Lecture 7 (2015)Apunte Inglés
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Ingrid Guarro Marzoa
Lecture 7: multilevel transcriptional control in human
DNA-binding domain and activating domain:
The specific transcription factors that bind enhancers (sequences that allow the
gene to be expressed) can bind and activate them because they have two
specific domains: DNA-binding domain and activating domain. The activating
domain can bind the basal complex and with this action transcription would
start. A connecting domain links both domains.
We have to take into account that transcription factors don’t need to be close to the promoter to start transcription because as DNA is a flexible molecule, by making a loop they can contact and activate or repress transcription. Thanks to the capability of these proteins to act in large distance, lots of proteins can bind the basal complex and cooperate and interact among each other to activate or repress gene expression.
The activating domain apart from binding the basal complex has other functions such as: o Histones have positive charge tails and this positive charge help the histones to bind the phosphate groups of DNA. If the tails are modified, the positive charges disappear and the histones go away from DNA allowing it to be transcribed. A function of the activating domains is also to recruit enzymes (co-activators) that will acetylate the tails of histones to relax chromatin, allowing the basal complex to initiate transcription.
o They also can recruit other proteins (co-repressors) that remove the acetyl groups, helping the chromatin to be packed and like this repressing transcription.
• Large number of proteins are involved in transcription: We don’t know the causes of many illnesses but in many tissues, we see that they lose the capability of functioning as they are supposed to function. Often this happens because cells are not able to differentiate properly to fulfil the specific tissue requirements. In many experiments it has been proved that the inactivation of a single transcription factor can cause severe distortions on the function of tissuesin multicellular organisms. Ex: the inactivation of one protein in the genome of a fly caused the duplication of the fly wings.
2014/2015 Molecular biology Ingrid Guarro Marzoa We can see how there are lots of proteins involved in transcription. The expression of a gene depends on the presence or absence of the specific transcriptional factors but this is not the only factor. If the presence or absence of specific transcriptional factors were the only mechanism implied in transcription, a circular argument would be created because the activation of a gene would imply the synthesis of a specific transcription factor which, in turn, would imply the activation of another gene and successively. To solve this circular argument, we need the presence of additional mechanisms.
• How transcription factors are regulated? Different examples: 1. Protein synthesis: the synthesis of a certain transcription factor causes the expression of the gene. Ex: homeoproteins.
2. Protein phosphorylation: there are some transcription factors that can’t bind DNA and they can only do it if they are phosphorylated. Phosphate groups cause strong structural changes that make the protein capable of binding DNA. Ex: HSTF 3. Protein dephosphorylation: there are some transcription factors that can’t bind DNA if they are phosphorylated so they need to be dephosphorylated to be capable of binding DNA.
4. Protein ligand binding: when the protein is outside the nucleus, it needs a special molecule (ligand) that binds the protein and transports it into the nucleus. Once in the nucleus, the protein will be able to bind DNA. Ex: steroid receptors 5. Protein cleavage to release active factor: when a protein is processed by a protease, the specific transcription factor is released and it can bind DNA to start transcription. Ex: sterol response.
6. Protein release by inhibitor: when a specific transcription factor is bound to an inhibitor factor it can’t bind DNA. When this inhibitor factor releases the protein, it can bind DNA and start transcription. Ex: NF-kB 7. Protein change in partner: when a specific transcription factor is bound to an inhibitor factor it can’t bind DNA. When an activating factor binds the protein and the inhibitor factor releases it, the protein can bind DNA and start transcription. Ex: HLH (MyoD/ID).
• How our cells react in front of sexual hormones 2014/2015 Molecular biology Ingrid Guarro Marzoa Steroid hormones are molecules that can cross membranes, entering to the cytosol. They don’t need special proteins outside to transport them. These molecules enter the cell and bind receptor proteins (a class of specific transcription factors) that are within the cell already. The binding of the hormone to the protein is direct. Once the hormone is bound, the receptor proteins can enter to the nucleus and bind DNA to start transcription.
The different receptors are capable of recognizing similar enhancers (sexual hormones) but the difference in bases of the different enhancers is sufficient for the proteins to distinguish one enhancer from each other. Each enhancer will activate different sets of genes and create specific responses.
• HLH proteins: They are specific transcription factors that are important in differentiation of cells during all our life. In tissues that are renewed very fast, we have lots of cells that are being differentiated and this differentiation is regulated by HLH proteins.
HLH proteins with basic DNA-binding domains bind to each other forming a dimer capable of binding DNA. If the HLH protein doesn’t has a basic DNAbinding domain, although it can form a dimer with another HLH protein with a basic DNA-binding domain, the dimer will not have enough affinity and it will not bind DNA.
2014/2015 • Molecular biology Ingrid Guarro Marzoa The algebra of the genes: Specific transcription factors may interact among them to define decision models offering a wide range of functional responses to fulfil extremely complex requirements in eukaryotes. We can find different situations: 1. The two specific transcription factors (A and B) must be present to activate transcription. A has the DNA-binding domain and the activation domain that will be in B.
2. Any of the two specific transcription factors (A or B) is able on itself to activate transcription.Both A and B can recognize the same enhancer and they both contain an activation domain.
3. Factor B doesn’t contain an activation domain and behaves as a repressor by replacing activating factor A at its DNA-binding site.
B can bind the enhancer but doesn’t have an activation domain, and substitutes the specific transcription factor A (that has enhancer and activating domain). Like this, transcription doesn’t occur.
• How can the tissues originate? All our tissues originate from one cell and this process mostly depends on transcription factors.
The scheme proposes that, in the first division of the embryonic cell, the two daughter cells are not exactly the same. Cell A is different from cell B by the only presence of 1 single transcription factor. In the second division, from A we will have two cells that are going to be different as in the first division. The two cells that come from B will have the transcription factors from the mother. Like this the number of transcription factors increases exponentially.
By this simple rule, we end up in three divisions with 8 cells with different combinations of transcription factors. We can obtain like this more than 10.000 different cells with just 25 different transcription factors.
2014/2015 Molecular biology Ingrid Guarro Marzoa Our genome has hundreds of transcription factors so it is not that difficult to produce such a complex organisms like ours.
• Methyl imprinting: Genetic imprinting is a phenomenon in which a gene is either expressed or not expressed in the offspring depending on which parent it is inherited from.
Genes in the X chromosome particularly show this behaviour in females.
Methylation is an important mechanism of transcription regulation.
In the early embryo, genes subject to imprinting are marked by methylation. In this way DNA methylation is used as a mark to distinguish two copies of a gene that may be otherwise identical. This methylation will affect the two strand of DNA and it usually takes place in promoters. In most cases, the methyl imprints silences nearby gene expression (acting as repressor). In some cases, the methyl imprint can activate expression of a gene.
Normally what happens is that the enzymes that add methyl groups are only expressed in the egg cells (germ cells) in females. A random combination will generate an embryo that will only have methyl groups that come from the female (maternal imprinting).
In other genes, the methylations occur only in the male germ cells (parental imprinting). So methylation is maintained over successive cell divisions and explains maternal or paternal genetic dominance. In these cases, the Mendel’s laws don’t work.
• Stability of mRNAs: The stability of mRNAs is a mechanism of gene control at a post-transcrptional level.
We can distinguish 3 different types of mRNAS: • Stable mRNA: If the mRNA has a final destination, it’s better to have an mRNA very stable because the cell will be able to use this mRNA to produce proteins for long periods of time.
• Unstable mRNA: If mRNA codes for a growth factor (proteins segregated by the immune cells that tell the cells that they have to proliferate), it is usually an unstable mRNA. When you want mRNA for a transient response, those mRNAs are very unstable. If this mRNA was stable, proliferation would occur during too much time and it could produce tumours.
2014/2015 • Molecular biology Ingrid Guarro Marzoa DNA synthesis modulates mRNA stability: a very extreme example is the histone mRNA. The requirements of histones of a cell are very special. Depending on the situation of the cell, you will need more or less histones. And that’s why the histone mRNAs only last 12 minutes when it’s not synthetizing histones but if it’s synthetizing histones, it last until 1 h so the DNA synthesis modulates its stability.