Dr Tata

Question 1

Why is it important to understand transcription factors in multicellular eukaryotes?

Answer 1

Important to understand how transcription factors aid development from a single cell; how gene expression patterns are maintained and how these factors signal between cells which can be used.

Question 2

Do all cells contain the same genetic information?

Answer 2

NO most cells contain identical genetic information however lymphocytes have each bits added to allow them to produce all the antibodies required within the body. Different cells however use different groups of genes.

Question 3

How are genes regulated in multicellular eukaryotes?

Answer 3

Many different specialised cell types so cells respond to internal environments such as signals from neighbouring cells or via widespread signalling by hormones.

Question 4

How does a hormone transmit a signal? Example?

Answer 4

Example of a ligand. They are produced by one cell and then bind to receptors either in the same cell or a different cell (Ligands include proteins (insulin) and steriods (biochemically derived from cholesterol).

Example is Oestrogen which prepare the chicken oviduct to produce an egg by changing the proteins a cell makes causing a change in physiological behaviour of the cell (Some cells produce ovalbumin (component of egg white), some produce avidin and some aid motility).

Question 5

How do steroid hormones directly regulate gene expression?

Answer 5

The hormone binds to a receptor within the cell causing a conformational change in the receptor. This hormone receptor complex is then able to bind to a specific sequence of DNA. This is an example of a transcription factor.

Question 6

How does signal transduction lead to gene regulation?

Answer 6

Ligand binds to a transmembrane receptor activating the intracellular domain. This leads to a sequence of kinases phosphorylating a number of different molecules within the cell (phospho-kinase cascade). These phosphorylations lead to the activation of a transcription factor which in turn switches a target gene on. This system is very important but if it becomes hyperactive the gene may be switched on permanently.

Question 7

Give examples of transcription factors with specialised domains with DNA.

Answer 7

Hetero and homo dimers (Leucine zipper) which either interact with transcriptional machinery or by modulating chromatin structure.

Question 8

Describe a basic eukaryotic RNA pol II promoter.

Answer 8

Produce RNA for most producing genes. TATA boxes are located 30 bp downstream of the target gene (may be AGAC at +32-+28). They help to position RNA pol II in the correct place before initiating transcription. Different motifs increase TATA box efficiency however they are only basic promoters and are uncontrolled.

Question 9

How do enhancers help in the expression of a gene?

Answer 9

Specific transcription factors bind to enhancers which makes TFIID more likely to bind to TATA boxes. TFIID is a TF which directly regulates expression and interacts with the RNA pol complex. An enhancer sequence may be located a distance from the promoter at either end of the gene; within the gene or orientation independent.

Question 10

How is transcription regulated?

Answer 10

DNA is looped to allow sequences to be brought together.

TF factors may bind repressing transcription.

Complex combinations of TF can set the transcriptional state of a gene.

Question 11

Why do most genes have multiple enhancers and activators?

Answer 11

Cooperativy between TF increase activation and by increasing specificity prevents accidental expression.

Different combinations set different transcription factors allowing genes to be activated by different stimuli/conditions.

Question 12

Describe chromatin

Answer 12

Nucleosome - DNA + histone complex (2 of each- H2A, H2B, H3 and H4);
146bp per histone complex wrapped into 1.67 superhelical turns.
Spacer DNA between nucleosomes is regulated by H1 protein.

Question 13

How is chromosome structure modulated?

Answer 13

Acetyl groups are added to the lysine found on the H4 tail loosening chromatin to allow access for transcription.

Histone acteyltransferase = initiation histone deacetyltransferase = inhibition.

Histones can also be methylated, phosphrylated or ubiquinated.

Question 14

How do transcription factors initiate remodelling?

Answer 14

Promoters are tightly bound in the chromatin so some TF disturb the local DNA-histone interactions allowing other proteins access to further loose the chromatin.

Question 15

How is gene expression inhibited (EPIGENETICS)?

Answer 15

Methylation is used in mammals to prevent transcriptional activation of genes. C is methylated when next to G inhibiting transcription and this is maintained throughout the cell cycle. Ovalbumin - oviduct = methylated all other tissues = unmethylated. Can also be seen in foetal globin gene.

Question 16

Explain autoregulation of a gene

Answer 16

If TFA activates gene A this is known as an autoregulatory loop. This maintains pattern of gene expression after cell division as the cytoplasm of daughter cell has TF indicating the importance of interactions between the nucleus and cytoplasm.

Question 17

Is a transcription factor able to regulate genes which code for regulatory proteins?

Answer 17

YES. This then leads to the inhibition or expression of further genes and amplifies the difference between cells.

Question 18

Describe an experiment used to show the importance of interactions between the nucleus and cytoplasm.

Answer 18

Cell fusion experiments. Rat muscle cell infected by a human liver cell forming a heterokaryon. After the cell has been left for a time, the human muscle genes have been switched on and liver genes switched off showing the importance of proteins produced in the cytoplasm and the effect they have on the genes. Also shows that some genes have been conserved over a long period of time as the rat TF able to bind to human enhancer sequence.

Question 19

How do hormones regulate sex development?

Answer 19

Cause major effects on the behaviour of cells and can be easily observed and manipulated. They also act as switches during development.

In XY humans, at 7 weeks the testis release testosterone which switches on the male-specific genes in target cells diverting development down another route. This can be show experimentally but removal of the testes in male Rabbits who then develop into female rabbits as a result.

Question 20

What is a clone?

Answer 20

Genetically identical cells from a single cell via binary fission;

Replica of DNA sequence reproduced by genetic engineering.

An individual grown from a single somatic cell of a parent and is genetically indentical.

Question 21

What is totipotency?

Answer 21

When a whole new organism can develop from a single cell. This can be seen in plant cells where the information can be easily reversed as all information is retained with the cell.

Question 22

How are animal clones produced?

Answer 22

By the transfer of an adult nucleus into an egg cell where the nucleus has been removed because the egg cell holds all the molecules required to initiate and support early development. (First used on frogs in 1950s).

Question 23

Why is it difficult to clone an adult nucleus?

Answer 23

The fate of cells is determined from an early stage during development so as development progresses alot of changes occur in the expression of genes within a cell making it progressively more restricting. To reveal totipotency must remove all modifications and reprogramme genes. (USE DOLLY THE SHEEP)

Question 24

What consequences were faced after Dolly the Sheep was cloned?

Answer 24

Inefficient and things go wrong on a number of occasions causing neonatal death. DNA taken from 6 year old sheep so had gone through a number of changes and the molecular clock on the chromosome shorter. Decreased fertility due to age of nucleus.

Question 25

What are stem cells?

Answer 25

Unspecialised to specialised (haematopoietic stem cells found in bone marrow produce all blood cell types). Embryonic stem cell - produce all cell types. Cells can be isolated when the embryo has reached the blastocyst stage and the inner cell mass is removed and cultured. They are difficult to then differentiate but is possible by the addition of signalling molecules which alter the pattern of gene expression.