Please Answer ALL questions. Thanks! 13) You have the following mutant strains o
ID: 204984 • Letter: P
Question
Please Answer ALL questions. Thanks!
13) You have the following mutant strains of Escherichia coli. Strain designation lacl- produces a defective lac repressor (does not work). Strain designation lacS produces a lac repressor that cannot bind lactose (allolactose) Strain designation crp+ produces CRP that is always active (always binds DNA) Strain designation cam+ produces an always active adenyl cyclase (always produces cAMP). Strain designation crpS produces CRP that cannot bind cAMP. Describe how you can use these mutants to prove the negative and postive controls of the lac operon (NOTE: It may help to first explain the regulation). 14) p21 is classified as a tumor suppressor in that it prohibits cell division, and is a gene associated with colon cancer. It is normally produced during a cell's division cycle to slow down the process to make sure the cell is healthy enough to divide (think of it as a check & balance system). The gene is not expressed at other times. Chromatin remodelling is the normal means of regulating this gene Trichostatin has been shown to increase expression of p21 through hyperacetylation of histones. Explain the concept of chromatin remodeling and how hyperacetylation of histones can result in arn increased expression of this gene.Explanation / Answer
13. Lac operon is cónsists of genes which are transcribed together It has a promoter region, where RNA polymerase binds, a operator region where the repressor protein binds to the DNA. In addition it has structural genes, lac Z, lacY and lacA. lacZ codes for beta.galactosidase, lacY codes for permease and lacA codes for transacetylase. When RNA polymerase binds to the promoter, the genes are transcribed and the operon is on. However, when repressor binds to the operator region, RNA polymerase can not bind anymore to the promoter region and there is no gene transcription takes place. Then we can say that the operon is in off state. The lac operon is also regulated by CAP protein . The protein is encoded by CRP gene .
Now lets see how lac operon is regulated in the presene and absence of lactose.When lactose is high and glucose is low , then, cAMP binds to inactive CAP protein and makes it active. This CAP protein next binds to the CAP binding region of the DNA which also increse RNA polymerase to bind with the promoter. Now as the lactose which is a inducer bind to the repressor, so repressor can not block the promoter site. As a result the operon is on state and structural genes are transcribed.
On the other hand, when lactose is low and glucose is high, cAMP can not bind to the CAP protein, and RNA polymerase also not bind to the promoter region. Additionally as there is no inducer present the repressor also blocks the promoter site and no transcription occures.
If we no go back the mutants, we can describe the negative regulation by the mutants lacI- and lacS. In lacI- mutant the repressor will not synthesized , as a result the RNA polyerase is able to bind to promoter and lac genes are transcribed. On the other hand as lacS mutant produces a defective repressor, this is also unable to block the promoter site. As a result in this mutant also, lac genes are transcribed. Thus we need active lac respressor protein to negatively regulate the lac operon.
Now, in strain crp+ where CRP is constituvely expressed all the time, the cMAP-CRP will bind to the promoter and together with RNA polymerase will carry in the transcription of the structural genes.
In the strain cam+ which always produce a cAMP which will bind to the CAP protein in the CAP binding side and increase the activaty the RNA polymerase to transcribe the lac genes,
On the other hand crpS produces CRP which can not bind anymore cAMP, thus rhe cAMP-CRP complex can not bind to DNA to activate RNA polymerase , thus the activity of RNA polymerase will decrease.
14. Chromatin remodelling is a method by which the condensed chromatin structure is modified. This modification will allow the DNA to ineract with the transcriptional regulators and as a result gene expression are regulated. In histone acetylation, acetyl groups are added to the lysine residues of the N terminal region of the histone protein by the enzyme histone acetyltransferase. Now, in normal condition, histone has a positive charge which can interact with the negative charge of the positive group present in the phosphate in DNA. So when acetylation occurs it will remove the positive charge from the histone, so it can not interact with the negatively charge phosphate group of DNA. As a result the condensed structure of chromatin becomes relaxed and DNA can interact strongly with the transcription factors to induce gene expression. Thus p21 also get hyperacetylated to increase the expression.