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Lecture

20. Translational and Post translational regulation.pdf

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Department
Biology (Sci)
Course
BIOL 200
Professor
Richard Roy
Semester
Fall

Description
Naveen Sooknanan McGill Fall 2011 Translational and Post-translational Regulation of Gene Expression: Logic would dictate that if there are more mRNA molecules within a cell, this should correlate to more proteins being synthesized. This is not, however, always the case.  While a Northern blot shows equal presence of a certain mRNA molecule over a period of time, a Western blot showing corresponding protein to this mRNA shows a decline in population over time o Details of Western blotting will be described later o This is because protein synthesis is under strict regulatory control by a series of cellular mechanisms o Quantative analysis, such as the creating of GFP fusion proteins, can determine abundance of any protein within a cell A first method of regulation can occur during translation by altering the stability of the mRNA molecules. An example is the mRNA for mammalian transferrin receptor, which is responsible for importing iron into the cell. The stability of mammalian mRNA molecules coding for the mammalian transferrin receptor (TfR) is mediated by an iron response element binding protein (IRE-BP)  Transferrin receptor does not need to be in high levels when the cell is saturated with iron, but needs to be in excess when the cell is short of iron o Regulation of mRNA stability is responsible for this fluctuation  IRE-BP, has two conformations which depend on iron concentration within the body o It is inactive in high iron concentration o It is active in low iron concentration  TfR mRNA contains AU rich sequence with secondary structure on its 3’ UTR called iron response elements (IREs) o These IREs are capable of binding IRE-BPs  In high iron concentrations, the IRE-BPs are inactive and cannot bind to the mRNA o The mRNA is therefore degraded by exonucleases and the transferrin receptor proteins cannot be expressed  In low iron concentration, there IRE-BPs become activated and bind to the mRNA’s IREs o This prevents exonucleic decay thus stabilizing the mRNAs and allowing higher expression of transferrin within the cell  This is an important physiological mechanism because, while iron is needed in the cell, it can also be toxic if allowed to flow freely in high concentrations o Iron concentration is therefore very highly monitored and kept within a very narrow range Another regulation mechanism involving the inhibition of translation can be seen using another iron regulation system through the ferritin mRNA. Ferritin is a storage unit which is able to bind free floating iron ions to prevent toxic effects.  Contrarily to transferrin, it is needed in high concentrations in high iron concentrations and is not needed in low iron concentrations 1Naveen Sooknanan McGill Fall 2011  In this case, IREs (same ones as before) are located in the 5’ UTR of the ferritin mRNA  When iron levels are high in the cell, IRE-BP (also same one as before) is inactive and does not bind to the IREs o Thus, when translation machinery sits down on the 5’ cap, EIF4A is able to quickly remove these IREs through its helicase activity and translation of ferritin can proceed at high levels o When ferritin is produced, it quickly enters the bloodstream and binds to any free floating iron ions it sees  When iron levels are low in the cell, IRE-BP becomes active and binds to the IREs o This time, cap binding proteins which activate translation cannot sit down on the 5’ cap, and therefore translation of ferritin is strongly inhibited in low iron concentrations o This prevents iron absorption which assures that cells have adequate iron to perform their cellular processes Inhibition of translation can also be seen in development, particularly well in the Drosophila (fruit fly) embryo. A simplified embryo contains two mRNAs which compete for protein expression: Nanos and hunchback  Both mRNAs are equally diffused throughout the embryo, meaning some translational mechanism much be regulating protein expression  It can be seen in wild-type embryos that Nanos is predominate in the anterior (front) section while hunchback is found most in the posterior (back) o This is because of an inhibition mechanism taking place between Nanos and the hunchback mRNA o This causes a gradient of Nanos and hunchback protein over the length of the embryo  In the anterior end, Nanos mRNA undergoes translation and Nanos protein is produced. This protein then acts as an inhibitor for hunchback mRNA, which cannot produce its protein o This explains the lower expression of hunchback in the posterior  In the posterior end, however, Nanos protein isn’t present, meaning that hunchback mRNA is free to translate  This has been proved in vitro by removing Nanos from the entire embryo, which causes hunchback to be present in equal concentrations all over the cell o This causes death of the embryo Post-translational regulation can occur by a wide array of mechanisms through protein folding, protein modification and protein stability  Protein folding mechanisms include: o Phosphorylation, addition of phosphate ions o Glycosylation, addition of sugar moieties 2Naveen Sooknanan McGill Fall 2011 o Lipid moieties  Help with the protein’s ability to interact with the lipid membrane o Small peptides  Protein folding occurs with the help of chaperones, which were stu
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