Zinc and DHA have opposing effects on the expression levels of histones H3 and H4 in human neuronal cells

Zn and DHA have putative neuroprotective effects and these two essential nutrients are known to interact biochemically. We aimed to identify novel protein candidates that are differentially expressed in human neuronal cell line M17 in response to Zn and DHA that would explain the molecular basis of this interaction. Two-dimensional gel electrophoresis and MS were applied to identify major protein expression changes in the protein lysates of human Ml7 neuronal cells that had been grown in the presence and absence of Zn and DHA. Proteomic findings were further investigated using Western immunoblot and real-time PCR analyses. Four protein spots, which had significant differential expression, were identified and selected for in-gel trypsin digestion followed by matrix-assisted laser desorption ionisation MS analysis. The resultant peptide mass fingerprint for each spot allowed their respective identities to be deduced. Two human histone variants H3 and H4 were identified. Both H3 and H4 were downregulated by Zn in the absence of DHA (Zn effect) and upregulated by DHA (DHA effect) in the presence of Zn (physiological condition). These proteomic findings were further supported by Western immunoblot and real-time PCR analyses using H3- and H4-specific monoclonal antibodies and oligonucleotide primers, respectively. We propose that dietary Zn and DHA cause a global effect on gene expression, which is mediated by histones. Such novel information provides possible clues to the molecular basis of neuroprotection by Zn and DHA that may contribute to the future treatment, prevention and management of neurodegenerative diseases such as Alzheimer’s disease. Zinc: n-3 Fatty acids: DHA: M17 human neuronal cells: Histone H3: Histone H4 Neuronal cell damage or death (apoptosis) is a key feature in the pathology of neurodegenerative disorders, such as ischaemic stroke, Alzheimer’s disease (AD) and Parkinson’s disease (1) . Therefore, neuronal cell survival is pivotal in preventing such neurodegenerative disorders. Importantly, identification and characterisation of the critical modulators of neuronal cell survival are of paramount significance to elucidate the mechanisms underlying neurological diseases and thus facilitate the development of novel treatment strategies. Another common feature seen in AD brains is lesions of extracellular amyloid plaques, which consist of protein aggregates of b-amyloid (2,3) . Significantly, free neuronal Zn has been shown to promote aggregation of the b-amyloid (4) , and increased levels of Zn are reportedly colocalised in the