In vivo screening of proteins likely to bind uranium in exposed rat kidney

Abstract

Uranium is a naturally abundant element which has been used in several industries. Internal exposure could occur via three main pathways that are ingestion, inhalation and wounds. It has been recently shown that chronic ingestion of uranium in drinking water induces an important uranium accumulation in kidney with a perturbation of iron metabolism in this organ.Whereas uranium speciation is a key parameter to elucidate the chemical reactivity and the mobility of an element, it remains poorly documented in most of environmental and biological media. A few examples of uranium complexation with biomolecules have been published recently but most of them are in vitro studies whereas in vivo experiments remain poorly investigated.In order to better understand possible competition of uranium towards metals involved in the metal-protein binding, i.e. iron, copper, calcium, a study on uranium speciation was investigated by doing an in vivo screening of target proteins likely to bind it in kidneys of exposed rats. Rats were chronically exposed via contaminated drinking water at 40 mg L −1 and killed 9 months after the beginning of exposure. Kidneys were dissected out and protein extract was prepared. Then, separation of renal proteins by isoelectric focusing gel electrophoresis (IEF) and two-dimensional gel electrophoresis (2-DE) followed by LA-ICPMS analysis were performed.IEF-LA-ICP MS showed that uranium could specifically bind few proteins in kidney whereas 2-DE-LA-ICP MS could indicate that uranium is not covalently bound to proteins in this organ. The results suggested that even at moderate concentrations of exposure, uranium can be observed chelated with some renal proteins that is very encouraging to understand the entry, storage and elimination of this element in kidneys.

Insights into the nature of uranium target proteins within zebrafish gills after chronic and acute waterborne exposures

“…We could therefore consider that there may be a mechanism involving the formation of ATP–U complexes with a potential impact on heat shock protein 70 functions. Beta‐actin is part of the actin protein family, which has already been suggested as a potential U target in rat kidney . It is a 41.7‐kDa protein involved in numerous structural functions of the cell through its polymerization into microfilament structures.…”

“…Complementarily, other in vivo screening research determined that U‐binding proteins are mainly acidic, often metal‐binding, proteins (e.g., Fe–ferritin, Cu–hemocyanin, Mn–superoxide dismutase) and also that they often bind with phosphorus groups .…”

Separative techniques for metalloproteomics require balance between separation and perturbation

“…Consequently, the use of SDS and other denaturing agents was shown to induce partial or total loss of metal from metalloproteins, such as superoxide dismutase (SOD), carbonic anhydrase (CA), cytochrome C or holotransferrin (holoTf) [35]. Similar results concerning the influence of SDS were found on uranyl-binding proteins from renal cells by Frelon et al [40], who reported two different causes for the loss of uranyl during the separation steps:…”

“…The combination of these two dimensions often reaches a high resolution. Some studies on real biological samples have been performed by transferring classical methods used for proteomics to metalloproteomics {i.e., combining classical 2D-GE with laser-ablation ICP-MS (LA-ICP-MS) for detection of metals [40,49,66]…”

Subcellular fractionation and chemical speciation of uranium to elucidate its fate in gills and hepatopancreas of crayfish Procambarus clarkii

“…In rat kidney uranium was mainly detected (after exposure) bound to acidic proteins (Frelon et al, 2009). In human kidney cell extracts, proteins referred to as hard Lewis cation (Ca 2+ , Zn 2+ , Mn 2+ or Mg 2+ ) binding proteins, proteins with phosphorylation sites and various biomolecules containing carrying phospho-ligands such as phospho-serine or threonine, were evoked as good candidates to complex uranyl cation (Dedieu et al, 2009).…”