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Reproduction (Cambridge, England) Mar 2024The fertilizing spermatozoa induce a Ca2+ oscillatory pattern, the universal hallmark of oocyte activation, in all sexually reproducing animals. Assisted reproductive...
The fertilizing spermatozoa induce a Ca2+ oscillatory pattern, the universal hallmark of oocyte activation, in all sexually reproducing animals. Assisted reproductive technologies (ARTs) like intracytoplasmic sperm injection (ICSI) bypass the physiological pathway; however, while a normal Ca2+ release pattern occurs in some species, particularly humans, artificial activation is compulsory for ICSI-fertilized oocytes to develop in most farm animals. Unlike the normal oscillatory pattern, most artificial activation protocols induce a single Ca2+ spike, undermining proper ICSI-derived embryo development in these species. Curiously, diploid parthenogenetic embryos activated by the same treatments develop normally at high frequencies and implant upon transfer in the uterus. We hypothesized that, at least in ruminant embryos, the oscillatory calcium waves late in the first cell cycle target preferentially the paternal pronucleus and are fundamentally important for paternal nuclear remodeling. We believe that Ca2+ signaling is central to full totipotency deployment of the paternal genome. Research in this area could highlight the asymmetry between the parental genome reprogramming timing/mechanisms in early development and impact ARTs like ICSI and cloning.
Topics: Animals; Female; Male; Humans; Calcium; Semen; Cytoplasm; Fertilization; Spermatozoa; Oocytes
PubMed: 38112585
DOI: 10.1530/REP-23-0360 -
Cell Reports. Medicine Nov 2023High-level MET amplification (METamp) is a primary driver in ∼1%-2% of non-small cell lung cancers (NSCLCs). Cohort B of the phase 2 VISION trial evaluates tepotinib,...
High-level MET amplification (METamp) is a primary driver in ∼1%-2% of non-small cell lung cancers (NSCLCs). Cohort B of the phase 2 VISION trial evaluates tepotinib, an oral MET inhibitor, in patients with advanced NSCLC with high-level METamp who were enrolled by liquid biopsy. While the study was halted before the enrollment of the planned 60 patients, the results of 24 enrolled patients are presented here. The objective response rate (ORR) is 41.7% (95% confidence interval [CI], 22.1-63.4), and the median duration of response is 14.3 months (95% CI, 2.8-not estimable). In exploratory biomarker analyses, focal METamp, RB1 wild-type, MYC diploidy, low circulating tumor DNA (ctDNA) burden at baseline, and early molecular response are associated with better outcomes. Adverse events include edema (composite term; any grade: 58.3%; grade 3: 12.5%) and constipation (any grade: 41.7%; grade 3: 4.2%). Tepotinib provides antitumor activity in high-level METamp NSCLC (ClinicalTrials.gov: NCT02864992).
Topics: Humans; Carcinoma, Non-Small-Cell Lung; Lung Neoplasms; Pyrimidines; Liquid Biopsy
PubMed: 37944528
DOI: 10.1016/j.xcrm.2023.101280 -
BMC Plant Biology Nov 2023Stomatal variation, including guard cell (GC) density, size and chloroplast number, is often used to differentiate polyploids from diploids. However, few works have...
BACKGROUND
Stomatal variation, including guard cell (GC) density, size and chloroplast number, is often used to differentiate polyploids from diploids. However, few works have focused on stomatal variation with respect to polyploidization, especially for consecutively different ploidy levels within a plant species. For example, Allium tuberosum, which is mainly a tetraploid (2n = 4x = 32), is also found at other ploidy levels which have not been widely studied yet.
RESULTS
We recently found cultivars with different ploidy levels, including those that are diploid (2n = 2x = 16), triploid (2n = 3x = 24), pseudopentaploid (2n = 34-42, mostly 40) and pseudohexaploid (2n = 44-50, mostly 48). GCs were evaluated for their density, size (length and width) and chloroplast number. There was no correspondence between ploidy level and stomatal density, in which anisopolyploids (approximately 57 and 53 stomata/mm in triploid and pseudopentaploid, respectively) had a higher stomatal density than isopolyploids (approximately 36, 43, and 44 stomata/mm in diploid, tetraploid and pseudohexaploid, respectively). There was a positive relationship between ploidy level and GC chloroplast number (approximately 44, 45, 51, 72 and 90 in diploid to pseudohexaploid, respectively). GC length and width also increased with ploidy level. However, the length increased approximately 1.22 times faster than the width during polyploidization.
CONCLUSIONS
This study shows that GC size increased with increasing DNA content, but the rate of increase differed between length and width. In the process of polyploidization, plants evolved longer and narrower stomata with more chloroplasts in the GCs.
Topics: Chive; Ploidies; Tetraploidy; Triploidy; Plant Stomata
PubMed: 38017401
DOI: 10.1186/s12870-023-04615-y -
American Journal of Physiology. Cell... Oct 2023Polyploidization of tubular cells (TC) is triggered by acute kidney injury (AKI) to allow survival in the early phase after AKI, but in the long run promotes fibrosis...
Polyploidization of tubular cells (TC) is triggered by acute kidney injury (AKI) to allow survival in the early phase after AKI, but in the long run promotes fibrosis and AKI-chronic kidney disease (CKD) transition. The molecular mechanism governing the link between polyploid TC and kidney fibrosis remains to be clarified. In this study, we demonstrate that immediately after AKI, expression of cell cycle markers mostly identifies a population of DNA-damaged polyploid TC. Using transgenic mouse models and single-cell RNA sequencing we show that, unlike diploid TC, polyploid TC accumulate DNA damage and survive, eventually resting in the G1 phase of the cell cycle. In vivo and in vitro single-cell RNA sequencing along with sorting of polyploid TC shows that these cells acquire a profibrotic phenotype culminating in transforming growth factor (TGF)-β1 expression and that TGF-β1 directly promotes polyploidization. This demonstrates that TC polyploidization is a self-sustained mechanism. Interactome analysis by single-cell RNA sequencing revealed that TGF-β1 signaling fosters a reciprocal activation loop among polyploid TC, macrophages, and fibroblasts to sustain kidney fibrosis and promote CKD progression. Collectively, this study contributes to the ongoing revision of the paradigm of kidney tubule response to AKI, supporting the existence of a tubulointerstitial cross talk mediated by TGF-β1 signaling produced by polyploid TC following DNA damage. Polyploidization in tubular epithelial cells has been neglected until recently. Here, we showed that polyploidization is a self-sustained mechanism that plays an important role during chronic kidney disease development, proving the existence of a cross talk between infiltrating cells and polyploid tubular cells. This study contributes to the ongoing revision of kidney adaptation to injury, posing polyploid tubular cells at the center of the process.
Topics: Animals; Mice; Transforming Growth Factor beta1; Acute Kidney Injury; Epithelial Cells; Polyploidy; Fibrosis
PubMed: 37642236
DOI: 10.1152/ajpcell.00081.2023 -
BMC Plant Biology Nov 2023Artificial induction of polyploidy is the most common and effective way to improve the biological properties of Populus and develop new varieties of this tree. In this...
BACKGROUND
Artificial induction of polyploidy is the most common and effective way to improve the biological properties of Populus and develop new varieties of this tree. In this study, in order to confirm and expand earlier findings, we established a protocol using colchicine and based on an efficient shoot regeneration system of leaf blades to induce tetraploidy in vitro in three genotypes from diploid Populus hopeiensis. The stomatal characteristics, leaf blade size, and growth were evaluated for diploids and tetraploids of three genotypes.
RESULTS
We found that genotype, preculture duration, colchicine concentration, and colchicine exposure time had highly significant effects on the tetraploid induction rate. The optimal protocol for inducing tetraploidy in P. hopeiensis was to preculture leaf blades for 7 days and then treat them for 4 days with 40 mg/L colchicine. The tetraploid induction rates of genotypes BT1, BT3, and BT8 were 21.2, 11.4 and 16.7%, respectively. A total of 136 tetraploids were identified by flow cytometry analysis and somatic chromosome counting. The stomatal length, width, and density of leaf blades significantly differed between diploid and tetraploid plants. Compared with their diploid counterparts, the tetraploids produced larger leaf blades and had a slower growth rate. Our findings further document the modified morphological characteristics of P. hopeiensis following whole-genome duplication (e.g., induced tetraploidy).
CONCLUSIONS
We established a protocol for in vitro induction of tetraploidy from diploid leaf blades treated with colchicine, which can be applied to different genotypes of P. hopeiensis.
Topics: Tetraploidy; Populus; Polyploidy; Diploidy; Biological Variation, Population; Colchicine
PubMed: 37957587
DOI: 10.1186/s12870-023-04578-0 -
Journal of Fungi (Basel, Switzerland) Feb 2024In the budding yeast , sporulation occurs during starvation of a diploid cell and results in the formation of four haploid spores forming within the mother cell ascus.... (Review)
Review
In the budding yeast , sporulation occurs during starvation of a diploid cell and results in the formation of four haploid spores forming within the mother cell ascus. Meiosis divides the genetic material that is encapsulated by the prospore membrane that grows to surround the haploid nuclei; this membrane will eventually become the plasma membrane of the haploid spore. Cellularization of the spores occurs when the prospore membrane closes to capture the haploid nucleus along with some cytoplasmic material from the mother cell, and thus, closure of the prospore membrane is the meiotic cytokinetic event. This cytokinetic event involves the removal of the leading-edge protein complex, a complex of proteins that localizes to the leading edge of the growing prospore membrane. The development and closure of the prospore membrane must be coordinated with other meiotic exit events such as spindle disassembly. Timing of the closure of the prospore membrane depends on the meiotic exit pathway, which utilizes Cdc15, a Hippo-like kinase, and Sps1, an STE20 family GCKIII kinase, acting in parallel to the E3 ligase Ama1-APC/C. This review describes the sporulation process and focuses on the development of the prospore membrane and the regulation of prospore membrane closure.
PubMed: 38392804
DOI: 10.3390/jof10020132 -
BioRxiv : the Preprint Server For... Nov 2023Cardiac regeneration in newborn rodents depends on the ability of pre-existing cardiomyocytes to proliferate and divide. This capacity is lost within the first week of...
Cardiac regeneration in newborn rodents depends on the ability of pre-existing cardiomyocytes to proliferate and divide. This capacity is lost within the first week of postnatal development when these cells rapidly switch from hyperplasia to hypertrophy, withdraw from the cell cycle, become binucleated, and increase in size. How these dynamic changes in size and ploidy impact cardiomyocyte proliferative potential is not well understood. In this study, we innovate the application of a commercially available digital holographic imaging microscope, the Holomonitor M4, to evaluate the proliferative responses of mononucleated diploid and binucleated tetraploid cardiomyocytes. This instrument coupled with the powerful Holomonitor App Suite software enables long-term label-free quantitative three-dimensional tracking of primary cardiomyocyte dynamics in real-time with single-cell resolution. Our digital holographic imaging results provide direct evidence that mononucleated cardiomyocytes retain significant proliferative potential as most can successfully divide with high frequency. In contrast, binucleated cardiomyocytes exhibit a blunted response to a proliferative stimulus with the majority not attempting to divide at all. Nevertheless, some binucleated cardiomyocytes were capable of complete division, suggesting that these cells still do retain limited proliferative capacity. By quantitatively tracking cardiomyocyte volume dynamics during these proliferative responses, we reveal that both mononucleated and binucleated cells reach a unique size threshold prior to attempted cell division. The absolute threshold is increased by binucleation, which may limit the ability of binucleated cardiomyocytes to divide. By defining the interrelationship between cardiomyocyte size, ploidy, and cell cycle control, we will better understand the cellular mechanisms that drive the loss of mammalian cardiac regenerative capacity after birth.
PubMed: 37961676
DOI: 10.1101/2023.11.02.565407 -
Cancer Reports (Hoboken, N.J.) Dec 2023Prostate cancer is morphologically and molecularly heterogeneous. Genomic heterogeneity might be mirrored by variability in DNA ploidy. Aneuploidy is a hallmark of...
BACKGROUND
Prostate cancer is morphologically and molecularly heterogeneous. Genomic heterogeneity might be mirrored by variability in DNA ploidy. Aneuploidy is a hallmark of genomic instability and associated with tumor aggressiveness. Little attention has been paid to the biological significance of the diploid tumor cell population that often coexists with aneuploid populations. Here, we investigated the role of DNA ploidy in tumor heterogeneity and clonal evolution.
METHODS
Three radical prostatectomy specimens with intratumoral heterogeneity based on nuclear features on H&E were selected. DNA content of each subpopulation was determined by DNA image cytometry and silver in situ hybridization (SISH). Genomic evolution was inferred from array comparative genomic hybridization (aCGH). Additionally, immunohistochemistry was used to examine the stemness-associated marker ALDH1A1.
RESULTS
Nuclear morphology reliably predicted DNA ploidy status in all three cases. In one case, aCGH analysis revealed several shared deletions and one amplification in both the diploid and the aneuploid population, suggesting that these populations could be related. In the other two cases, a statement about relatedness was not possible. Furthermore, ALDH1A1 was expressed in 2/3 cases and exclusively observed in their diploid populations.
CONCLUSIONS
In this proof-of-concept study, we demonstrate the feasibility to predict the DNA ploidy status of distinct populations within one tumor by H&E morphology. Future studies are needed to further investigate the clonal relationship between the diploid and the aneuploid subpopulation and test the hypothesis that the aneuploid population is derived from the diploid one. Finally, our analyses pointed to an enrichment of the stemness-associated marker ALDH1A1 in diploid populations, which warrants further investigation in future studies.
PubMed: 38148577
DOI: 10.1002/cnr2.1953 -
BMC Cancer Nov 2023Fringe is a glycosyltransferase involved in tumor occurrence and metastasis. However, a comprehensive analysis of the Fringe family members lunatic fringe (LFNG), manic...
BACKGROUND
Fringe is a glycosyltransferase involved in tumor occurrence and metastasis. However, a comprehensive analysis of the Fringe family members lunatic fringe (LFNG), manic fringe (MFNG), radical fringe (RFNG) in human cancers is lacking.
METHODS
In this study, we performed a pan-cancer analysis of Fringe family members in 33 cancer types with transcriptomic, genomic, methylation data from The Cancer Genome Atlas (TCGA) project. The correlation between Fringe family member expression and patient overall survival, copy number variation, methylation, Gene Ontology enrichment, and tumor-infiltrating lymphocytes (TILs) was investigated by using multiple databases, such as cBioPortal, Human Protein Atlas, GeneCards, STRING, MSigDB, TISIDB, and TIMER2. In vitro experiments and immunohistochemical assays were performed to validate our findings.
RESULTS
High expression levels of LFNG, MFNG, RFNG were closely associated with poor overall survival in multiple cancers, particularly in pancreatic adenocarcinoma (PAAD), uveal melanoma (UVM), and brain lower-grade glioma (LGG). Copy number variation analysis revealed that diploid and gain mutations of LFNG was significantly increased in PAAD and stomach adenocarcinoma (STAD), and significantly associated with the methylation levels in promoter regions. Significant differential genes between high and low expression groups of these Fringe family members were found to be consistently enriched in immune response and T cell activation pathway, extracellular matrix adhesion pathway, RNA splicing and ion transport pathways. Correlation between the abundance of tumor-infiltrating lymphocytes (TILs) and LFNG, MFNG, and RFNG expression showed that high LFNG expression was associated with lower TIL levels, particularly in PAAD. In vitro experiment by using pancreatic cancer PANC1 cells showed that LFNG overexpression promoted cell proliferation and invasion. Immunohistochemical assay in 90 PAAD patients verified the expression level of LFNG and its relationship with the prognosis.
CONCLUSIONS
Our study provides a relatively comprehensive understanding of the expression, mutation, copy number, promoter methylation level changes along with prognosis values of LFNG, MFNG, and RFNG in different tumors. High LFNG expression may serve as a poor prognosis molecular marker for PAAD.
Topics: Humans; Proteins; Adenocarcinoma; DNA Copy Number Variations; Pancreatic Neoplasms; Prognosis; Tumor Microenvironment
PubMed: 37932706
DOI: 10.1186/s12885-023-11545-3 -
Nature Biotechnology Oct 2023The Telomere-to-Telomere consortium recently assembled the first truly complete sequence of a human genome. To resolve the most complex repeats, this project relied on...
The Telomere-to-Telomere consortium recently assembled the first truly complete sequence of a human genome. To resolve the most complex repeats, this project relied on manual integration of ultra-long Oxford Nanopore sequencing reads with a high-resolution assembly graph built from long, accurate PacBio high-fidelity reads. We have improved and automated this strategy in Verkko, an iterative, graph-based pipeline for assembling complete, diploid genomes. Verkko begins with a multiplex de Bruijn graph built from long, accurate reads and progressively simplifies this graph by integrating ultra-long reads and haplotype-specific markers. The result is a phased, diploid assembly of both haplotypes, with many chromosomes automatically assembled from telomere to telomere. Running Verkko on the HG002 human genome resulted in 20 of 46 diploid chromosomes assembled without gaps at 99.9997% accuracy. The complete assembly of diploid genomes is a critical step towards the construction of comprehensive pangenome databases and chromosome-scale comparative genomics.
Topics: Humans; Sequence Analysis, DNA; Diploidy; Genomics; Genome, Human; Telomere; High-Throughput Nucleotide Sequencing
PubMed: 36797493
DOI: 10.1038/s41587-023-01662-6