An H1-like protein from the sperm chromatin of 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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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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