CONTRIBUTION TO THE STUDY OF MOLECULAR EFFECTS OF COPPER ON MYTILUS GALLOPROVINCIALIS DURING REPRODUCTIVE PERIOD

Successful reproduction is a determining factor for the survival of species. In this century, major risks affecting reproductive health are for those species that fertilize externally, such as several marine invertebrates, since several marine ecosystems are polluted by different types of xenobiotics. Among the xenobiotics present in seawater, a relevant role is played by heavy metals whose release into the marine environment has increased their levels to large extents within the past few decades. Copper is one of the most interesting heavy metals, because in small quantities, it is essential, being involved in several physiological functions, such as redox reactions, oxygen transport, cellular respiration, free radical defense, neurotransmitter synthesis, and neuronal myelination, but can be toxic above certain threshold concentrations. In this study, we have evaluated the effects induced by 24 hr exposure to a subtoxic copper concentration on the reproductive system (gonads, spermatozoa, and protamine‐like proteins) of Mytilus galloprovincialis. Protamine‐like proteins represent the main basic protein component of sperm chromatin of this organism. After exposure, we found accumulation of this metal in gonads, spermatozoa, and protamine‐like proteins of exposed mussels, as indicated by inductively coupled plasma–mass spectrometry analyses. Moreover, altered expression levels of mt10 and protamine‐like proteins genes in spermatozoa and gonads, respectively, of exposed mussels were registered by real‐time polymerase chain reaction analyses. We analyzed also the DNA binding affinity of protamine‐like proteins extracted from spermatozoa of exposed mussels. The results showed a higher DNA binding affinity and a different DNA binding mode in exposed mussels. Moreover, an increased amount of NaCl was required for the release from sperm nuclei of PL‐III, the main protamine‐like proteins component. Interestingly, protamine‐like proteins extracted from spermatozoa of exposed mussels promoted DNA oxidative damage in the presence of H2O2. These results demonstrate that also tolerable copper amount could affect the properties of protamine‐like proteins and determine negative effects on Mytilus galloprovincialis reproductive system. Finally, these analyses could be useful to develop quick and efficient chromatin‐based genotoxicity tests for pollution biomonitoring programs.