Biochemical and ultrastructural study of the sperm chromatin from Mytilus galloprovincialis

Mercury is one of the most dangerous environmental pollutants. In this work, we analysed the effects of exposure of Mytilus galloprovincialis to 1, 10 and 100 pM HgCl2 for 24 h on the gonadal morphology and on the expression level of three stress genes: mt10, hsp70 and πgst. In this tissue we also evaluated the level of steroidogenic enzymes 3β-HSD and 17β-HSD and the expression of PL protein genes. Finally, we determined difference in sperm chromatin accessibility to micrococcal nuclease. We found alterations in gonadal morphology especially after exposure to 10 and 100 pM HgCl2 and hypo-expression of the three stress genes, particularly for hsp70. Furthermore, decreased labelling with both 3β-HSD and 17β-HSD antibodies was observed following exposure to 1 and 10 pM HgCl2 and complete absence at 100 pM HgCl2 exposure. Gonads of mussels exposed to all HgCl2 doses showed decreased expression of PL protein genes especially for PLIII. Finally, micrococcal nuclease digestions showed that all doses of HgCl2 exposure resulted in increased sperm chromatin accessibility to this enzyme, indicative of improper sperm chromatin structure. All of these changes provide preliminary data of the potential toxicity of mercury on the reproductive health of this mussel.

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Histochemical and ultrastructural study on the innervation of the byssus glands of Mytilus galloprovincialis

Cell and Tissue Research ◽

10.1007/bf00238306 ◽

1983 ◽

Vol 233 (2) ◽

pp. 403-413 ◽

Cited By ~ 9

Author(s):

Laura Vitellaro-Zuccarello ◽

Silvia De Biasi ◽

Isabella Blum

Keyword(s):

Ultrastructural Study ◽

Mytilus Galloprovincialis

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Spermatozoa Transcriptional Response and Alterations in PL Proteins Properties after Exposure of Mytilus galloprovincialis to Mercury

International Journal of Molecular Sciences ◽

10.3390/ijms22041618 ◽

2021 ◽

Vol 22 (4) ◽

pp. 1618

Author(s):

Gennaro Lettieri ◽

Rosaria Notariale ◽

Alessia Ambrosino ◽

Alfredo Di Bonito ◽

Antonella Giarra ◽

...

Keyword(s):

Inductively Coupled Plasma ◽

Mytilus Galloprovincialis ◽

Binding Capacity ◽

Transcriptional Response ◽

Environmental Pollutant ◽

Sperm Chromatin ◽

Altered Expression ◽

Inductively Coupled ◽

Sds Page ◽

Plasma Mass Spectrometry

Mercury (Hg) is an environmental pollutant that impacts human and ecosystem health. In our previous works, we reported alterations in the properties of Mytilus galloprovincialis protamine-like (PL) proteins after 24 h of exposure to subtoxic doses of toxic metals such as copper and cadmium. The present work aims to assess the effects of 24 h of exposure to 1, 10, and 100 pM HgCl2 on spermatozoa and PL proteins of Mytilus galloprovincialis. Inductively coupled plasma–mass spectrometry indicated accumulation of this metal in the gonads of exposed mussels. Further, RT-qPCR analyses showed altered expression levels of spermatozoa mt10 and hsp70 genes. In Mytilus galloprovincialis, PL proteins represent the major basic component of sperm chromatin. These proteins, following exposure of mussels to HgCl2, appeared, by SDS-PAGE, partly as aggregates and showed a decreased DNA-binding capacity that rendered them unable to prevent DNA damage, in the presence of CuCl2 and H2O2. These results demonstrate that even these doses of HgCl2 exposure could affect the properties of PL proteins and result in adverse effects on the reproductive system of this organism. These analyses could be useful in developing rapid and efficient chromatin-based genotoxicity assays for pollution biomonitoring programs.

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New Insights into Alterations in PL Proteins Affecting Their Binding to DNA after Exposure of Mytilus galloprovincialis to Mercury—A Possible Risk to Sperm Chromatin Structure?

International Journal of Molecular Sciences ◽

10.3390/ijms22115893 ◽

2021 ◽

Vol 22 (11) ◽

pp. 5893

Author(s):

Gennaro Lettieri ◽

Rosaria Notariale ◽

Nadia Carusone ◽

Antonella Giarra ◽

Marco Trifuoggi ◽

...

Keyword(s):

Fluorescence Spectroscopy ◽

Conformational Changes ◽

Mytilus Galloprovincialis ◽

Polyacrylamide Gel Electrophoresis ◽

Sodium Dodecyl ◽

Reproductive Activity ◽

Structural Rearrangement ◽

Sperm Chromatin ◽

Sperm Nuclei ◽

Protein Alterations

Mercury (Hg) is a highly toxic and widespread pollutant. We previously reported that the exposure of Mytilus galloprovincialis for 24 h to doses of HgCl2 similar to those found in seawater (range 1–100 pM) produced alterations in the properties of protamine-like (PL) proteins that rendered them unable to bind and protect DNA from oxidative damage. In the present work, to deepen our studies, we analyzed PL proteins by turbidimetry and fluorescence spectroscopy and performed salt-induced release analyses of these proteins from sperm nuclei after the exposure of mussels to HgCl2 at the same doses. Turbidity assays indicated that mercury, at these doses, induced PL protein aggregates, whereas fluorescence spectroscopy measurements showed mercury-induced conformational changes. Indeed, the mobility of the PLII band changed in sodium dodecyl sulphate-polyacrylamide gel electrophoresis, particularly after exposure to 10-pM HgCl2, confirming the mercury-induced structural rearrangement. Finally, exposure to HgCl2 at all doses produced alterations in PL-DNA binding, detectable by DNA absorption spectra after the PL protein addition and by a decreased release of PLII and PLIII from the sperm nuclei. In conclusion, in this paper, we reported Hg-induced PL protein alterations that could adversely affect mussel reproductive activity, providing an insight into the molecular mechanism of Hg-related infertility.

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Protamine-like proteins have bactericidal activity. The first evidence in Mytilus galloprovincialis

Acta Biochimica Polonica ◽

10.18388/abp.2018_2638 ◽

2018 ◽

Cited By ~ 1

Author(s):

Rosaria Notariale ◽

Adriana Basile ◽

Elena Montana ◽

Nunzia Colonna Romano ◽

Maria Grazia Cacciapuoti ◽

...

Keyword(s):

Bactericidal Activity ◽

Mytilus Galloprovincialis ◽

Bacterial Species ◽

Enterobacter Aerogenes ◽

Sperm Chromatin ◽

Bacterial Strains ◽

Gram Positive ◽

Gram Negative ◽

Intermediate Group

The major acid-soluble protein components of the mussel Mytilus galloprovincialis sperm chromatin consist of the protamine-like proteins PL-II, PL-III and PL-IV, an intermediate group of sperm nuclear basic proteins between histones and protamines. The aim of this study was to investigate the bactericidal activity of these proteins since, to date, there are reports on bactericidal activity of protamines and histones, but not on protamine-like proteins. We tested the bactericidal activity of these proteins against Gram-positive bacteria: Enterococcus faecalis and two different strains of Staphylococcus aureus, as well as Gram-negative bacteria: Proteus mirabilis, Proteus vulgaris, Pseudomonas aeruginosa, Salmonella typhmurium, Enterobacter aerogenes, Enterobacter cloacae, and Escherichia coli. Clinical isolates of the same bacterial species were also used to compare their sensitivity to these proteins. The results show that Mytilus galloprovincialis protamine-like proteins exhibited bactericidal activity against all bacterial strains tested with different minimum bactericidal concentration values, ranging from 15.7 to 250 µg/mL. Furthermore, these proteins were active against some bacterial strains tested that are resistant to conventional antibiotics. These proteins showed very low toxicity as judged by red blood cell lysis and viability MTT assays and seem to act both at the membrane level and within the bacterial cell. We also tested the bactericidal activity of the product obtained from an in vitro model of gastrointestinal digestion of protamine-like proteins on a Gram-positive and a Gram-negative strain, and obtained the same results with respect to undigested protamine-like proteins on the Gram-positive bacterium. These results provide the first evidence of bactericidal activity of protamine-like-proteins.

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Tumor Diagnosis

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100053966 ◽

1982 ◽

Vol 40 ◽

pp. 262-265

Author(s):

Bruce Mackay

Keyword(s):

Electron Microscopy ◽

Transmission Electron Microscopy ◽

Differential Diagnosis ◽

Light Microscopy ◽

Clinical Examination ◽

Ultrastructural Study ◽

Diagnostic Procedures ◽

Specific Diagnosis ◽

Transmission Electron ◽

Microscopic Diagnosis

The broadest application of transmission electron microscopy (EM) in diagnostic medicine is the identification of tumors that cannot be classified by routine light microscopy. EM is useful in the evaluation of approximately 10% of human neoplasms, but the extent of its contribution varies considerably. It may provide a specific diagnosis that can not be reached by other means, but in contrast, the information obtained from ultrastructural study of some 10% of tumors does not significantly add to that available from light microscopy. Most cases fall somewhere between these two extremes: EM may correct a light microscopic diagnosis, or serve to narrow a differential diagnosis by excluding some of the possibilities considered by light microscopy. It is particularly important to correlate the EM findings with data from light microscopy, clinical examination, and other diagnostic procedures.

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Further Observations on the Virus of Epizootic Diarrhea of Infant Mice. An Electron Microscopic Study

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100060568 ◽

1967 ◽

Vol 25 ◽

pp. 110-111

Author(s):

W. G. Banfield ◽

G. Kasnic ◽

J. H. Blackwell

Keyword(s):

Electron Microscope ◽

Infected Cell ◽

Microscopic Study ◽

Intestinal Epithelium ◽

Ultrastructural Study ◽

Osmium Tetroxide ◽

Lead Citrate ◽

Electron Microscopic ◽

Virus Particles ◽

Infected Cells

An ultrastructural study of the intestinal epithelium of mice infected with the agent of epizootic diarrhea of infant mice (EDIM virus) was first performed by Adams and Kraft. We have extended their observations and have found developmental forms of the virus and associated structures not reported by them.Three-day-old NLM strain mice were infected with EDIM virus and killed 48 to 168 hours later. Specimens of bowel were fixed in glutaraldehyde, post fixed in osmium tetroxide and embedded in epon. Sections were stained with uranyl magnesium acetate followed by lead citrate and examined in an updated RCA EMU-3F electron microscope.The cells containing virus particles (infected) are at the tips of the villi and occur throughout the intestine from duodenum through colon. All developmental forms of the virus are present from 48 to 168 hours after infection. Figure 1 is of cells without virus particles and figure 2 is of an infected cell. The nucleus and cytoplasm of the infected cells appear clearer than the cells without virus particles.

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Glycogen in guinea pig neutrophils: Ultrastructural study of neutrophils at the intradermal site of BCG injection

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100077207 ◽

1983 ◽

Vol 41 ◽

pp. 726-727

Author(s):

Corazon D. Bucana

Keyword(s):

Bone Marrow ◽

Guinea Pig ◽

Electron Density ◽

Peritoneal Cavity ◽

Ultrastructural Study ◽

Guinea Pigs ◽

Random Distribution ◽

Glycogen Content ◽

Polymorphonuclear Neutrophils ◽

Ultrastructural Studies

In the circulating blood of man and guinea pigs, glycogen occurs primarily in polymorphonuclear neutrophils and platelets. The amount of glycogen in neutrophils increases with time after the cells leave the bone marrow, and the distribution of glycogen in neutrophils changes from an apparently random distribution to large clumps when these cells move out of the circulation to the site of inflammation in the peritoneal cavity. The objective of this study was to further investigate changes in glycogen content and distribution in neutrophils. I chose an intradermal site because it allows study of neutrophils at various stages of extravasation.Initially, osmium ferrocyanide and osmium ferricyanide were used to fix glycogen in the neutrophils for ultrastructural studies. My findings confirmed previous reports that showed that glycogen is well preserved by both these fixatives and that osmium ferricyanide protects glycogen from solubilization by uranyl acetate.I found that osmium ferrocyanide similarly protected glycogen. My studies showed, however, that the electron density of mitochondria and other cytoplasmic organelles was lower in samples fixed with osmium ferrocyanide than in samples fixed with osmium ferricyanide.

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An ultrastructural study of vegetative incompatibility in plants

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100083047 ◽

1978 ◽

Vol 36 (2) ◽

pp. 436-437

Author(s):

Randy Moore

Keyword(s):

Ultrastructural Study ◽

Cell Walls ◽

Growth And Development ◽

Solanum Pennellii ◽

Initial Product ◽

Basic Aspect ◽

Tissue Interactions ◽

Graft Interface ◽

Antigen Antibody ◽

Standard Fixation

Cell and tissue interactions are a basic aspect of eukaryotic growth and development. While cell-to-cell interactions involving recognition and incompatibility have been studied extensively in animals, there is no known antigen-antibody reaction in plants and the recognition mechanisms operating in plant grafts have been virtually neglected.An ultrastructural study of the Sedum telephoides/Solanum pennellii graft was undertaken to define possible mechanisms of plant graft incompatibility. Grafts were surgically dissected from greenhouse grown plants at various times over 1-4 weeks and prepared for EM employing variations in the standard fixation and embedding procedure. Stock and scion adhere within 6 days after grafting. Following progressive cell senescence in both Sedum and Solanum, the graft interface appears as a band of 8-11 crushed cells after 2 weeks (Fig. 1, I). Trapped between the buckled cell walls are densely staining cytoplasmic remnants and residual starch grains, an initial product of wound reactions in plants.

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