分子生物学教学课件：CHAPTER 17 Gene Regulation in Eukaryotes

1、 A lot more regulator bindings sites in multicellular organisms reflects the more extensive signal integration Fig. 17-1 The regulatory elements of a bacterial, yeast, and human gene. Insulators nMechanisms of Eukaryotic Regulators nSignal Transduction and the Control of Transcriptional Regulators n

2、Gene “Silencing” by Modification of Histones and DNA nEukaryotic Gene Regulation at Steps after Transcription Initiation Structure of Eukaryotic Regulators Transcriptional Activation Transcriptional Repressors Structure of Eukaryotic Regulators 1. The more elaborate transcriptional machinery 2. The

3、nucleosome and their modifiers typical of eukaryotes Activators/Repressors nSeparated DNA Binding and Activation Functions nDNA-Binding Domains nActivating Regions Activators Have Separate DNA Binding and Activation Functions Gal4 is the most studied eukaryotic activator Gal4 activates transcription

4、 of the galactose genes in the yeast S. cerevisae. Gal4 binds to four sites upstream of GAL1, and activates transcription 1,000-fold in the presence of galactose Fig; 17-3 The regulatory sequences of the yeast GAL1 gene. N-terminal region (DNA- binding domain) Domain swap experiment) Domain swap exp

5、eriment Moving domains among proteins, proving that domains can be dissected into separate parts of the proteins. Many similar experiments shows that DNA binding domains and activating regions are separable. Bactrial regulatory proteins 1.Most use the helix-turn-helix motif to bind DNA target 2.Most

6、 bind as dimers to DNA sequence: each monomer inserts an a helix into the major groove. Eukaryotic regulatory proteins 1.Recognize the DNA using the similar principles, with some variations in detail. 2.Some form heterodimers to recognize DNA, extending the range of DNA-binding specificity. DNA bind

7、ing domains Zinc containing DNA-binding domains finger domain: Zinc finger proteins (TFIIIA) Zinc cluster domain (Gal4) Leucine Zipper Motif: The Motif combines dimerization and DNA-binding surfaces within a single structural unit. Dimerization is mediated by hydrophobic interactions between the app

8、ropriately-spaced leucine to form a coiled coil structure Helix-Loop-Helix motif: Helix-loop-helix proteins. An extended helical region from each of two monomers insets into the major groove of the DNA. Because the region of the a-helix that binds DNA contains baisc amino acids residues, Leucine zip

9、per and HLH proteins are often called basic zipper and basic HLH proteins. Both of these proteins use hydrophobic amino acid residues for dimerization. The activating regions are grouped on the basis of amino acids content Acidic activation domains Glutamine-rich domains Proline-rich domains nMechan

10、isms of Eukaryotic Regulators Structure of Eukaryotic Regulators Mechanisms of Eukaryotic Activator Mechanisms of Eukaryotic repressor nSignal Transduction and the Control of Transcriptional Regulators nGene “Silencing” by Modification of Histones and DNA nEukaryotic Gene Regulation at Steps after T

11、ranscription Initiation Mechanisms of Eukaryotic Activator Directly fuse the bacterial DNA- binding protein LexA protein to the mediator complex Gal11 to activate GAL1 expression. 2. Activators also recruit Nuleosome modifiers that help the transcription machinery bind at the promoter Two types of N

12、ucleosome modifiers : 1. Those add chemical groups to the tails of histones, such as histone acetyl transferases (HATs) 2. Those remodel the nucleosomes, such as the ATP-dependent activity of SWI5/SNF Modification of the N-terminal tails of the histones Modification of the histone N- terminal tails

13、alters the function of chromatin Fig 7-35 Nucleosome movement catalyzed by nucleosome remodeling complexes remodling Effect of histone tail modification 3. Action at a distance: loops and insulators Insulators block activation by enhancers Specific elements called insulators control the actions of a

14、ctivators, preventing the activating the non-specific genes Transcriptional Silencing nSilencing is a specializes form of repression that can spread along chromatin, switching off multiple genes without the need for each to bear binding sites for specific repressor. nInsulator elements can block thi

15、s spreading, so insulators protect genes from both indiscriminate activation and repression. nMechanisms of Eukaryotic Regulators Structure of Eukaryotic Regulators Mechanisms of Eukaryotic Activator Mechanisms of Eukaryotic repressor nSignal Transduction and the Control of Transcriptional Regulator

16、s nGene “Silencing” by Modification of Histones and DNA nEukaryotic Gene Regulation at Steps after Transcription Initiation Signal Integration and Combinatorial Control 1. integrate signals nIn eukaryotic cells, numerous signals are often required to switch a gene on. So at many genes multiple activ

17、ators must work together. nThey do these by working synergistically: two activators working together is greater than the sum of each of them working alone. Three strategies of synergy : 1.Two activators recruit a single complex 2. Activators help each other binding cooperativity 3. One activator rec

18、ruit something that helps the second activator bind a.“Classical” cooperative binding b. Both proteins interacting with a third protein c. A protein recruits a remodeller to reveal a binding site for another protein d. Binding a protein unwinds the DNA from nucleosome a little, revealing the binding

19、 site for another protein nSignal integration: the HO gene is controlled by two regulators; one recruits nucleosome modifiers and the other recruits mediator nSignal integration: Cooperative binding of activators at the human b-interferon gene. The examples of integrate signals alter the nucleosome

20、Active only at correct stage of cell cycle NFB, IRF, and Jun/ATF They bind cooperatively to sites within an enhancer, form a structure called Enhanceosome. nCombinatory control lies at the heart of the complexity and diversity of eukaryotes nCombinatory control of the mating-type genes from S. cerev

21、isiae 2.Combinatory control In complex multicellular organisms, combinatorial control involves many more regulators and genes than shown above, and repressors as well as activators can be involved. Four signals Three signals nThe yeast S.cerevisiae exists in three forms: two haploid cells of differe

22、nt mating types a and a and the diploid formed when an a and an a cell mate and fuse. nCells of the two mating types differ because they express different sets of genes : a specific genes and a specific genes. Combinatory control of the mating-type genes from S. cerevisiae na cell make the regulator

23、y protein a1,a cell make the protein a1 and a2. A fourth regulator protein Mcm1 is also involved in regulatory the mating-type specific genes and is present in both cell types. nIn eukaryotes, repressors dont work by binding to sites that overlap the promoter and thus block binding of polymerase, bu

24、t most common work by recruiting nucleosome modifiers. nFor example, histone deacetylases repress transcription by removing actetyl groups from the tails of histone Transcriptional Repressors Ways in which eukaryotic repressor Work a and b Ways in which eukaryotic repressor Work c and d Figure 17-19

25、 nIn the presence of glucose, Mig1 binds a site between the USAG and the GAL1 promoter. By recruiting the Tup1 repressing complex, Mig1 represses expression of GAL1. nTwo mechanisms have been proposed to explain the repressing effect of Tup1. First, Tup1 recruits histone deaxetylases. Second, Tup1 i

26、nteracts directly with the transcription machinery at the promoter and inhibits initiation. nMechanisms of Eukaryotic Regulators nSignal Transduction and the Control of Transcriptional Regulators nGene “Silencing” by Modification of Histones and DNA nEukaryotic Gene Regulation at Steps after Transcr

27、iption Initiation Structure of Eukaryotic Regulators Transcriptional Activation Transcriptional Repressors 1 .Signals are often communicated to transcriptional regulators through signal transduction pathway 2 .Signals control the activities of eukaryotic transcriptional regulators in a variety of wa

28、ys 1.signal transduction involve STAT The JAK/STAT Signaling PathwayThe JAK/STAT Signaling Pathway 2. The MAPK signalling pathways The RAS-activated MAPK pathway: RAS-RAF-MEK-MAPK represents the first example where all the steps in a complete signalling cascade from the cell surface receptor PTK(pro

29、tein tyrosine kinase ), to the nuclear transcription is known. Signals control the activities of eukaryotic transcriptional regulators in a variety of ways Once a signal has been communicated, directly or indirectly, to a transcriptional regulator, how does it control the activity of that regulator

30、? In eukaryotes, transcriptional regulators are not typically controlled at the level of DNA binding. They are usually controlled in one of two basic ways : nUnmasking an activating region n Transport in or out of the nucleus Activator Gal4 is regulated by masking protein Gal80 The signalling ligand

31、 causes activators (or repressors) to move to the nucleus where they act from cytoplasm. nMechanisms of Eukaryotic Regulators nSignal Transduction and the Control of Transcriptional Regulators nGene “Silencing” by Modification of Histones and DNA nEukaryotic Gene Regulation at Steps after Transcript

32、ion Initiation Gene “silencing” is a position effecta gene is silenced because of where it is located, not in response to a specific environmental signal. The most common form of silencing is associated with a dense form of chromatin called heterochromatin. It is frequently associated with particular regions of the chromosome, notably the telomeres, and the centromeres. nGene “Silencing” by Modification of Histones and DNA