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. 2020 Sep 13;21(18):6710.
doi: 10.3390/ijms21186710.

Molecular Alterations in Spermatozoa of a Family Case Living in the Land of Fires. A First Look at Possible Transgenerational Effects of Pollutants

Gennaro Lettieri  1 , Federica Marra  1 , Claudia Moriello  1 , Marina Prisco  1 , Tiziana Notari  2 , Marco Trifuoggi  3 , Antonella Giarra  3 , Liana Bosco  4 , Luigi Montano  5 , Marina Piscopo  1
Affiliations

Molecular Alterations in Spermatozoa of a Family Case Living in the Land of Fires. A First Look at Possible Transgenerational Effects of Pollutants

Gennaro Lettieri et al. Int J Mol Sci. .

Abstract

In our previous work, we reported alterations in protamines/histones ratio, in DNA binding of these proteins and their involvement in DNA oxidative damage in 84% of the young men living in the Land of Fires. In the present work, we extended our findings, evaluating any alterations in spermatozoa of a family case, a father and son, living in this area, to also give a first look at the possibility of transgenerational inherited effects of environmental contaminants on the molecular alterations of sperm nuclear basic proteins (SNBP), DNA and semen parameters. In the father and son, we found a diverse excess of copper and chromium in the semen, different alterations in SNBP content and low DNA binding affinity of these proteins. In addition, DNA damage, in the presence of CuCl2 and H2O2, increased by adding both the father and son SNBP. Interestingly, son SNBP, unlike his father, showed an unstable DNA binding and were able to produce DNA damage even without external addition of CuCl2, in line with a lower seminal antioxidant activity than the father. The peculiarity of some characteristics of son semen could be a basis for possible future studies on transgenerational effects of pollutants on fertility.

Keywords: DNA oxidative damage; EMSA; Land of Fires; heavy metals; human protamines; human spermatozoa; protein-DNA binding; transgenerational effects.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Aniline blue staining from control 1 sample (a), son (b) and father (c).
Figure 2
Figure 2
Characterization of human SNBP from samples belonging to two individuals of L-group (controls) and a father and his son living in the “Land of Fires”. AU-PAGE of SNBP of: rabbit total histones(lane 1); sea urchin total histones (lane 2); human control 1 and 2 samples, showing the CP/Hr (lanes 3–4); son sample showing the nCP/Hr (lane 5) and father sample showing only-H (lane 6); calf thymus H1 histone (lane 7). SNBP: sperm nuclear basic proteins; L-group: low impact area group; CP/Hr: canonical protamines/histones ratio; nCP/Hr: non-canonical protamines/histones ratio; only-H: only histones and other basic proteins.
Figure 3
Figure 3
DNA binding ability of SNBP obtained from: control 1 (a); son (b) and father (c) analyzed by EMSA on 1% agarose gel. Bands on gel representing the state of pGEM3 plasmid DNA incubated in a w/w ratio with increasing amount of SNBP from samples containing: CP/Hr (a); nCP/Hr (b); only–H (c). Panel a from supplementary material of Lettieri et al., 2020 [11]. L-group: low impact area group; EMSA: electrophoresis mobility shift Assay; SNBP: sperm nuclear basic proteins; CP/Hr: canonical protamines/histones ratio; nCP/Hr: not canonical protamines/histones ratio; only-H: only histones and other basic proteins.
Figure 4
Figure 4
Chromium and copper accumulation in the semen of control 1, son and father.
Figure 5
Figure 5
Analysis on 1% agarose gel of pGEM3 plamid DNA breakage induced by H2O2. In the presence of SNBP of the son (a) and father (b). SNBP: sperm nuclear basic proteins.
Figure 6
Figure 6
DNA protection analysis on 1% agarose gel of pGEM3 plasmid DNA in the presence of increasing (0.4, 0.6 and 0.8) controls (lanes 6, 9 and 12), son (lanes 4, 7 and 10) and father (5, 8 and 11) SNBP/DNA ratios. SNBP: sperm nuclear basic proteins.
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