Transport of a neurotoxicant by molecular mimicry: the methylmercury–l-cysteine complex is a substrate for human L-type large neutral amino acid transporter (LAT) 1 and LAT2

Abstract

Methylmercury (MeHg) readily crosses cell membrane barriers to reach its target tissue, the brain. Although it is generally assumed that this rapid transport is due to simple diffusion, recent studies have demonstrated that MeHg is transported as a hydrophilic complex, and possibly as an L-cysteine complex on the ubiquitous L-type large neutral amino acid transporters (LATs). To test this hypothesis, studies were carried out in Xenopus laevis oocytes expressing two of the major L-type carriers in humans, LAT1-4F2 heavy chain (4F2hc) and LAT2-4F2hc. Oocytes expressing LAT1-4F2hc or LAT2-4F2hc demonstrated enhanced uptake of [(14)C]MeHg when administered as the L-cysteine or D,L-homocysteine complexes, but not when administered as the D-cysteine, N -acetyl-L-cysteine, penicillamine or GSH complexes. Kinetic analysis of transport indicated that the apparent affinities ( K (m)) of MeHg-L-cysteine uptake by LAT1 and LAT2 (98+/-8 and 64+/-8 microM respectively) were comparable with those for methionine (99+/-9 and 161+/-11 microM), whereas the V (max) values were higher for MeHg-L-cysteine, indicating that it may be a better substrate than the endogenous amino acid. Uptake and efflux of [(3)H]methionine and [(14)C]MeHg-L-cysteine were trans -stimulated by leucine and phenylalanine, but not by glutamate, indicating that MeHg-L-cysteine is both a cis - and trans -substrate. In addition, [(3)H]methionine efflux was trans -stimulated by leucine and phenylalanine even in the presence of an inwardly directed methionine gradient, demonstrating concentrative transport by both LAT1 and LAT2. The present results describe a major molecular mechanism by which MeHg is transported across cell membranes and indicate that metal complexes may form a novel class of substrates for amino acid carriers. These transport proteins may therefore participate in metal ion homoeostasis and toxicity.

Molecular and ionic mimicry and the transport of toxic metals

“…In recent years, a number of carrier proteins have been implicated in the transport of some toxic metals. In particular, amino acid transporters (i.e., system b 0,+ , system L; Simmons-Willis et al, 2002) and organic anion transporters (i.e., OAT1 and OAT3; Aslamkhan et al, 2003; have been implicated in the absorptive transport of inorganic and organic forms of Hg in renal epithelial cells, endothelial cells and glial cells. Molecular mimicry has been implicated as the primary means for the entry of certain metals via these transporters.…”

“…With this knowledge, it has become possible to identify and characterize further the specific mechanisms involved in the uptake of CH 3 Hg-S-Cys. To illustrate this point, Simmons-Willis et al (2002) utilized oocytes from Xenopus laevis to study directly the involvement of LAT1 and LAT2 in the transport of this conjugate. These investigators provided the first line of direct molecular evidence implicating CH 3 Hg-S-Cys as a transportable substrate of LAT 1 and 2 (Simmons- .…”

Localizing organomercury uptake and accumulation in zebrafish larvae at the tissue and cellular level

“…Quantities of Hg per pixel were calibrated using two certified highly uniform thin film standards on 6.3 m-thick mylar substrates (Micromatter Co.), containing 16.3 and 17.1 g/cm 2 Au and TlCl, respectively. The scatter-corrected Au and Tl L␣1,2 fluorescence intensities were used to interpolate a Hg L␣1,2 fluorescence intensity and this was applied to the scatter-corrected Hg distribution maps to obtain the quantities of Hg per pixel in g/cm 2 . Quantification was not possible for images of intact fish because the thickness is not known accurately.…”

Collate the literature on toxicity data on mercury in experimental animals and humans (Part I – Data on organic mercury)

“…The MeHg-glutathione complex seems to be a substrate for proteins that mediate cellular export of glutathione-S-conjugates, like the multidrug resistance-associated protein 2 (MRP2) (Ballatori, 2002). Simmons-Willis et al (2002) reported that MeHg is probably transported as a hydrophilic complex, and possibly as an Lcysteine complex on the ubiquitous L-type large neutral amino acid transporters (LATs).…”