Expression of constitutive and metal-inducible metallothioneins in oyster embryos (Crassostrea virginica)

doi:10.1016/0305-0491(93)90127-Q

Comparative Biochemistry and Physiology Part B: Comparative Biochemistry

Volume 106, Issue 3, November 1993, Pages 523-529

https://doi.org/10.1016/0305-0491(93)90127-Q Get rights and content

Abstract

1.
1. Putative Zn and CuZn metallothioneins (MTs) were identified from control oyster embryos (Crassostrea virginica). These constitutive MT isoforms are probably involved in Zn and Cu homeostasis during development.
2.
2. During single metal exposures (Cd or Cu), additional MT isoforms (Cd MT and CuCd MT) were induced that probably function in detoxification and sequestration.
3.
3. When embryos were exposed to Cd and Cu simultaneously, inhibition of Cd MT expression suggests that there are complex competitive interactions that affect metal-specific regulation of transcription.

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Metallothionein role in the kinetic model of copper accumulation and elimination in the clam Ruditapes decussatus
2009, Environmental Research
In order to clarify the role of metallothioneins (MT) in copper (Cu) toxicity, this work aimed to assess the involvement of this protein in the accumulation and elimination strategies of Cu in the clam Ruditapes decussatus exposed to two sublethal concentrations (25 and 50 μg Cu l⁻¹). The behaviour of MT in three different tissues of clams during the accumulation and depuration processes was also followed by gel-filtration chromatography to assess if Cu was bound to MT or to other cytosolic components.
The 96 h LC₅₀ for water-borne copper was 715 μg L⁻¹ in R. decussatus. The Cu accumulation pattern was dependent on Cu exposure concentrations. In clams exposed to 25 μg l⁻¹, total Cu accumulation in the three tissues increased linearly during the exposure period, while in those exposed to 50 μg l⁻¹ it followed the first order kinetic model. The greatest amount of Cu accumulated in all tissues is associated to the low molecular weight cytosolic fraction (>50%). The chromatographic assay indicated that Cu in the cytosolic fraction is bound to MT and MT levels increase with the increase of Cu exposure confirming the binding affinity of Cu to MT in all tissues. However, a smaller percentage of Cu seems to be bond to other ligands, such as GSH. Copper was exponentially eliminated (only studied in clams exposed to 25 μg l⁻¹) and the estimated half-life was tissue dependent (9, 5 and 14 days for the gills, digestive gland and remaining tissues, respectively). Copper bound to the thermostable compounds was eliminated more quickly (t_1/2=4–7 days) in all tissues than those bound to the thermolabile compounds (t_1/2=7–18 days). Interestingly, MT is rapidly degraded (t_1/2=7 and 18 days), suggesting that this protein is actively involved in the elimination of this metal, through the Cu–MT complex since MT and Cu are turning over simultaneously.
Therefore, when Cu exposure is low, the clam can cope efficiently with the excess of Cu levels by increasing MT induction as well as rapidly eliminating this metal via the MT–Cu complex. Copper toxicity in the clam R. decussatus is associated to the limited capacity of MT induction at higher and environmental unrealistic Cu exposures especially in the gills and remaining tissues.
Metallothioneins in aquatic invertebrates: Their role in metal detoxification and their use as biomarkers
2006, Aquatic Toxicology
The literature on metallothioneins (MT) and metallothionein-like proteins (MTLP) in aquatic invertebrates is large and increasing, and yet inconsistencies and confusion remain, not least over the physiological role of MT and their use as biomarkers in environmental monitoring programmes. We have collated published information on MT in three important groups of aquatic invertebrates—the molluscs, crustaceans and annelid worms, and attempted to seek explanations for some of the apparent inconsistencies present in the dataset. MTs can be induced by the essential metals Cu and Zn and the non-essential metals Cd, Ag and Hg in both vertebrates and invertebrates, but their induction is variable. Such variation is intraspecific and interspecific, and is down to a variety of reasons environmental and physiological explored here. Against this background of variability MTs do appear to play roles both in the routine metabolic handling of essential Cu and Zn, but also in the detoxification of excess amounts intracellularly of these metals and of non-essential Cd, Ag and Hg. Different isoforms of MT probably play different physiological roles, and the dependence on MT in detoxification processes varies environmentally and between zoological groups. MTs can be used as biomarkers if used wisely in well-designed environmental monitoring programmes.
Glutathione and metallothionein status in an acute response by Mercenaria mercenaria brown cells to copper in vivo
2002, Ecotoxicology and Environmental Safety
Red glands of Mercenaria mercenaria comprise brown cells that accumulate, detoxify, and excrete copper. Brown cell involvement in metal detoxification is due in part to endogenous glutathione (GSH) and metallothionein (MT). The intent of this study was to test the hypotheses that brown cell GSH functions in protection against Cu2²⁺ toxicity, that brown cell GSH provides the initial defense against Cu2²⁺ prior to brown cell MT induction, and that MT variants (MTI, MTII), if present are unequal in response to Cu2²⁺. Brown cells were analyzed for GSH and MT after 0.25, 1, 2, 3, and 4 days of treatment of Mercenaria with 0.01 and 0.05 ppm Cu2²⁺. Glutathione initiated the brown cell acute response (within the first day of treatment) to both the 0.01 and 0.05 ppm Cu2²⁺ treatments. Metallothionein in brown cells increased to Day 4 during treatment with 0.01 ppm Cu2²⁺, whereas MT concentration was greatest at Day 2 after which it decreased to Day 4 with treatment of 0.05 ppm Cu2²⁺. The change in MTII relative to its control was greater than that of MTI in the brown cell acute response to 0.01 ppm Cu2²⁺ and also for Days 0.25 to 2 in response to the 0.05 ppm Cu2²⁺ treatment. At Days 3 and 4 with the 0.05 ppm Cu2²⁺ the change in MTI/MTII ratio was due to a greater change in MTI than MTII relative to their respective controls. The variants of brown cell MT appear to respond differently to Cu2²⁺ depending on the Cu2²⁺ treatment concentration.
Effects of glutathione depletion on copper cytotoxicity in oysters (Crassostrea virginica)
2000, Aquatic Toxicology
Glutathione is a tripeptide that plays an important role in ameliorating metal toxicity. Depletion of glutathione has been associated with an increased risk of metal toxicity in mammals. An understanding of the toxicological significance of glutathione depletion in oysters would be of considerable importance given the widespread use of bivalves in biological monitoring. Laboratory studies were conducted by using an inhibitor of glutathione synthesis (buthionine sulfoximine) to investigate the effects of glutathione depletion on metallothionein expression, lysosomal membrane destabilization, and lipid peroxidation in Cu-exposed oysters. In oysters exposed to Cu (20 and 80 μg/l) and buthionine sulfoximine (20 mg/l), metallothionein induction was suppressed and cellular stress responses were frequently higher than those observed in oysters exposed singly to Cu. Together, these results suggest that environmental conditions that cause glutathione depletion may increase the potential for adverse effects to pollutants during in situ exposures.
In vivo metallothionein and glutathione status in an acute response to cadmium in Mercenaria mercenaria brown cells
2000, Comparative Biochemistry and Physiology - C Pharmacology Toxicology and Endocrinology
Brown cells that are found in the red glands of Mercenaria mercenaria accumulate, detoxify and excrete cadmium. Brown cell involvement in metal detoxification was due in part to endogenous glutathione (GSH) and protein sulfhydryl. Metallothionein (MT) and GSH have been shown to play an important role in metal detoxification in bivalve molluscs. This study showed that the protein sulfhydryl in brown cells of Mercenaria was in fact MT, that brown cell GSH functioned in acute protection against Cd²⁺ toxicity, that GSH provided the initial defense against Cd²⁺ toxicity prior to MT induction and that MT variants were unequal in response to Cd²⁺. During treatment of Mercenaria with 0.5 and 1.0 ppm Cd²⁺, brown cells were analyzed for MT by capillary electrophoresis and GSH colorimetrically after 0.25, 1, 2, 3, and 4 days. The data indicated that the cadmium-binding protein was MT with an apparent molecular weight of 9 kDa determined by gel filtration or 6 kDa as indicated by capillary electrophoresis. Glutathione appeared to prevail in the brown cell acute response to 0.5 ppm Cd²⁺, whereas MT appeared to prevail in the acute response to 1.0 ppm Cd²⁺. Capillary electrophoresis can be used to monitor and quantify MT and its variants in brown cells without need for prior separation of cytosolic components by chromatography. The change in MT-II was greater relative to the change in MT-I in the brown cell acute response to 0.5 ppm Cd²⁺, whereas the change in MT-I was greater relative to the change in MT-II in the acute response to 1.0 ppm Cd²⁺. The variants of brown cell MT appeared to respond differentially to Cd²⁺ depending upon the Cd²⁺ treatment concentration.
Accumulation and cytosolic partitioning of metals in the American oyster Crassostrea virginica
1999, Marine Environmental Research
The ability of the oyster to accumulate trace metals, particularly the transition metals (Cd, Cu, Zn, and Hg), from food and water has been well documented. There is little information, however, on how these metals interact to affect whole animal retention, partitioning, and binding to metallothionein (Mt). In this study oysters were exposed to Cd and Cu, both alone and in combination, and significant effects have been demonstrated on subcellular partitioning. These studies showed that Cu can displace Cd from Mt, but the Cd is not lost from the animal. Two metal-binding protein peaks have been separated by gel chromatography (<10 K and 24 K daltons). The lower molecular weight protein peak has been characterized as Mt, consisting of two isoproteins, but the higher molecular weight protein peak appears to be an aggregation of metal-binding proteins, some of which may be Mt or a Mt dimer. In environments that are contaminated with trace metals, Mt binds primarily Hg, Cu, and/or Cd and Zn, but in noncontaminated areas Mt binds primarily Zn on one of the two isoMts.

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^∗: Present address and requests for reprints: Marine Resources Research Institute, P.O. Box 12559, Charleston, SC 29412, U.S.A.

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Expression of constitutive and metal-inducible metallothioneins in oyster embryos (Crassostrea virginica)

Abstract

Devl Biol.

J. Inorg. Biochem.

Archs Biochem. Biophys.

Mar. Environ. Res.

Comp. Biochem. Physiol.

Adv. Mar. Biol.

Mar. Poll. Bull.

Devl Biol.

J. biol. Chem.

Comp. Biochem. Physiol.

Devl Biol.

Archs Biochem. Biophys.

Biochim. biophys. Acta

Mar. Poll. Bull.

Metallothionein and metallothionein-like proteins: physiological importance

Adv. comp. Environ. Physiol.

Purification and characterization studies of cadmium-binding proteins from the American oyster, Crassostrea virginica

Environ. Hlth Perspect.