and CDK2/CyclinA, respectively. The remote docking site (Arg488-Tyr497) and the second active site (Lys538-Arg544) of CDC25B were observed to have high fluctuations in the CDC25BR492L-CDK2/CyclinA system with post-analysis, where the high fluctuation of these two regions resulted in weak interactions between CD25B and CDK2. In addition, Asp38-Glu42 and Asp206-Asp210 of CDK2 showed the slightly descending fluctuation, and CDK2 revealed an enhanced the self-interaction, which made CDK2 keep a relatively stable state in the CDC25BR492L-CDK2/CyclinA system. Finally, Leu492 of CDC25B was speculated to be the key residue, which had great effects on the binding between CDC25BR492L and CDK2 in the CDC25BR492L-CDK2/CyclinA system. Consequently, overall analyses appeared in this study ultimately offered a helpful understanding of the weak interactions between CDC25BR492L and CDK2.

Diversification of LytM Protein Functions in Polar Elongation and Cell Division of Agrobacterium tumefaciens.

LytM-domain containing proteins are LAS peptidases (lysostaphin-type enzymes, D-Ala-D-Ala metallopeptidases, and sonic hedgehog) and are known to play diverse roles throughout the bacterial cell cycle through direct or indirect hydrolysis of the bacterial cell wall. A subset of the LytM factors are catalytically inactive but regulate the activity of other cell wall hydrolases and are classically described as cell separation factors NlpD and EnvC. Here, we explore the function of four LytM factors in the alphaproteobacterial plant pathogen Agrobacterium tumefaciens. An LmdC ortholog (Atu1832) and a MepM ortholog (Atu4178) are predicted to be catalytically active. While Atu1832 does not have an obvious function in cell growth or division, Atu4178 is essential for polar growth and likely functions as a space-making endopeptidase that cleaves amide bonds in the peptidoglycan cell wall during elongation. The remaining LytM factors are degenerate EnvC and NlpD orthologs. Absence of these proteins results in striking phenotypes indicative of misregulation of cell division and growth pole establishment. The deletion of an amidase, AmiC, closely phenocopies the deletion of envC suggesting that EnvC might regulate AmiC activity. The NlpD ortholog DipM is unprecedently essential for viability and depletion results in the misregulation of early stages of cell division, contrasting with the canonical view of DipM as a cell separation factor. Finally, we make the surprising observation that absence of AmiC relieves the toxicity induced by dipM overexpression. Together, these results suggest EnvC and DipM may function as regulatory hubs with multiple partners to promote proper cell division and establishment of polarity.

Disentangling the link between leaf photosynthesis and turgor in fruit growth.

Despite the importance of understanding plant growth, the mechanisms underlying how plant and fruit growth declines during drought remain poorly understood. Specifically, it remains unresolved whether carbon or water factors are responsible for limiting growth as drought progresses. We examine questions regarding the relative importance of water and carbon to fruit growth depending on the water deficit level and the fruit growth stage by measuring fruit diameter, leaf photosynthesis, and a proxy of cell turgor in olive (Olea europaea). Flow cytometry was also applied to determine the fruit cell division stage. We found that photosynthesis and turgor were related to fruit growth; specifically, the relative importance of photosynthesis was higher during periods of more intense cell division, while turgor had higher relative importance in periods where cell division comes close to ceasing and fruit growth is dependent mainly on cell expansion. This pattern was found regardless of the water deficit level, although turgor and growth ceased at more similar values of leaf water potential than photosynthesis. Cell division occurred even when fruit growth seemed to stop under water deficit conditions, which likely helped fruits to grow disproportionately when trees were hydrated again, compensating for periods with low turgor. As a result, the final fruit size was not severely penalized. We conclude that carbon and water processes are able to explain fruit growth, with importance placed on the combination of cell division and expansion. However, the major limitation to growth is turgor, which adds evidence to the sink limitation hypothesis.

An electrical model with microtubules, impedance measurements and COMSOL simulations for single MDA-MB-231 cells under extremely low frequency electromagnetic fields

This paper presents an electrical equivalent model with microtubules (MTs) for use in analyzing the division process of human breast cancer cells (MDA-MB-231). A three-dimensional cell capture and sensing array electrode (3D ECCSA) and an extremely-low-frequency electromagnetic field (ELF-EMF) were integrated for the real-time monitoring of human breast cancer cells for 12 h. The relationship between the cellular processes (cell-spreading stage, cell mitosis stage, and cell division stage) and the ELF-EMF can be analyzed using the electrical equivalent model, the impedance measurements in the experiment, and COMSOL simulations. This was the first time that a circuit model combined with MTs was used to analyze cancer cells under an ELF-EMF. MT impedance contains MT–cell membrane contact resistance(RMct), MT resistance (RMT), and MT capacitance (CMT). At the cell-spreading stage, the ELF-EMF-reduced assembly of the MTs impedes the cell spread, which leads to lower impedance variation. At the cell mitosis stage, the MT volume variation in the ELF-EMF group decreases faster than that in the control group, which indicates that the ELF-EMF influences the assembly of the MTs. In addition, the ELF-EMF impedes the assembly of the MTs between the mitotic spindles, and then the cells prolong the cell mitosis period, which can be analyzed through the cell impedance and our model. At cell division, the impedance increases because two daughter cells block more current. Our study showed that the novel model with MTs and a 3D ECCSA could analyze the cell mitosis process under an ELF-EMF.

Unchaining miniBacillus Strain PG10: Relief of FlgM-Mediated Repression of Autolysin Genes.

Cell chaining in Bacillus subtilis is naturally observed in a subset of cells during exponential growth and during biofilm formation. However, the recently constructed large-scale genome-minimized B. subtilis strain PG10 displays a severe and permanent defect in cell separation, as it exclusively grows in the form of long filaments of nonseparated cells. In this study, we investigated the underlying mechanisms responsible for the incomplete cell division of PG10 by genomic and transcriptomic analyses. Repression of the SigD regulon, including the major autolysin gene lytF, was identified as the cause for the cell separation problem of PG10. It appeared that SigD-regulated genes are downregulated in PG10 due to the absence of the flagellar export apparatus, which normally is responsible for secretion of FlgM, the anti-sigma factor of SigD. Although mild negative effects on growth and cell morphology were observed, deletion of flgM could revert the aberrant cell-chaining phenotype and increased transformation efficiency. Interestingly, our work also demonstrates the occurrence of increased antisense transcription of slrR, a transcriptional repressor of autolysin genes, in PG10 and provides further understanding for this observation. In addition to revealing the molecular basis of the cell separation defect in PG10, our work provides novel targets for subsequent genome reduction efforts and future directions for further optimization of miniBacillus PG10. IMPORTANCE Reduction of the size of bacterial genomes is relevant for understanding the minimal requirements for cellular life as well as from a biotechnological point of view. Although the genome-minimized Bacillus subtilis strain PG10 displays several beneficial traits as a microbial cell factory compared to its parental strain, a defect at the final stage of cell division was introduced during the genome reduction process. By genetic and transcriptomic analyses, we identified the underlying reasons for the cell separation problem of PG10. In addition to enabling PG10 to grow in a way similar to that of B. subtilis wild-type strains, our work points toward subsequent targets for fine-tuning and further reduction of the genome of PG10. Moreover, solving the cell separation defect facilitates laboratory handling of PG10 by increasing the transformation efficiency, among other means. Overall, our work contributes to understanding and improving biotechnologically attractive minimal bacterial cell factories.

Abstract 1435: The distribution of Tumor Treating Fields is affected by cell confluence and pores in the membrane

Abstract Introduction: Better understanding of the interaction between Tumor Treating Fields (TTFields) and living cells would help to elaborate on its mechanism of action. Numerical simulations investigating the electric field distribution within isolated cells have shown that during metaphase a uniform electric field forms within the rounded cells, and during cytokinesis a non-uniform field forms at the furrow, leading to strong dielectrophoretic (DEP) forces that can disrupt cell division. However, preclinical studies have shown that TTFields influence cells during earlier stages of cell division. Hence, field non-uniformity within the cell during cytokinesis cannot provide a full explanation for how TTFields exerts an anti-mitotic effect on cells. We hypothesized that strong DEP forces could arise at the boundaries between cells and in pores on the membrane when applying TTFields to cell culture. We tested this hypothesis by using numerical simulations to investigate how the clustering of cells and pores within the membrane influence TTFields distribution. Methods: COMSOL was used to numerically simulate delivery of TTFields to clusters of round cells placed in a hexagonal arrangement. The influence of the distance between the cells on field distribution was investigated. The effect of pores in the cell membrane on field distribution was also investigated. A generic analytical model was developed for predicting conditions for field amplification in the TTFields frequencies. Results: Placing round cells in clusters separated by ~10 nm resulted in regions of highly non-uniform fields within the cells and very strong DEP forces at the intercellular level. Maximum field intensities between cells were observed at 200 kHz. Very strong gradients in the electric field were observed around pores placed in the membrane, and a strong field amplification was observed at 200 kHz regardless of the pore size. For a 10 nm pore, up to 450 V/cm is generated at the pore's vicinity. 100 nm pores generate more than 130 V/cm for an applied external field of 1 V/cm. Electroporation generates enough energy for overcoming the internal KT energy at pores that are ≤30 nm and may allow for dipole alignment. Conclusions: Cells in close proximity to one another creates gradients in the electric field, and are associated with strong DEP forces that enhance the effects of TTFields on cells. Strong DEP forces in the membrane may provide a physical mechanism by which TTFields enhance membrane permeability. The strong field amplification could cause local heating, particle diffusion, and impact the cytoskeleton in the vicinity of membrane pores. These simulations have validated a generic model that predicts the conditions for strong field amplifications in cells at TTFields frequencies, allowing further elaboration of the mechanism of action. Our model fits well with other analytical models; the furrow model, cells in confluence, or electroporation. Citation Format: Tal Marciano, Shay Levi, Eduard Fedorov, Zeev Bomzon. The distribution of Tumor Treating Fields is affected by cell confluence and pores in the membrane [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1435.

The Last Chance Saloon.

Accurate chromosome segregation requires the removal of all chromatin bridges, which link chromosomes before cell division. When chromatin bridges fail to be removed, cell cycle progression may halt, or cytokinesis failure and ensuing polyploidization may occur. Conversely, the inappropriate severing of chromatin bridges leads to chromosome fragmentation, excessive genome instability at breakpoints, micronucleus formation, and chromothripsis. In this mini-review, we first describe the origins of chromatin bridges, the toxic processing of chromatin bridges by mechanical force, and the TREX1 exonuclease. We then focus on the abscission checkpoint (NoCut) which can confer a transient delay in cytokinesis progression to facilitate bridge resolution. Finally, we describe a recently identified mechanism uncovered in C. elegans where the conserved midbody associated endonuclease LEM-3/ANKLE1 is able to resolve chromatin bridges generated by various perturbations of DNA metabolism at the final stage of cell division. We also discuss how LEM-3 dependent chromatin bridge resolution may be coordinated with abscission checkpoint (NoCut) to achieve an error-free cleavage, therefore acting as a “last chance saloon” to facilitate genome integrity and organismal survival.

Secreted midbody remnants are a class of extracellular vesicles molecularly distinct from exosomes and microparticles

During the final stages of cell division, newly-formed daughter cells remain connected by a thin intercellular bridge containing the midbody (MB), a microtubule-rich organelle responsible for cytokinetic abscission. Following cell division the MB is asymmetrically inherited by one daughter cell where it persists as a midbody remnant (MB-R). Accumulating evidence shows MB-Rs are secreted (sMB-Rs) into the extracellular medium and engulfed by neighbouring non-sister cells. While much is known about intracellular MB-Rs, sMB-Rs are poorly understood. Here, we report the large-scale purification and biochemical characterisation of sMB-Rs released from colon cancer cells, including profiling of their proteome using mass spectrometry. We show sMB-Rs are an abundant class of membrane-encapsulated extracellular vesicle (200-600 nm) enriched in core cytokinetic proteins and molecularly distinct from exosomes and microparticles. Functional dissection of sMB-Rs demonstrated that they are engulfed by, and accumulate in, quiescent fibroblasts where they promote cellular transformation and an invasive phenotype.

Allelopathic activities of three carotenoids, neoxanthin, crocin and β-carotene, assayed using protoplast co-culture method with digital image analysis.

Allelopathic activities of three carotenoids of a natural pigment group, neoxanthin, crocin and β-carotene, were assayed by the protoplast co-culture method with digital image analysis (DIA-PP method). Effects on three different growth stages of lettuce protoplasts, i.e., cell wall formation, cell division, and yellow pigment accumulation, were investigated using 96-well culture plates. Cell division was inhibited 65-95% by all three carotenoids at 33-100 µM. Inhibition of cell division stage was stronger than at the cell wall formation stage in neoxanthin, and the water-soluble carotenoid, crocin, whose yellow pigment was incorporated into the vacuole of lettuce protoplasts. Neoxanthin at 33 µM and crocin at higher than 100 µM inhibited more than 100% of the yellow pigment accumulation. By contrast, at low concentrations (0.01-1 µM) β-carotene stimulated growth at the cell division stage. At high concentrations of β-carotene (100-500 µM), inhibition was prominent at all three stages, and also in neighboring wells of zero control, which suggested emission of a volatile compound by β-carotene. They were compared with the report of the volatile compound, tulipalin A. Differences in patterns of inhibition of carotenoids on lettuce protoplast growth were compared with reports of another natural pigment, anthocyanin, and anthocyanin-containing red callus cultured in the light, and with that of neoxanthin-containing yellow callus cultured in the dark.

Linking Mineral Nutrition and Fruit Quality to Growth Intensity and Crop Load in Apple

Diagnostic of the nutritional status in fruit trees can be estimated using different models. However, these models usually do not fully match the character of the perennial crops and their accuracy needs to be improved. This study aimed at deepening the understanding of the aspects controlling mineral content of macro- and micronutrients and fruit quality of the apple trees with respect to the crop load and vegetative shoot growth. The focus was given on description of the nutrient content variability in time and space, its relation to important physiological events, and conditions altering their gradient in leaf and fruit. The evaluation was carried out with six years old ’Golden Delicious’ apple trees on M9, planted in an experimental orchard. Flowering rate, total yield, shoot growth intensity, mean fruit weight and quality along with mineral content of leaves, and fruit in different development stages were analysed according to intensity of natural or regulated flowering and fruit set, respectively. We found that aside of macroelements, iron, and manganese depended on the crop load. For nitrogen, phosphorus, and potassium content, their gradient increased up to 10–25% within the sufficiency range as the trees tried to maintain certain level of these nutrients in leaves and fruit simultaneously during deficit. In contrary, the higher content of Mg, Fe, and Mn related to the avoidance of deficit through up to 20–30% higher uptake in trees with higher crop load. Phosphorus, potassium, and calcium content in short fruiting shoots was more stage dependent, showing relation with the timing of cessation of particular meristematic growth activity. If no intense deficit occurred, the trees kept unrestricted initial investment of nutrients in the fruit during cell division stage whatever the crop load was. Their content at the later development stages appeared to be more dependent upon the consumption by particular sink. These findings allowed to improve the interpretation of the fruit trees nutrition status due to adaptation of the reference nutrient content in low cropping trees by a reduction of 5–10% N and P, 10–15% Ca, as well as by an increase of 15–20% K content in leaves at the stage with fruit size of 20 mm, and without change in P content at the stage of about 70% of final fruit size.

A centriole's subdistal appendages: contributions to cell division, ciliogenesis and differentiation.

The centrosome is a highly conserved structure composed of two centrioles surrounded by pericentriolar material. The mother, and inherently older, centriole has distal and subdistal appendages, whereas the daughter centriole is devoid of these appendage structures. Both appendages have been primarily linked to functions in cilia formation. However, subdistal appendages present with a variety of potential functions that include spindle placement, chromosome alignment, the final stage of cell division (abscission) and potentially cell differentiation. Subdistal appendages are particularly interesting in that they do not always display a conserved ninefold symmetry in appendage organization on the mother centriole across eukaryotic species, unlike distal appendages. In this review, we aim to differentiate both the morphology and role of the distal and subdistal appendages, with a particular focus on subdistal appendages.

Dinoroseobacter shibae Outer Membrane Vesicles Are Enriched for the Chromosome Dimer Resolution Site dif.

Outer membrane vesicles (OMVs) are universally produced by prokaryotes and play important roles in symbiotic and pathogenic interactions. They often contain DNA, but a mechanism for its incorporation is lacking. Here, we show that Dinoroseobacter shibae, a dinoflagellate symbiont, constitutively secretes OMVs containing DNA. Time-lapse microscopy captured instances of multiple OMV production at the septum during cell division. DNA from the vesicle lumen was up to 22-fold enriched for the region around the terminus of replication (ter). The peak of coverage was located at dif, a conserved 28-bp palindromic sequence required for binding of the site-specific tyrosine recombinases XerC/XerD. These enzymes are activated at the last stage of cell division immediately prior to septum formation when they are bound by the divisome protein FtsK. We suggest that overreplicated regions around the terminus have been repaired by the FtsK-dif-XerC/XerD molecular machinery. The vesicle proteome was clearly dominated by outer membrane and periplasmic proteins. Some of the most abundant vesicle membrane proteins were predicted to be required for direct interaction with peptidoglycan during cell division (LysM, Tol-Pal, Spol, lytic murein transglycosylase). OMVs were 15-fold enriched for the saturated fatty acid 16:00. We hypothesize that constitutive OMV secretion in D. shibae is coupled to cell division. The footprint of the FtsK-dif-XerC/XerD molecular machinery suggests a novel potentially highly conserved route for incorporation of DNA into OMVs. Clearing the division site from small DNA fragments might be an important function of vesicles produced during exponential growth under optimal conditions.IMPORTANCE Gram-negative bacteria continually form vesicles from their outer membrane (outer membrane vesicles [OMVs]) during normal growth. OMVs frequently contain DNA, and it is unclear how DNA can be shuffled from the cytoplasm to the OMVs. We studied OMV cargo in Dinoroseobacter shibae, a symbiont of dinoflagellates, using microscopy and a multi-omics approach. We found that vesicles formed during undisturbed exponential growth contain DNA which is enriched for genes around the replication terminus, specifically, the binding site for an enzyme complex that is activated at the last stage of cell division. We suggest that the enriched genes are the result of overreplication which is repaired by their excision and excretion via membrane vesicles to clear the divisome from waste DNA.

Strong Allelopathic Activities of Leaves and Cultured Cells of <i>Spiraea thunbergii</i> Assayed by the Protoplast Co-Culture Method with Digital Image Analysis: Evaluation of <i>Cis</i>- and <i>Trans</i>-Cinnamic Acid as Allelochemicals

The inhibitory allelopathic activities of leaves and leaf-origin suspension cultured cells of Spiraea thunbergii, and putative allelochemicals, cis- and trans-cinnamic acid, were investigated by the protoplast co-culture method with digital image analysis (DIA-PP method) using lettuce as a recipient. The optimal conditions of 2,4-dichlorophenoxyacetic acid (2,4-D) and benzyladenine (BA) for the cell division of S. thunbergii protoplasts were first examined using 50 μL liquid MS basal medium containing 3% sucrose, and 0.8 M mannitol in a 96-well culture plate. The hormonal condition for co-culture, 1 μM 2,4-D plus 0.1 μM BA, which was optimal for lettuce protoplast growth, was sub-optimal for S. thunbergii protoplasts. Effects of co-culture on the three stages of lettuce protoplast growth, i.e., cell wall formation, cell division, and yellow pigment accumulation, were examined. Protoplasts of leaf and suspension cells of S. thunbergii strongly inhibited lettuce protoplast growth at the cell division stage (100% inhibition at 80 - 100 × 103 mL-1), but not so much at the other two stages. Both cis- and trans-cinnamic acid, showed the strongest inhibition at the cell wall formation stage, and 100% inhibition at the cell division stage at 100 μM. These results were compared with those obtained in a lettuce seedling growth test, using different allelopathic plants, and their allelochemicals were studied by the DIA-PP method.

Application of Calcium Chloride at Different Phenological Stages Alleviates Chilling Injury and Delays Climacteric Ripening in Peach Fruit during Low-Temperature Storage

ABSTRACT Owing to the high degree of perishability of the fruit, the peach industry faces serious challenges tackling physiological disorders, microbial/chemical decay, and the suboptimal quality of the fruit during and following cold storage. The present study was aimed to investigate the effects of spray applications of calcium chloride (CaCl2) at various physiological stages of fruit growth and development, on ripening, flesh softening, chilling injury (CI) and nutritional quality of peach fruit cv “Flordaking” during low-temperature storage. The aqueous solutions of CaCl2 [0, 1, 2, and 3% (w/v)] were sprayed on peach trees during cell division, pit hardening and cell expansion stages in 2011 and 2012. At commercial harvest, the fruit were hand-picked and were subjected to cold storage at 1 ± 1°C (RH: 90 ± 5%) for 6 weeks. The data on CI index and other quality characteristics of the fruit were recorded at harvest and then weekly for up to 6 weeks. The results showed that the pre-harvest spray application of CaCl2 substantially reduced the incidence of chilling injury (CI) and delayed climacteric ripening seemingly by lowering the rates of ethylene production and fruit softening during storage as compared to control, regardless the doses of CaCl2 and its application time, however higher doses of calcium chloride proved to be toxic causing blemishes on the surface of “Flordaking” peaches. In conclusion, pre-harvest spray application of CaCl2 @ 1% may be a successful production management tool to maintain the quality and to extend storability of peach fruit during cold storage.

Organization of peptidoglycan synthesis in nodes and separate rings at different stages of cell division of Streptococcus pneumoniae.

Bacterial peptidoglycan (PG) synthesis requires strict spatiotemporal organization to reproduce specific cell shapes. In ovoid-shaped Streptococcus pneumoniae (Spn), septal and peripheral (elongation) PG synthesis occur simultaneously at midcell. To uncover the organization of proteins and activities that carry out these two modes of PG synthesis, we examined Spn cells vertically oriented onto their poles to image the division plane at the high lateral resolution of 3D-SIM (structured-illumination microscopy). Labeling with fluorescent D-amino acids (FDAA) showed that areas of new transpeptidase (TP) activity catalyzed by penicillin-binding proteins (PBPs) separate into a pair of concentric rings early in division, representing peripheral PG (pPG) synthesis (outer ring) and the leading-edge (inner ring) of septal PG (sPG) synthesis. Fluorescently tagged PBP2x or FtsZ locate primarily to the inner FDAA-marked ring, whereas PBP2b and FtsX remain in the outer ring, suggesting roles in sPG or pPG synthesis, respectively. Pulses of FDAA labeling revealed an arrangement of separate regularly spaced "nodes" of TP activity around the division site of predivisional cells. Tagged PBP2x, PBP2b, and FtsX proteins also exhibited nodal patterns with spacing comparable to that of FDAA labeling. Together, these results reveal new aspects of spatially ordered PG synthesis in ovococcal bacteria during cell division.

Influence of Lactic Acid on Cell Cycle Progressions in Lactobacillus bulgaricus During Batch Culture.

Lactic acid has been proved to inhibit the proliferation of lactic acid bacteria in the fermentation process. To shed light on the cell cycle alterations in acidic conditions, the cell division of Lactobacillus bulgaricus sp1.1 in batch culture was analyzed directly by implementing of the intracellular fluorescent tracking assay in different pH adjusted by lactic acid. Cell proliferation and cell division were investigated to be negatively controlled by the decrease of pH, and pH4.1 was the critical condition of downregulating cell division but retains cell culturability. The cell area and cell length in pH4.1 were examined by using fluorescent labeling, and they reduced to about 29.18-34.89% and 32.67-40% of cells cultured in the unacidified medium, respectively. The DNA replication initiation was undergoing prompted by the low extent of DNA condensation and higher expression of the dnaA gene in this critical pH. The results indicated that the cell cycle progressions of Lactobacillus bulgaricus sp1.1 in acidic conditions were arrested at intracellular biomass accumulation and cell division stage. These findings provide fundamental insight into cell cycle control of the acidic environment in Lactobacillus bulgaricus sp1.1.

Lotus japonicus Nuclear Factor YA1, a nodule emergence stage-specific regulator of auxin signalling.

Summary Organogenesis of legume root nodules begins with the nodulation factor‐dependent stimulation of compatible root cells to initiate divisions, signifying an early nodule primordium formation event. This is followed by cellular differentiation, including cell expansion and vascular bundle formation, and we previously showed that Lotus japonicus NF‐YA1 is essential for this process, presumably by regulating three members of the SHORT INTERNODES/STYLISH (STY) transcription factor gene family.In this study, we used combined genetics, genomics and cell biology approaches to characterize the role of STY genes during root nodule formation and to test a hypothesis that they mediate nodule development by stimulating auxin signalling.We show here that L. japonicus STYs are required for nodule emergence. This is attributed to the NF‐YA1‐dependent regulatory cascade, comprising STY genes and their downstream targets, YUCCA1 and YUCCA11, involved in a local auxin biosynthesis at the post‐initial cell division stage. An analogous NF‐YA1/STY regulatory module seems to operate in Medicago truncatula in association with the indeterminate nodule patterning.Our data define L. japonicus and M. truncatula NF‐YA1 genes as important nodule emergence stage‐specific regulators of auxin signalling while indicating that the inductive stage and subsequent formation of early nodule primordia are mediated through an independent mechanism(s).

Mitosis in Cancer Cell Increases Immune Resistance via High Expression of HLA-G and PD-L1.

Simple SummaryCancer results from “aggressive division” of cells, that is unchecked by the immune cells. Cancer cells express several proteins to evade the immune response. One of the main treatment against cancer is to target and block their cell division. We found that, the expression of these proteins is increased during the mitosis stage of cell division. Therefore the blockade of cell cycle in the mitotic phase may results in increased immune resistance that can further help the cancer cells in evading immune attack.Cancer is a result of “aggressive” division and uncontrolled proliferation of the abnormal cells that survive attack by immune cells. We investigated the expression of HLA-G and PD-L1 with the different stages of cancer cell division along with their role in the interaction of immune cells in vitro. Ovarian cancer (OVCAR-3) and chronic myeloid leukemia cell line (K-562) are used for this study. The correlation of protein expression with percentage of cells in each phase (G1, S and G2 phase) was evaluated through FACS. Cells were synchronized in G1, G2 and mitotic phase to evaluate gene (RT-qPCR) and protein expression (FACS). Real-time immune cell attack (RTICA) analysis with PBMCs (peripheral blood mono-nuclear cells) and cancer cells were performed. We found that cells expressing higher levels of HLA-G and PD-L1 are mainly in G2 phase and those expressing lower levels are mainly in G1 phase. Evidently, the higher expression of the two proteins was observed when synchronized in mitotic phase as compared to low expression when synchronized in G1 phase. RTICA analysis showed the presence of HLA-G delayed the lysis of the cells. In conclusion, the cancer cell can escape from immune cells in division stage that suggests the impact of mitosis index for cancer immunotherapy.

Divergent Impact of Actin Isoforms on Division of Epithelial Cells

We investigated distribution and functions of beta- and gamma-cytoplasmic actins (CYAs) at different stages of non-neoplastic epithelial cell division using laser scanning microscopy (LSM). Here, we demonstrated that beta- and gamma-CYAs are spatially segregated in the early prophase, anaphase, telophase, and cytokinesis. Small interfering RNA (siRNA) experiments revealed that in both beta-CYA- and gamma-CYA-depleted cells, the number of cells was significantly reduced compared with the siRNA controls. Beta-CYA depletion resulted in an enlargement of the cell area in metaphase and high percentage of polynuclear cells compared with the siRNA control, indicating a potential failure of cytokinesis. Gamma-CYA depletion resulted in a reduced percentage of mitotic cells. We also observed the interdependence between the actin isoforms and the microtubule system in mitosis: (i) a decrease in the gamma-CYA led to impaired mitotic spindle organization; (ii) suppression of tubulin polymerization caused impaired beta-CYA reorganization, as incubation with colcemid blocked the transfer of short beta-actin polymers from the basal to the cortical compartment. We conclude that both actin isoforms are essential for proper cell division, but each isoform has its own specific functional role in this process.