102. CHARACTERISATION OF THE GTPase DYNAMIN THROUGHOUT MURINE SPERM MATURATION

2010 ◽  
Vol 22 (9) ◽  
pp. 20
Author(s):  
A. T. Reid ◽  
S. D. Roman ◽  
R. Aitken ◽  
B. Nixon
Keyword(s):  
Protein Trafficking ◽  
Molecular Mechanisms ◽  
Protein Translocation ◽  
Morphological Changes ◽  
Sperm Maturation ◽  
The Novel ◽  
Epididymal Maturation ◽  
Distinct Cell ◽  
Molecular Machinery

Throughout sperm maturation distinct remodelling events occur that imbue the cells with both the ability to bind the zona pellucida and undergo the acrosome reaction. Of long standing interest to our laboratory is the elucidation of the molecular mechanisms that underpin the attainment of these key functional attributes. This process begins with a complex range of morphological changes that accompany spermatogenesis, and is continued through post-testicular phases of maturation in both the male (epididymal maturation) and female (capacitation) reproductive tracts. However, among these changes only those occurring during the initial stages of spermatogenesis are intrinsically driven. The fact that the majority of sperm remodelling is extrinsically stimulated, and occurs in the absence of new protein synthesis, highlights the potential importance of processes such as intracellular protein trafficking. This has directed our focus towards the dynamin family of protein traffickers. The GTPase dynamin exists in three isoforms, namely Dnm1, Dnm2 and Dnm3 and is an integral part of the molecular machinery required for vesicle mediated protein translocation. Recent research from our laboratory has demonstrated the presence of these three isoforms in distinct, cell-specific locations during murine spermatogenesis. Immunofluorescence on mouse testis revealed that both Dnm1 and 2 are present within a region corresponding to the developing acrosome in maturing sperm, whilst Dnm3 appears to reside solely within pre-meiotic germ cells. Interestingly, Dnm1 and Dnm2 are both retained within the peri-acrosomal region of the sperm head in mature spermatozoa. Additionally, upon the induction of capacitation in vitro, staining for both Dnm1 and 2 becomes significantly reduced. Collectively these data support the novel hypothesis that dynamin not only participates in sperm remodelling events during spermatogenesis but may also have a previously unappreciated role in capacitation-associated priming of the sperm surface for interaction with the oocyte.

scholarly journals Platelet-derived lysophosphatidic acid mediated LPAR1 activation as a therapeutic target for osteosarcoma metastasis

Oncogene ◽  
2021 ◽  
Author(s):  
Satoshi Takagi ◽  
Yuki Sasaki ◽  
Sumie Koike ◽  
Ai Takemoto ◽  
Yosuke Seto ◽  
...  
Keyword(s):  
Platelet Activation ◽  
Pulmonary Metastasis ◽  
Lysophosphatidic Acid ◽  
Therapeutic Intervention ◽  
Molecular Mechanisms ◽  
Morphological Changes ◽  
Osteosarcoma Cells ◽  
Invasion And Metastasis ◽  
Platelet Releasate

AbstractOsteosarcoma is the most common primary malignant bone cancer, with high rates of pulmonary metastasis. Osteosarcoma patients with pulmonary metastasis have worse prognosis than those with localized disease, leading to dramatically reduced survival rates. Therefore, understanding the biological characteristics of metastatic osteosarcoma and the molecular mechanisms of invasion and metastasis of osteosarcoma cells will lead to the development of innovative therapeutic intervention for advanced osteosarcoma. Here, we identified that osteosarcoma cells commonly exhibit high platelet activation-inducing characteristics, and molecules released from activated platelets promote the invasiveness of osteosarcoma cells. Given that heat-denatured platelet releasate maintained the ability to promote osteosarcoma invasion, we focused on heat-tolerant molecules, such as lipid mediators in the platelet releasate. Osteosarcoma-induced platelet activation leads to abundant lysophosphatidic acid (LPA) release. Exposure to LPA or platelet releasate induced morphological changes and increased invasiveness of osteosarcoma cells. By analyzing publicly available transcriptome datasets and our in-house osteosarcoma patient-derived xenograft tumors, we found that LPA receptor 1 (LPAR1) is notably upregulated in osteosarcoma. LPAR1 gene KO in osteosarcoma cells abolished the platelet-mediated osteosarcoma invasion in vitro and the formation of early pulmonary metastatic foci in experimental pulmonary metastasis models. Of note, the pharmacological inhibition of LPAR1 by the orally available LPAR1 antagonist, ONO-7300243, prevented pulmonary metastasis of osteosarcoma in the mouse models. These results indicate that the LPA–LPAR1 axis is essential for the osteosarcoma invasion and metastasis, and targeting LPAR1 would be a promising therapeutic intervention for advanced osteosarcoma.

Formation, translocation and resolution of Holliday junctions during homologous genetic recombination

1995 ◽  
Vol 347 (1319) ◽  
pp. 21-25 ◽  
Keyword(s):  
Molecular Mechanisms ◽  
Genetic Recombination ◽  
Holliday Junctions ◽  
The Novel ◽  
E Coli ◽  
The Past ◽  
Endonucleolytic Cleavage ◽  
Insight Into ◽  
Made In

Over the past three or four years, great strides have been made in our understanding of the proteins involved in recombination and the mechanisms by which recombinant molecules are formed. This review summarizes our current understanding of the process by focusing on recent studies of proteins involved in the later steps of recombination in bacteria. In particular, biochemical investigation of the in vitro properties of the E. coli RuvA, RuvB and RuvC proteins have provided our first insight into the novel molecular mechanisms by which Holliday junctions are moved along DNA and then resolved by endonucleolytic cleavage.

Maintaining the permeability barrier during protein trafficking at the endoplasmic reticulum membrane

2003 ◽  
Vol 31 (6) ◽  
pp. 1227-1231 ◽  
Author(s):  
A.E. Johnson
Keyword(s):  
Endoplasmic Reticulum ◽  
Small Molecule ◽  
Protein Trafficking ◽  
Molecular Mechanisms ◽  
Protein Translocation ◽  
Cellular Membrane ◽  
Permeability Barrier ◽  
Endoplasmic Reticulum Membrane ◽  
Ion Diffusion

Many proteins are translocated across or integrated into a cellular membrane without disrupting its integrity, although it is difficult to imagine how such macromolecular transmembrane movement can occur without simultaneously allowing significant small-molecule and ion diffusion across the bilayer. Recent studies have identified some molecular mechanisms that are involved in maintaining the permeability barrier of the endoplasmic reticulum membrane during co-translational protein translocation and integration. These mechanisms are both simple and direct in concept, but are operationally complex and require the co-ordinated and regulated interaction of several multicomponent complexes.

scholarly journals Role of Oxidative Stress in the Senescence Pattern of Auditory Cells in Age-Related Hearing Loss

Antioxidants ◽  
2021 ◽  
Vol 10 (9) ◽  
pp. 1497
Author(s):  
Luz del Mar Rivas-Chacón ◽  
Sofía Martínez-Rodríguez ◽  
Raquel Madrid-García ◽  
Joaquín Yanes-Díaz ◽  
Juan Ignacio Riestra-Ayora ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Hearing Loss ◽  
Molecular Mechanisms ◽  
Morphological Changes ◽  
Cell Damage ◽  
Stria Vascularis ◽  
Organ Of Corti ◽  
Age Related ◽  
Age Related Hearing Loss

Age-related hearing loss (ARHL) is an increasing and gradual sensorineural hearing dysfunction. Oxidative stress is an essential factor in developing ARHL; additionally, premature senescence of auditory cells induced by oxidative stress can produce hearing loss. Hydrogen peroxide (H2O2) represents a method commonly used to generate cellular senescence in vitro. The objective of the present paper is to study H2O2-induced senescence patterns in three auditory cell lines (House Ear Institute-Organ of Corti 1, HEI-OC1; organ of Corti, OC-k3, and stria vascularis, SV-k1 cells) to elucidate the intrinsic mechanisms responsible for ARHL. The auditory cells were exposed to H2O2 at different concentrations and times. The results obtained show different responses of the hearing cells concerning cell growth, β-galactosidase activity, morphological changes, mitochondrial activation, levels of oxidative stress, and other markers of cell damage (Forkhead box O3a, FoxO3a, and 8-oxoguanine, 8-oxoG). Comparison between the responses of these auditory cells to H2O2 is a helpful method to evaluate the molecular mechanisms responsible for these auditory cells’ senescence. Furthermore, this in vitro model could help develop anti-senescent therapeutic strategies for the treatment of AHRL.

Do mycoplasmas cause human cancer?

2001 ◽  
Vol 47 (8) ◽  
pp. 691-697 ◽  
Author(s):  
Nevio Cimolai
Keyword(s):  
Tissue Culture ◽  
Molecular Mechanisms ◽  
Human Cancer ◽  
Epidemiological Studies ◽  
Respiratory Pathogen ◽  
The Novel ◽  
The 1960S ◽  
In Vitro Data

A linkage between mycoplasmas and malignancy was mainly proposed in the 1960s when human-associated mycoplasmas were becoming of interest given the novel characterization of the human respiratory pathogen Mycoplasma pneumoniae. Associations with leukemia and other malignancies, however, were largely ascribed to tissue-culture contamination, which is now recognized as a significant potential problem in molecular biology circles. A few epidemiological studies, however, continue to raise concern over such a linkage. As well, in vitro data have demonstrated the potential for some mycoplasmas to induce karyotypic changes and malignant transformation during chronic tissue-culture infestation. As cellular and molecular mechanisms for such transformation become studied, a resurgence of interest in this area is inevitable. A role for mycoplasmas in malignancy of any sort is conjectural, but there remains a need to continue with focussed epidemiological and laboratory investigations.Key words: mycoplasma, cancer, oncogenesis, leukemia.

scholarly journals Localization and distribution of nickel and other elements in in-vitro grown Alyssum corsicum exhibiting morphological changes in trichomes: initial insights into molecular mechanisms of nickel hyperaccumulation

2013 ◽  
Vol 37 ◽  
pp. 1115-1124 ◽  
Author(s):  
Seniha Selcen BABAOĞLU AYDAŞ ◽  
Leyla AÇIK ◽  
Danika LEDUC ◽  
Nezaket ADIGÜZEL ◽  
Şeküre Şebnem ELLİALTIOĞLU ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Morphological Changes ◽  
Nickel Hyperaccumulation

scholarly journals New insights into the mechanisms underlying 5-fluorouracil-induced intestinal toxicity based on transcriptomic and metabolomic responses in human intestinal organoids

2021 ◽  
Author(s):  
Daniela Rodrigues ◽  
Terezinha de Souza ◽  
Luke Coyle ◽  
Matteo Di Piazza ◽  
Bram Herpers ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Molecular Mechanisms ◽  
Morphological Changes ◽  
Cell Signalling ◽  
Mrna Translation ◽  
Atp Synthesis ◽  
Metabolome Analysis ◽  
Tissue Specific ◽  
Intestinal Toxicity

Abstract5-Fluorouracil (5-FU) is a widely used chemotherapeutical that induces acute toxicity in the small and large intestine of patients. Symptoms can be severe and lead to the interruption of cancer treatments. However, there is limited understanding of the molecular mechanisms underlying 5-FU-induced intestinal toxicity. In this study, well-established 3D organoid models of human colon and small intestine (SI) were used to characterize 5-FU transcriptomic and metabolomic responses. Clinically relevant 5-FU concentrations for in vitro testing in organoids were established using physiologically based pharmacokinetic simulation of dosing regimens recommended for cancer patients, resulting in exposures to 10, 100 and 1000 µM. After treatment, different measurements were performed: cell viability and apoptosis; image analysis of cell morphological changes; RNA sequencing; and metabolome analysis of supernatant from organoids cultures. Based on analysis of the differentially expressed genes, the most prominent molecular pathways affected by 5-FU included cell cycle, p53 signalling, mitochondrial ATP synthesis and apoptosis. Short time-series expression miner demonstrated tissue-specific mechanisms affected by 5-FU, namely biosynthesis and transport of small molecules, and mRNA translation for colon; cell signalling mediated by Rho GTPases and fork-head box transcription factors for SI. Metabolomic analysis showed that in addition to the effects on TCA cycle and oxidative stress in both organoids, tissue-specific metabolic alterations were also induced by 5-FU. Multi-omics integration identified transcription factor E2F1, a regulator of cell cycle and apoptosis, as the best key node across all samples. These results provide new insights into 5-FU toxicity mechanisms and underline the relevance of human organoid models in the safety assessment in drug development.

scholarly journals Changes in glycosaminoglycan biosynthesis during differentiation in vitro of human monocytes

1983 ◽  
Vol 210 (3) ◽  
pp. 661-667 ◽  
Author(s):  
S O Kolset ◽  
L Kjellén ◽  
R Seljelid ◽  
U Lindahl
Keyword(s):  
Culture Medium ◽  
Negative Charge ◽  
Morphological Changes ◽  
Paper Electrophoresis ◽  
Exchange Chromatography ◽  
Adherent Cells ◽  
The Novel ◽  
Dermatan Sulphate ◽  
Negative Charge Density

Monocytes isolated from human blood were maintained in vitro on plastic culture dishes. After 3-4 days, adherent cells displayed morphological changes previously attributed to differentiation of the cells into histiocytes. 35S-labelled glycosaminoglycans were isolated after incubation of the cells with inorganic [35S]sulphate. Polysaccharide recovered from the culture medium after labelling from day 0 to day 2 or from day 5 to day 7 in vitro was approximately 90% galactosaminoglycan (resistant to deamination by HNO2), irrespective of labelling period. Whereas day-0-2 material was extensively degraded to disaccharide on incubation with the bacterial eliminase chondroitinase AC, a significant portion, about 30%, of the day-5-7 material resisted degradation under the same conditions. The resistant portion was readily depolymerized by treatment with chondroitinase ABC and may be dermatan sulphate. Paper electrophoresis and paper chromatography of the disaccharides obtained by eliminase digestion identified the day-0-2 labelled galactosaminoglycan as chondroitin 4-sulphate. In contrast, the corresponding day-5-7 material yielded approximately 20% disulphated disaccharide, both on digestion with chondroitinase AC and on subsequent enzymic degradation of the chondroitinase AC-resistant fraction. Further treatment of the disulphated disaccharide with chondro-4-sulphatase and chondro-6-sulphatase indicated that both sulphate groups were located on the N-acetylgalactosamine residue. In accordance with these findings, the day-5-7 polysaccharide showed a higher negative charge density than the day-0-2 material on ion-exchange chromatography. It is concluded that the novel properties acquired by the monocyte during prolonged culturing on plastic include the ability to synthesize glycosaminoglycan(s) containing 4,6-disulphated N-acetylgalactosamine units.

scholarly journals Convergence of Notch and β-catenin signaling induces arterial fate in vascular progenitors

2010 ◽  
Vol 189 (2) ◽  
pp. 325-338 ◽  
Author(s):  
Kohei Yamamizu ◽  
Taichi Matsunaga ◽  
Hideki Uosaki ◽  
Hiroyuki Fukushima ◽  
Shiori Katayama ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Embryonic Stem ◽  
Gene Promoter ◽  
Stem Cell Differentiation ◽  
Embryonic Stem Cell Differentiation ◽  
Target Gene Promoter ◽  
Vascular Progenitor Cells ◽  
Molecular Machinery

Molecular mechanisms controlling arterial–venous specification have not been fully elucidated. Previously, we established an embryonic stem cell differentiation system and demonstrated that activation of cAMP signaling together with VEGF induces arterial endothelial cells (ECs) from Flk1+ vascular progenitor cells. Here, we show novel arterial specification machinery regulated by Notch and β-catenin signaling. Notch and GSK3β-mediated β-catenin signaling were activated downstream of cAMP through phosphatidylinositol-3 kinase. Forced activation of Notch and β-catenin with VEGF completely reconstituted cAMP-elicited arterial EC induction, and synergistically enhanced target gene promoter activity in vitro and arterial gene expression during in vivo angiogenesis. A protein complex with RBP-J, the intracellular domain of Notch, and β-catenin was formed on RBP-J binding sites of arterial genes in arterial, but not venous ECs. This molecular machinery for arterial specification leads to an integrated and more comprehensive understanding of vascular signaling.

scholarly journals Redox regulation by TXNRD3 during epididymal maturation underlies capacitation-associated mitochondrial activation and sperm motility in mice

2021 ◽  
Author(s):  
Huafeng Wang ◽  
Qianhui Dou ◽  
Kyung Jo Jung ◽  
Jungmin Choi ◽  
Vadim N. Gladyshev ◽  
...  
Keyword(s):  
Redox Regulation ◽  
Molecular Mechanisms ◽  
Redox Homeostasis ◽  
Thioredoxin Reductase ◽  
Redox Status ◽  
Male Reproduction ◽  
Sperm Maturation ◽  
Epididymal Maturation ◽  
Sperm Development ◽  
Sheath Formation

During epididymal transit, redox remodeling protects mammalian spermatozoa, preparing them for survival in the subsequent journey to fertilization. However, molecular mechanisms of the redox regulation in sperm development and maturation remain largely elusive. In this study, we report thioredoxin reductase 3 (TXNRD3) - a thioredoxin reductase family member particularly abundant in elongating spermatids at the site of mitochondrial sheath formation - contributes to regulate redox homeostasis in male reproduction. Using Txnrd3-/- mice, our biochemical, ultrastructural, and live cell imaging analyses revealed impairments in sperm morphology and motility in absence of TXNRD3. Absence of TXNRD3 alters redox status in both the head and tail during sperm maturation and capacitation, resulting in defective mitochondrial ultrastructure and activity under capacitating conditions. These findings provide insights into molecular mechanisms of redox homeostasis and bioenergetics during sperm maturation, capacitation, and fertilization.

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