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- Christina Aiu
- Loyola Marymount University
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- I am annotating a paper for a class called "Expression of metallothionein mRNAs by in situ hybridization in the gills of Mytilus galloprovincialis from natural polluted environments." []
The RT-PCR results can be visualized in Figure 3, []which illustrates mRNA expression of MTs in mussels from the Goro and Faro areas. Recall that Goro was characterized by basal conditions while Faro is known to contain high concentrations of environmental contaminants. On the right side of the gel, actin, which is known to remain unaltered by environmental conditions, was used as a loading control. The band intensities of the actin control clearly display that equal amounts of each sample were loaded into the columns. The left side of the gel in Figure 3a identifies the presence of the MT-10 isogene in both areas. From the equal band intensities, authors determined that MT-10 expression was the same in both samples from Goro and Faro. Contrastingly, Figure 3b shows that there was no MT-20 isogene expression in mussels from Goro while mussels from Faro showed elevated MT-20 expression. Again, actin was used as a loading control and the band intensities illustrate the equivalent amounts of sample that were loaded. Figure 5 []which illustrates the results of fluorescent in situ hybridization for MTs on histological sections of Mytilus galloprovincialis, further confirms the results that were obtained from RT-PCR. The fluorescent probe highlights the areas in the tissue cross-sections where MT-10 and MT-20 isogenes were present. Figures 5a and 5b display a clear equivalency in the presence of the MT-10 isogene (highlighted in green) in mussels from both Goro and Faro. However, Figures 5c and 5d show that MT-20 (highlighted in red) was identified only in mussels from Faro and not in those from Goro.
A recent study carried out on the expression of both MT isogenes in specimens of M. galloprovincialis exposed to sublethal concentrations of heavy metals in vivo have shown that the MT-10 gene transcription was inducible by Cd, Zn, Cu, and Hg. The study also showed that MT-20 transcription level was very low under basal conditions while it’s mRNA expression was dramatically increased after Cd and Hg exposure. In fact, after exposure to such metals, MT-20 expression reached levels similar to normal MT-10 expression (Dondero et al., 2005 ) In this study, heavy metal analyses, testing for presence of Pb, Cd, Zn, Cu, and Cr, were performed on lyophilized (frozen) tissues. Metal concentrations were determined by atomic absorption spectrometry using a graphite furnace, a technique that uses a graphite-coated furnace to vaporize a sample and determine light absorbance. If the light absorbance is close to the absorbance of the metal of interest, one can conclude presence of that particular element within the sample. Figure 7  illustrates the results of heavy metal analysis for concentrations (in units mg/Kg d.w.) and standard deviation of Cr, Pb, and Cd detected in the gills of Mytilus galloprovincialis from Faro (polluted environment) and Goro (control environment). From the graph, one can clearly notice that specimens collected from Faro contained significantly larger concentrations of Cr, Pb, and Cd in comparison to Goro (p<0.05). There was a particularly greater difference in concentration of Cr levels between the two sites. Cr also shows a much larger standard deviation than the measurements taken from the other metals. Figure 8 shows the results of heavy metal testing for Zn and Cu. There is also a clear difference here between the concentrations of heavy metals found in Faro in comparison to Goro (<0.05). These results support the hypothesis that the presence of MT’s serve as a possible defense mechanism that allows the organism to maintain homeostasis while under stress from polluted environments. Previous literature also suggests that transcription of MT-20 is possibly Cd-induced (Lemione et al., 2000 ).
- Interest 3
- Goldbeter A and Koshland DE Jr. An amplified sensitivity arising from covalent modification in biological systems. Proc Natl Acad Sci U S A. 1981 Nov;78(11):6840-4.
- JACOB F and MONOD J. Genetic regulatory mechanisms in the synthesis of proteins. J Mol Biol. 1961 Jun;3:318-56.
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- Mark Ptashne. A genetic switch. Cold Spring Harbor, N.Y.: Cold Spring Harbor Laboratory Press, 2004. ISBN:0879697164