Stages Of Cell Division Research Articles

The Isolation and Characterization of Perlucin in Pacific Abalone, Haliotis discus hannai: A Shell Morphogenic Protein with Potential Responses to Thermal Stress and Starvation.

Perlucin is a shell matrix protein that plays a significant role in regulating shell biomineralization. This study aimed to isolate and characterize the perlucin gene and analyze its expression to explore its role in shell formation, regeneration, and responses to thermal stress and starvation in Pacific abalone. The isolated full-length cDNA sequence of Hdh-Perlucin is 1002 bp long, encoding a 163-amino-acid polypeptide with a signal peptide. The mature peptide of Hdh-Perlucin contains a C-type lectin domain with signature motif and six conserved cysteine residues. Gene Ontology analysis suggests that Hdh-Perlucin exhibits carbohydrate-binding activity. Significantly higher expression of Hdh-Perlucin was observed during the juvenile, veliger, and trochophore stages, compared with cell division stage during early development. Upregulated expression was recorded from slow to rapid growth phases and during shell biomineralization, while downregulated expression was noted during starvation. Under thermal stress, expression peaked at 30 °C and 25 °C for 6 and 12 h, respectively, while consistently higher levels were observed at 15 °C throughout the experiment. This study provides the first comprehensive structural and expression analysis of Hdh-Perlucin, highlighting its roles in metamorphosis, shell formation and regeneration, and responses to heat stress and starvation in abalone.

Role of the antiparallel double-stranded filament form of FtsA in activating the Escherichia coli divisome.

The actin-like FtsA protein is essential for function of the cell division machinery, or divisome, in many bacteria including Escherichia coli. Previous in vitro studies demonstrated that purified wild-type FtsA assembles into closed mini-rings on lipid membranes, but oligomeric variants of FtsA such as FtsAR286W and FtsAG50E can bypass certain divisome defects and form arc and double-stranded (DS) oligomeric states, respectively, which may reflect conversion of an inactive to an active form of FtsA. However, it remains unproven which oligomeric forms of FtsA are responsible for assembling and activating the divisome. Here, we used an in vivo crosslinking assay for FtsA DS filaments to show that they largely depend on proper divisome assembly and are prevalent at later stages of cell division. We also used a previously reported variant that fails to assemble DS filaments, FtsAM96E R153D, to investigate the roles of FtsA oligomeric states in divisome assembly and activation. We show that FtsAM96E R153D cannot form DS filaments in vivo, fails to replace native FtsA, and confers a dominant negative phenotype, underscoring the importance of the DS filament stage for FtsA function. Surprisingly, however, activation of the divisome through the ftsL* or ftsW* superfission alleles suppressed the dominant negative phenotype and rescued the functionality of FtsAM96E R153D. Our results suggest that FtsA DS filaments are needed for divisome activation once it is assembled, but they are not essential for divisome assembly or guiding septum synthesis.IMPORTANCECell division is fundamental for cellular duplication. In simple cells like Escherichia coli bacteria, the actin homolog FtsA is essential for cell division and assembles into a variety of protein filaments at the cytoplasmic membrane. These filaments not only help tether polymers of the tubulin-like FtsZ to the membrane at early stages of cell division but also play crucial roles in recruiting other cell division proteins to a complex called the divisome. Once assembled, the E. coli divisome subsequently activates synthesis of the division septum that splits the cell in two. One recently discovered oligomeric conformation of FtsA is an antiparallel double-stranded filament. Using a combination of in vivo crosslinking and genetics, we provide evidence suggesting that these FtsA double filaments have a crucial role in activating the septum synthesis enzymes.

Cytokinetic abscission in Toxoplasma gondii is governed by protein phosphatase 2A and the daughter cell scaffold complex

Cytokinetic abscission marks the final stage of cell division, during which the daughter cells physically separate through the generation of new barriers, such as the plasma membrane or cell wall. While the contractile ring plays a central role during cytokinesis in bacteria, fungi and animal cells, the process diverges in Apicomplexa. In Toxoplasma gondii, two daughter cells are formed within the mother cell by endodyogeny. The mechanism by which the progeny cells acquire their plasma membrane during the disassembly of the mother cell, allowing daughter cells to emerge, remains unknown. Here we identify and characterize five T. gondii proteins, including three protein phosphatase 2A subunits, which exhibit a distinct and dynamic localization pattern during parasite division. Individual downregulation of these proteins prevents the accumulation of plasma membrane at the division plane, preventing the completion of cellular abscission. Remarkably, the absence of cytokinetic abscission does not hinder the completion of subsequent division cycles. The resulting progeny are able to egress from the infected cells but fail to glide and invade, except in cases of conjoined twin parasites.

Role of microRNAs in triple‑negative breast cancer and new therapeutic concepts (Review).

Breast cancer has surpassed lung cancer as the most prevalent malignancy affecting women worldwide. Triple-negative breast cancer (TNBC) is the type of breast cancer with the worst prognosis. As a heterogeneous disease, TNBC has a pathogenesis that involves multiple oncogenic pathways, including involvement of gene mutations and alterations in signaling pathways. MicroRNAs (miRNAs) are small endogenous, single-stranded non-coding RNAs that bind to the 3' untranslated region of target cell mRNAs to negatively regulate the gene expression of these specific mRNAs. Therefore, miRNAs are involved in cell growth, development, division and differentiation stages. miRNAs are also involved in gene targeting in tumorigenesis, tumor growth and the regulation of metastasis, including in breast cancer. Meanwhile, miRNAs also regulate components of signaling pathways. In this review, the role of miRNAs in the TNBC signaling pathway discovered in recent years is described in detail. The new concept of bi-targeted therapy for breast cancer using miRNA and artificial intelligence is also discussed.

Role of the antiparallel double-stranded filament form of FtsA in activating the Escherichia coli divisome.

The actin-like FtsA protein is essential for function of the cell division machinery, or divisome, in many bacteria including Escherichia coli. Previous in vitro studies demonstrated that purified wild-type FtsA assembles into closed mini-rings on lipid membranes, but oligomeric variants of FtsA such as FtsAR286W and FtsAG50E can bypass certain divisome defects and form arc and double-stranded (DS) oligomeric states, respectively, which may reflect conversion of an inactive to an active form of FtsA. Yet, it remains unproven which oligomeric forms of FtsA are responsible for assembling and activating the divisome. Here we used an in vivo crosslinking assay for FtsA DS filaments to show that they largely depend on proper divisome assembly and are prevalent at later stages of cell division. We also used a previously reported variant that fails to assemble DS filaments, FtsAM96E R153D, to investigate the roles of FtsA oligomeric states in divisome assembly and activation. We show that FtsAM96E R153D cannot form DS filaments in vivo, fails to replace native FtsA, and confers a dominant negative phenotype, underscoring the importance of the DS filament stage for FtsA function. Surprisingly, however, activation of the divisome through the ftsL* or ftsW* superfission alleles suppressed the dominant negative phenotype and rescued the functionallity of FtsAM96E R153D. Our results suggest that FtsA DS filaments are needed for divisome activation once it is assembled, but they are not essential for divisome assembly or guiding septum synthesis.

De novo TANGLED1 recruitment from the phragmoplast to aberrant cell plate fusion sites in maize.

Division plane positioning is crucial for proper growth and development in many organisms. In plants, the division plane is established before mitosis, by accumulation of a cytoskeletal structure called the preprophase band (PPB). The PPB is thought to be essential for recruitment of division site-localized proteins, which remain at the division site after the PPB disassembles. Here, we show that the division site-localized protein TANGLED1 (TAN1) is recruited independently of the PPB to the cell cortex by the plant cytokinetic machinery, the phragmoplast, from experiments using both the PPB-defective mutant discordia1 (dcd1) and chemical treatments that disrupt the phragmoplast in maize. TAN1 recruitment to de novo sites on the cortex is partially dependent on intact actin filaments and the myosin XI motor protein OPAQUE1 (O1). These data imply a yet unknown role for TAN1 and possibly other division site-localized proteins during the last stages of cell division when the phragmoplast touches the cell cortex to complete cytokinesis.

The thrombin receptor (PAR1) is associated with microtubules, mitosis and process formation in glioma cells

The cell surface protease-activated receptor 1 (PAR1) is overexpressed in glioblastoma multiforme (GBM). We studied the function and structure of intracellular microtubule (MT) and PAR1 in a tubulin-mediated process. We found that exposure to thrombin increased the percentage of proliferative, S, and M phases cells, affected morphology, and increased process elongation. PAR1 antagonist inversely affects these measures, increases tubulin end-binding protein 3 (EB3) mRNA expression in C6 cells, and reduces EB3 comet length, track length, and duration in neuroblastoma cells. In addition, immunofluorescence staining suggests that PAR1 is in close association with the MT α-tubulin and with coagulation cascade proteins during cell division stages. Our findings support PAR1 involvement in MT dynamics.

The remnant of the midbody as a cellular signaling mechanism

Cell signaling mechanisms are the basis for intercellular integration and regulation of proliferation and differentiation processes at the systemic level. One of the most plausible ways to control cell-to-cell interaction and targeted distribution of genetic information is for cells to use their own structures that are formed during mitosis and carry RNA-dependent signaling molecules that affect the mechanisms of control of intercellular interaction, cell proliferation and differentiation. The midbody remnant is a microtubule-rich structure that forms between dividing cells in the last stages of cytokinesis. Previously, it was thought to be only a temporary structure of the intercellular bridge during cytokinesis, which served to connect two future daughter cells. This structure is a key regulator of abscission and functions as a signaling platform that coordinates the cytoskeleton and endosomal dynamics during the terminal stages of cell division. The midbody is a subcellular structure that is formed during cell division, during penetration into the cleavage sulcus, when the microtubules of the central spindle are compacted and cross-linked by a thin intracellular bridge connecting the two daughter cells. The midbody plays a key role in organizing cytokinesis by recruiting a variety of mitotic kinases such as Aurora B and Plk1, as well as sulcus endosomes containing Rab11/FIP3, the membrane-rupturing ESCRT complex and the microtubule-rupturing enzyme spastin, all of which are responsible for mediated rupture during the later stages of cytokinesis. The midbodies can serve as extracellular and intracellular polarity signals during early embryogenesis, as well as during epithelialization and polarization of neurons. The molecular mechanism that governs the positioning of the middle body and how it transmits signals to neurons during differentiation or epithelium remains unknown. Importantly, the remains of the middle bodies can also function as intracellular signaling scaffolds that regulate proliferation and fate postmitotic cells. Since these structures can be released outside cells and taken up by other non-mitotic cells, it is suggested that they may function as vehicles for alternative transmission of complex sets of signaling molecules and/or receptors between cells, thus profoundly affecting signaling in general.

Genotoxicity of Gamma Radiation Against Lymphocytes of Radiation Workers: The Cytokinesis-Block Micronucleus Assay.

<b>Background and Objective:</b> Gamma irradiation induces genotoxicity, characterized by the formation of extra-nuclear bodies and left behind during the anaphase stage of cell division, often referred to as a micronucleus (MN). The present work aims to monitor exposure to ionizing radiation as a genotoxic agent in the lymphocytes of workers at radiation energy centers. <b>Materials and Methods:</b> The lymphocyte cytokinesis block micronucleus assay used and analyzed the correlation between the Nuclear Division Index (NDI), age, blood type and the number of micronuclei (MN). Blood samples were collected from 20 volunteers in heparin tubes, exposed to 2 Gy gamma rays and cultured <i>in vitro</i>. <b>Results:</b> A significant difference in the number of micronuclei between blood group A and blood groups A, B and AB. The Nuclear Division Index (NDI) value for lymphocytes of radiation energy center workers after gamma radiation was significant (1.74±0.1) but still within the normal range. Neither MN frequency nor NDI values correlated with age, but MN frequency showed a correlation with blood type. <b>Conclusion:</b> The gamma irradiation did not induce a cytostatic effect but proved genotoxic to the lymphocytes of radiation energy center workers. Notably, blood type A demonstrated higher sensitivity to gamma radiation.

The Neuropeptide HGAP Regulates Growth, Reproduction, Metamorphosis, Tissue Damage Repair, and Response against Starvation in Pacific Abalone

Introduction: Neuropeptides regulate vital physiological processes in multicellular organisms, including growth, reproduction, metamorphosis, and feeding. Recent transcriptome analyses have revealed neuropeptide genes with potential roles in vertebrate and invertebrate growth and reproduction. Among these genes, haliotid growth-associated peptide (HGAP) was identified as a novel gene in abalone. Methods: This study focused on HGAP in Pacific abalone (Haliotis discus hannai), where the complete cDNA sequence named Hdh-HGAP was identified and characterized. Samples from different experiments, such as metamorphosis, juvenile abalone growth, gonad development stages, muscle remodeling, and starvation, were collected for mRNA expression analysis. Results: The sequence spans 552 bp, encoding 96 amino acids with a molecular weight of 10.96 kDa. Expression analysis revealed that Hdh-HGAP exhibited higher levels in muscle tissue. Notably, during metamorphosis, Hdh-HGAP exhibited greater expression in the trochophore, veliger, and juvenile stages than in the cell division stages. Regarding growth patterns, Hdh-HGAP was highly expressed during rapid growth compared to stunted, minimal, and normal growth. In gonadal development, Hdh-HGAP mRNA reached its highest expression level during the ripening stage, indicating a potential role in gonadal cell proliferation and maturation. The in vivo effects of GnRH on gonad development and the expression of the Hdh-HGAP neuropeptide indicate its involvement in regulating reproduction in Pacific abalone. While tissue remodeling is primarily governed by immune genes, Hdh-HGAP was also upregulated during muscle tissue remodeling. Conversely, Hdh-HGAP was downregulated during prolonged starvation. Conclusion: This study marks the first comprehensive exploration of the Hdh-HGAP neuropeptide gene in Pacific abalone, shedding light on its involvement in growth, reproduction, metamorphosis, tissue remodeling, and response to starvation, although regulatory mechanisms are mostly unknown.

Testing effects of vapor pressure deficit on fruit growth: a comparative approach using peach, mango, olive, orange, and loquat.

Determining the influence of vapor pressure deficit (VPD) on fruit growth is a key issue under a changing climate scenario. Using a comparative approach across different fruit tree species may provide solid indications of common or contrasting plant responses to environmental factors. Knowing fruit growth responses to VPD may also be useful to optimize horticultural management practices under specific atmospheric conditions. Climate data to calculate VPD and fruit relative growth rates (RGR) by fruit gauges were monitored in peach at cell division, pit hardening and cell expansion stages; in two mango cultivars at cell division, cell expansion and maturation stages; in two olive cultivars, either full irrigated or rainfed, at early and late cell expansion stages; in 'Valencia' orange at early and late cell division stage, before and after mature fruit harvest; in loquat at cell expansion and maturation stages. At the fruit cell division stage, sensitivity of fruit growth to VPD seems to vary with species, time, and probably soil and atmospheric water deficit. 'Keitt' mango and 'Valencia' orange fruit growth responded to VPD in opposite ways, and this could be due to very different time of the year and VPD levels in the monitoring periods of the two species. At pit hardening stage of peach fruit growth, a relatively weak relationship was observed between VPD and RGR, and this is not surprising as fruit growth in size at this stage slows down significantly. A consistent and marked negative relationship between VPD and RGR was observed at cell expansion stage, when fruit growth is directly depending on water intake driving cell turgor. Another behavior common to all observed species was the gradual loss of relationship between VPD and RGR at the onset of fruit maturation, when fruit growth in size is generally programmed to stop. Finally, regardless of fruit type, VPD may have a significant effect on fruit growth and could be a useful parameter to be monitored for tree water management mainly when the cell expansion process prevails during fruit growth.

#25 : The Prediction Value of EEVA in the Timelapse GERI Incubator for the Blastocyst Morphology

Background and Aims: Time-lapse monitoring of embryo development may represent a superior way to culture and select embryos in vitro. The EEVA Test records the development of each embryo with a cell-tracking system and predicts the potential (High, Medium or Low) that an embryo will form a blastocyst based on automated detection and analysis of time-lapse imaging information of early cell division stage. The blastocyst grade will help clinicians in selecting the best embryos for transfer to increase pregnancy rates and reduce multiple pregnancies. The aim of this study is to determine the correlation between EEVA score and some blastocyst morphological parameters. Methods: Retrospective analysis study on 2522 embryos with indication for prolonging embryo culture to the blastocyst stage in time-lapse GERI incubator from January 2020 to June 2022. Results: Embryos with E1 and E2 score had significant higher morphological ratios of ICM A, ICM B, TE A, TE B, blastocoele expansion ratios of BL5,6 and lower morphological ratios of ICM D, TE D,compared with embryos having E3, E4, E5 score, p<0.01. Embryos with E3 score had significant higher morphological ratios of ICM A, ICM B, TE A, TE B, blastocoele expansion ratios of BL5,6 and lower morphological ratios of ICM D, TE D, compared with embryos having E4, E5 score, p<0.01. Conclusions: The blastocyst morphological parameters were associcated with the EEVA score. The rate of blastocyst with the ICM A, ICM B, TE A, TE B, BL 5,6 tended to decrease and the rate of blastocyst with ICM D; TE D tended to increase for embryos with low EEVA score. In other words, the good quality blastocyst rate tended to decrease, and the bad quality blastocyst rate tended to increase for embryos with low EEVA score.

Dynamin A as a one-component division machinery for synthetic cells.

Membrane abscission, the final cut of the last connection between emerging daughter cells, is an indispensable event in the last stage of cell division and in other cellular processes such as endocytosis, virus release or bacterial sporulation. However, its mechanism remains poorly understood, impeding its application as a cell-division machinery for synthetic cells. Here we use fluorescence microscopy and fluorescence recovery after photobleaching measurements to study the in vitro reconstitution of the bacterial protein dynamin A inside liposomes. Upon external reshaping of the liposomes into dumbbells, dynamin A self-assembles at the membrane neck, resulting in membrane hemi-scission and even full scission. Dynamin A proteins constitute a simple one-component division machinery capable of splitting dumbbell-shaped liposomes, marking an important step towards building a synthetic cell.

Putrescine, in Combination with Gibberellic Acid and Salicylic Acid, Improves Date Palm Fruit Quality via Triggering Protein and Carbohydrate Accumulation and Enhancing Mineral, Amino Acid, Sugar, and Phytohormone Acquisition

AbstractPutrescine (Put), gibberellic acid (GA3), and salicylic acid (SA) are involved in improving fruit growth and development. This is the first study investigating the effect of 1 mM Put, 100 mg L-1 GA3, and 100 mg L-1 SA on Phoenix dactylifera, cv. Zaghloul fruits. The experiment was conducted in a completely randomized block design during two successive seasons and five treatments [control (distilled water spray), Put, Put + GA3, Put + SA, and Put + GA3 + SA] were sprayed at the Hababouk (cell division) stage and Kimri [unripe green (cell elongation)] stage. Our results showed that all treatments significantly improved the yield and quality of ‘Zaghloul’ fruit by increasing the dry matter, crude fiber, ash, total soluble solids, (reducing, non-reducing, and total soluble) sugars, carbohydrate, protein, nitrogen, phosphorus, potassium, calcium, magnesium, sodium, zinc, iron, and manganese content as well as the peroxidase and catalase activity. Compared with the control treatment, exogenous applications also enhanced the amino acid (glutamic acid, aspartic acid, proline, glycine, alanine, arginine, cysteine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, serine, threonine, tyrosine, valine) acquisition and phytohormone (indole-3-acetic acid, cytokinins, GA3, SA) content, while decreasing the percentage of moisture, total acidity, total phenols, and tannins. The novel evidence indicates that among all treatments, application of diamine (Put), in combination with phytohormones (GA3 and SA), has the greatest effect on improving ‘Zaghloul’ fruit yield by up-regulating the nutrient acquisition, sugar accumulation, amino acid profile, antioxidant response, and phytohormone performance. These findings support the use of Put in conjunction with GA3 and SA to improve fruit yield and quality.

Microfluidic single-cell measurements of oxidative stress as a function of cell cycle position.

Single-cell measurements routinely demonstrate high levels of variation between cells, but fewer studies provide insight into the analytical and biological sources of this variation. This is particularly true of chemical cytometry, in which individual cells are lysed and their contents separated, compared to more established single-cell measurements of the genome and transcriptome. To characterize population-level variation and its sources, we analyzed oxidative stress levels in 1278 individual Dictyostelium discoideum cells as a function of exogenous stress level and cell cycle position. Cells were exposed to varying levels of oxidative stress via singlet oxygen generation using the photosensitizer Rose Bengal. Single-cell data reproduced the dose-response observed in ensemble measurements by CE-LIF, superimposed with high levels of heterogeneity. Through experiments and data analysis, we explored possible biological sources of this heterogeneity. No trend was observed between population variation and oxidative stress level, but cell cycle position was a major contributor to heterogeneity in oxidative stress. Cells synchronized to the same stage of cell division were less heterogeneous than unsynchronized cells (RSD of 37-51% vs 93%), and mitotic cells had higher levels of reactive oxygen species than interphase cells. While past research has proposed changes in cell size during the cell cycle as a source of biological noise, the measurements presented here use an internal standard to normalize for effects of cell volume, suggesting a more complex contribution of cell cycle to heterogeneity of oxidative stress.

In vitro Isolation and Characterization of Multipotent Postnatal Stem Cells from Human Dental Pulp: An Approach for Regeneration of Neural Crest Tissue.

Postnatal dental pulp tissues give the proper justification of the stem cell assimilation and characteristic of the multipotent of the stem cells. Researchers use an in vitro isolation process for clarifying the different stages of staining and cell division. Data collected from various sources helps in understanding how the stem cells help in tissue regeneration. It highlights the immunological phenotypes with the synthesis with cDNA for mentioning molecular immunology. Study also mentions the mitochondrial consistency to measure the potentiality regarding the immunology and the way it differs from 0 to 21 days. Researchers also mention the way for the future development by utilizing the key advantages and definite multipotent of the dental stem cells.

Strong Allelopathic Activity in Purple Leaves of Transgenic Spiraea cantoniensis Containing Cyanidin 3-Glucoside Assayed by the Protoplast Co-culture Method with Digital Image Analysis

The protoplast co-culture method with digital image analysis (DIA-PP method), was used for in vitro bioassay of allelopathic activity in the leaves of in vitro cultured plants of Spiraea cantoniensis. Purple leaves of transgenic S. cantoniensis, containing an anthocyanin, cyanidin 3-glucoside showed strong inhibitory allelopathic activity, 100% inhibition of recipient lettuce protoplast division by 80 × 103 mL-1 of co-cultured protoplasts, while green leaves of non-transgenic S. cantoniensis showed only 20% inhibition. Cyanidin 3-glucoside which inhibited lettuce protoplast division 80% at 100 µM was evaluated as an allelochemical of transgenic S. cantoniensis. Inhibition was stronger at the cell division stage of lettuce protoplast growth than that at the other stages, i.e., cell wall formation stage and yellow pigment accumulation stage. The results were discussed comparatively with the strong inhibitory activity of non-transgenic S. thunbergii and other allelopathic plant species containing different anthocyanin or carotenoid pigments as putative allelochemicals.

The restriction of calcium influx in metaphase and post-metaphase stages of cell division revealed by imaging secondary ion mass spectrometry (SIMS).

A secondary ion mass spectrometry (SIMS)-based isotopic imaging technique of ion microscopy was used for observing calcium influx in single renal epithelial LLC-PK1 cells. The CAMECA IMS-3f SIMS instrument, used in the study, is capable of producing isotopic images of single cells at 500nm spatial resolution. Due to the high-vacuum requirements of the instrument the cells were prepared cryogenically with a freeze-fracture method and frozen freeze-dried cells were used for SIMS analysis. The influx of calcium was imaged directly by exposure of cells to 44 Ca stable isotope in the extracellular buffer for 10min. The 44 Ca influx was measured at mass 44 and the distribution of endogenous calcium at mass 40 (40 Ca) in the same cell. A direct comparison of interphase cells to cells undergoing division revealed that calcium influx is restricted in metaphase and post-metaphase stages of cell division. This restriction is lifted in late cytokinesis. The net influx of 44 Ca in 10min was approximately half under calcium influx restriction in comparison to interphase cells. Under calcium influx restriction the 44 Ca concentration was the same between the mitotic chromosome and the cytoplasm. These observations indicate that the endoplasmic reticulum (ER) calcium uptake is compromised under calcium influx restriction in cells undergoing division.

Proliferative and preparative cell divisions in wing discs of the last larval instar are regulated by different hormones and determine the size and differentiation of the wing of Bombyx mori

Through investigating the two different enhanced cell division stages, we tried to clarify the switch from the growth to differentiation in the wing disc of the last larval instar of Bombyx mori. The response to insulin and 20E in vitro was stage specific. Bmmyc expression in V1 wing discs showed differences after being cultured with and without insulin. Bmmyc expression in V5 wing discs also showed differences after being cultured with and without 20E. Cell cycle–related genes, BmE2F1 and BmcycE, were upregulated with insulin or 20E in cultured wing discs of V1 or V5, respectively. Bmwnt1 and Bmras1 showed upregulation with 20E in cultured wing discs. Bmwnt1 showed upregulation with insulin in cultured wing discs, but Bmras1 did not show clear upregulation with insulin treatment. In contrast, Bmdpp showed upregulation with insulin, but did not show clear upregulation with 20E. The addition of PI3K or TOR inhibitors inhibited the upregulation of Bmmyc expression that was upregulated with insulin or 20E. The upregulation of Bmmyc and Bmwnt1 with insulin or 20E was inhibited with the addition of Myc or Wnt inhibitors, respectively. Genes related to matrix metalloprotease showed upregulation with 20E, and the upregulation was inhibited by the addition of Myc or Wnt inhibitors. From the present results, we concluded that cell division during the feeding stage occurred through PI3K/TOR cascade, and that at the wandering stage occurred through ecdysone and PI3K/TOR cascade; the former is for growth and the latter for differentiation.

Novel insights into molecular mechanisms of vegetative cell cycle and resting cyst formation in Apodileptus cf. visscheri (Alveolata, Ciliophora).

Ciliates usually with big cell sizes, complex morphological structures, and diverse life cycles, are good model organisms for studying cell proliferation regulation of eukaryotes. Up to date, the molecular regulation mechanisms for the vegetative cell cycle and encystment of these ciliates are poorly understood. Here, transcriptomes of Apodileptus cf. visscheri, which has an asexual vegetative cell cycle and is apt to encyst when environmental conditions become unfavorable, were sequenced to enrich our related knowledge. In this study, three replicates were sequenced for each of four cell stages, including initial period of growth, morphogenesis, cell division, and resting cyst. The significant transcription differences, involving cell cycle, biosynthesis, and energy metabolism pathways, were revealed between the resting cyst and vegetative cell cycle. Further investigations showed that the cell cycle pathway was enriched during morphogenesis stage and cell division stage. Compared to the initial period of growth stage, the differentially expressed genes involved in cellular components and molecular function were significantly enriched during cell division stage, while cellular components and biological processes were significantly enriched during morphogenesis stage. These provide novel insights into a comprehensive understanding at the molecular level of the survival and adaptive mechanism of unicellular eukaryotes.