The changes of heavy metal and metallothionein distribution in testis induced by cadmium exposure

Cadmium (Cd) is known to cause various disorders in the testis, and metallothionein (MT) is known as a protein, which has a detoxification function for heavy metals. However, the changes of Fe, Cu, and Zn distribution in the testis induced by Cd exposure have not been well examined. Moreover, only a few studies have been reported on the localization of MT after Cd exposure. In this study, we have investigated the changes of Fe, Cu, and Zn distribution in Cd-exposed testis by a newly developed in air micro-Particle Induced X-ray Emission (PIXE) method. Also, we examined the distribution of MT expression in testis. In the testis of Cd-treated rats with significant increases of lipid peroxidation, the sertoli cell tight junction was damaged by Cd exposure, resulting from disintegration of the blood testis barrier (BTB). Evaluation by in air micro-PIXE method revealed that Cd and Fe distribution were increased in the interstitial tissues and seminiferous tubules. The histological findings indicated that the testicular tissue damage was advanced, which may have been caused by Fe flowing into seminiferous tubules followed by disintegration of the BTB. As a result, Fe was considered to enhance the tissue damage caused by Cd exposure. MT was detected in spermatogonia, spermatocytes, and Sertoli’s cells in the testis of Cd-treated rats, but was not detected in interstitial tissues. These results suggested that MT was induced by Cd in spermatogonia, spermatocytes, and Sertoli’s cells, and was involved in the resistance to tissue damage induced by Cd.     

Cadmium binding studies to the earthworm Lumbricus rubellus metallothionein by electrospray mass spectrometry and circular dichroism spectroscopy

The earthworm Lumbricus rubellus has been found to inhabit cadmium-rich soils and accumulate cadmium within its tissues. Two metallothionein (MT) isoforms (1 and 2) have been identified and cloned from L. rubellus. In this study, we address the metalation status, metal coordination, and structure of recombinant MT-2 from L. rubellus using electrospray ionization mass spectrometry (ESI-MS), UV absorption, and circular dichroism (CD) spectroscopy. This is the first study to show the detailed mass and CD spectral properties for the important cadmium-containing earthworm MT. We report that the 20-cysteine L. rubellus MT-2 binds seven Cd(2+) ions. UV absorption and CD spectroscopy and ESI-MS pH titrations show a distinct biphasic demetalation reaction, which we propose results from the presence of two metal-thiolate binding domains. We propose stoichiometries of Cd(3)Cys(9) and Cd(4)Cys(11) based on the presence of 20 cysteines split into two isolated regions of the sequence with 11 cysteines in the N-terminal and 9 cysteines in the C-terminal. The CD spectrum reported is distinctly different from any other metallothionein known suggesting quite different binding site structure for the peptide.     

Comparative genomics analysis of metallothioneins in twelve Drosophila species

Metallothioneins (MTs) are a superfamily of Cys-rich polypeptides that bind heavy metal ions, both for physiological and detoxification purposes. They are present in all organisms, but their origin is probably polyphyletic, so that MT evolutionary studies are rather scarce. We present a thorough search and analysis of the MT coding sequences in the 12 Drosophila genomes completely sequenced, taking as reference the features reported for D. melanogaster, where four isogenes (MtnA to MtnD) are known and deeply characterized. We include a fifth isoform in this study, named MtnE, and recently annotated. The MTs polymorphism pattern is essentially the same for the 12 Drosophila species. Invariably, a MtnA form and an MtnB-cluster, comprising the MtnB-to-MtnE forms in tandem array, are observed. The whole set of genes are kept in the same synteny element (Muller E), but implicated in rearrangement events (mainly inversions), encompassing all or some of the isogenes. Gene exon/intron architecture, and cDNA and protein sequences appear highly conserved through Drosophila speciation, concordantly with an essential function for MT isoforms in flies, even for those previously considered as minor products. Data presented here will be comprehensively analyzed to provide a valuable guide for future MT evolutionary, structure and function studies.     

Evidence for operation of the direct zinc ligand exchange mechanism for trafficking, transport, and reactivity of zinc in mammalian cells

In addition to its critical role in normal cell function, growth, and metabolism, zinc is implicated as a major factor in the development and progression of many pathological conditions and diseases. Despite this importance of zinc, many important factors, processes, and mechanisms of the physiology, biochemistry, and molecular biology of zinc remain unknown. Especially important is the unresolved issue regarding the mechanism and process of the trafficking, transport, and reactivity of zinc in cells; especially in mammalian cells. This presentation focuses on the concept that, due to the existence of a negligible pool of free Zn²⁺ ions in the mammalian cell environment, the trafficking, transport and reactivity of zinc occurs via a direct exchange of zinc from donor Zn-ligands to acceptor ligands. This Zn exchange process occurs without the requirement for production of free Zn²⁺ ions. The direct evidence from mammalian cell studies is presented in support of the operation of the direct Zn-ligand exchange mechanism. The paper also provides important information and conditions that should be considered and employed in the conduct of studies regarding the role and effects of zinc in biological/biomedical research; and in its clinical interpretation and application.     

Variation in metallothionein gene expression is associated with adaptation to copper in the earthworm Dendrobaena octaedra

Evolution of resistance to heavy metals has been reported for several populations of soil living organisms occurring at metal contaminated sites. Such genetically based and heritable resistance contribute to the persistence of populations in contaminated areas. Here we report on molecular responses to experimental copper in populations of the earthworm, Dendrobaena octaedra, originating from copper contaminated soil near Gusum (Sweden) where heavy metal pollution has been present for several decades. We studied gene expression of six genes potentially involved in resistance to copper toxicity using F2-generations of D. octaedra populations, originating from reference sites and contaminated (High, Medium and Low) sites around Gusum. The main result was different expression patterns of genes encoding for two different isoforms (mt1 and mt2) of metallothionein proteins during experimental exposure to copper contaminated soil. Expression of mt1 showed a fast and significant upregulation in the High population and a slower, albeit significant, upregulation in Medium and Low populations. However, in the three reference populations no upregulation were seen. In comparison, a fast upregulation was also seen for the High population in the isoform mt2, whereas, gene expression of all other populations, including reference populations, showed slower upregulation in response to experimental copper. The results indicate that copper resistance in D. octaedra from contaminated areas is related to an increased expression of metallothioneins.     

Cyanobacterial metallothioneins: Biochemistry and molecular genetics

Summary Metallothioneins have been extensively studied in many different eukaryotes where they sequester, and hence detoxify, excess amounts of certain metal ions. However, the precise functions of many of these molecules are not fully understood. This article reviews literature concerning their namesakes in prokaryotes.Abbreviations MT metallothionein - MRE metal regulatory element     

以FITC作为荧光探针研究金属硫蛋白的构象变化

本文以异硫氰基荧光素（ＦＩＴＣ）作为荧光探针标记于金属硫蛋白分子上,用荧光光谱研究了Ｃｄ＾２＋及Ａｇ＾＋离子与ＺｎＭＴ２－ＦＩＴＣ进行金属交换及与ＡｐｏＭＴ２－ＦＩＴＣ进行金属重组时的构象变化。结果表明,标记后ＭＴ与Ｃｄ＾２＋离子进行金属交换及金属重组时不具有明显的结构域特征,而Ａｇ＾＋离子进行金属交换及金属重组时,分别在Ａｇ６ＭＴ、Ａｇ１２ＭＴ及Ａｇ１８ＭＴ处具有明显的结构域形成特征。此外高温下