Download Developmental Biology BY1101 Cellular differentiation and the

Developmental Biology BY1101
P. Murphy
Lecture 7
Cellular differentiation and the regulation of gene expression.
In this lecture we looked at two main questions:
How is gene expression regulated? (revision of some material in chapters 11, 17 and
18) and
How does this relate to cellular differentiation?
As you heard in lecture 1, Cellular differentiation is one of the 3 main processes needed
to form a complex organism from a single fertilized egg cell (Cell division, cell
differentiation and morphogenesis).
A complex organism requires many hundreds of different cell types to form structures
and carry out specific functions. For example, red blood cells are required to carry
oxygen, muscle cells are required for movement, neurons are required to receive
and transmit nerve signals.
If all the cells arise from a single fertilised egg cell and all contain the same DNA in their
nuclei, how do they become different to each other or ”differentiate”? This is
what we call cellular differentiation.
Cellular differentiation is brought about by differential gene expression: the cells become
different because they express different genes.
e.g. Muscle cells must express the myosin gene so that they have one of the structural
proteins needed (myosin) to enable a muscle fiber to contract and red blood cells must
express globin genes in order to produce haemoglobin to transport oxygen. Red blood
cells do not express the myosin gene and muscle cells do not express globin genes.- these
different cell types follow different differentiation programmes.
So in order to understand how this can be brought about, we recapped on what it means
to express a gene (turn it on) and how the decision to be expressed (or not) is controlled
in a cell.
How are genes turned on and off?
= How are genes regulated?
Some terms revised:
DNA (deoxyribonucleic acid): The substance that constitutes the hereditary material of
an organism. It resides in the nucleus of all eukaryotic cells, organised into linear units
called chromosomes. It is a double stranded polymer of deoxyribonucleotides of which
there are 4 types (see genetic code below)
A gene: A unit of hereditary information consisting of a particular nucleotide sequence of
DNA (generally). Many genes are organised along a chromosome.
(There are many definitions of a gene depending on the perspective you take)
The genetic code: DNA is made up of 4 types of nucleotide: A (adenine), C (cytosine), G
(guanine) and T (thymine). The sequence in which these nucleotides occur determines the
protein that a gene will encode.
Transcription: When a gene is “turned on” its DNA sequence is used as a template for the
synthesis of a complimentary RNA (RiboNucleic Acid) molecule (messenger RNA;
mRNA) by a process called transcription.
mRNA: A single stranded polymer of ribonucleotides produced by transcription of a
gene. It directs the production of a protein during translation. The sequence of
ribonucleotides is complimentary to the DNA sequence being transcribed. (U instead of
T)
Translation: The production of protein from RNA, the sequence of amino acids that make
up the protein depending on the genetic code carried by the RNA.
See Campbell and Reece Fig 17.4 for an illustration of transcription and translation
To view what happens when a gene is “on” (being expressed) in a eukaryotic cell, see
Campbell and Reece figure 17.3.
A gene includes more than coding sequences (sequences that are transcribed and
translated). It also includes regulatory sequences that determine which cells express that
gene and when they turn it on. Remember this when you need to define or describe a
gene; a gene is not just coding sequence but also the regulatory sequences that determine
when and where it will be expressed (turned on).
Regulatory sequences at the start of the coding sequence (Promoters) are needed for the
transcriptional machinery to assemble and begin to transcribe the DNA sequence into an
RNA message or transcript (mRNA). These are similar in all eukaryotic genes.
See Campbell and Reece figure 17.8
Other regulatory sequences are gene specific and these determine when and where a gene
will be turned on.
They can be situated close to the coding sequence (proximal control elements) or at large
distances (distal control elements). These are often called enhancers and can be
positioned upstream (before) or downstream (after) or within the coding sequence (in an
intron)
See Campbell and Reece figure 18.8
______________________________
Transcription initiation is controlled by proteins that interact with DNA (regulatory
sequences) and with each other- see figure 18.11
•
•
These proteins are called transcription factors and operate by binding to the
specific regulatory sequence elements (control elements- enhancers) described
above.
Cell specific transcription factors influence the efficiency with which the general
transcription factors (transcription initiation complex) assemble on the promoter
sequence and initiates expression of the gene.
Distant control elements, enhancers, may be thousands of nucleotides away from the
promoter or even downstream of the gene or within an intron.
Factors that turn a gene on in this way are called activators.
Eukaryotic genes can also be influenced by repressor proteins that bind to DNA
regulatory sequences and tend to destabilize transcription and turn the gene off (can also
be called silencers).
So to recap:
Activator
If the balance is favourable
Transcription
Three important points to note about cellular differentiation:
Point 1
–
Cellular differentiation is usually a result of transcriptional regulation:
turning genes on and off.
Point 2:
–
During embryonic development, cells become obviously different in
structure and function as they differentiate. But differentiation does not
happen suddenly. Differentiation happens progressively as the embryo
develops.
–
–
When differentiated cells appear they already produce the proteins that allow
them to carry out their specialised roles in the organism e.g eye lens cells,
80% of their capacity for protein synthesis makes crystallins.
However changes will be taking place inside a cell long before it visibly
“differentiates”. These include a gradual reprogramming of the genes
that are expressed. This would show up only at the molecular level.
We looked at the example of progressive myoblast differentiation under the control
of the cell specific transcription factor MyoD to illustrate progressive
differentiation. This is well covered in the text book: See Campbell and Reece Fig
18.18 and from bottom of page 414
Point 3:
The genes that encode transcription factors that control cellular differentiation (e.g.
MyoD) are called “Master regulatory genes”. These control the expression of sets of
target genes (downstream genes), the products of which are needed for the cell to
differentiate.
Many of the downstream genes may also be regulatory genes controlling the expression
of more target genes. This is how a cascade event along a differentiation pathway may be
controlled and explains why differentiation is progressive.
These master regulatory genes, or developmental regulators, are the genes of most
interest to developmental biologists.
-----------------------------------------------------This leaves much unexplained
How is the pathway initiated?
How are the master regulators (e.g. MyoD) spurred into action?
How do cells receive instructions about which master regulators to turn on?
We will begin to address these questions in lecture 8
___________________________________________________
Key concepts in lecture 7
1. Cellular differentiation, one of the three major processes that must take place during
development, is brought about by different cells expressing different sets of genes.
2. The genes that are expressed in a cell give it its special characteristics and allow it to
carry out its particular functions, e.g. muscle must contract, neurons must receive and
transmit signals, lens cells must transmit and focus light, and blood cells must transport
oxygen.
3. Consideration of what a gene is and how its expression is regulated is therefore
fundamental to working out how development is controlled. The basic facts about gene
expression were therefore revised.
4. The primary level at which gene expression is controlled is transcription: the decision
about whether or not to make an mRNA copy of the coding sequence of the gene.
5. Regulatory sequences (control regions) outside the coding sequence of the gene
determine when and where a gene is transcribed. They do this by acting as binding sites
for regulatory proteins called transcription factors. Most transcription factors are
activators of transcription but some can act as repressors.
6. The genes that are expressed in a cell therefore depend on the transcription factors that
are present. Muscle structural genes are therefore active in muscle cells because the cells
possess the right transcription factors (e.g. MyoD) to turn them on.
Lecture 7: Learning outcomes: you should be able to
A) Define cellular differentiation and describe its importance during embryonic
development giving examples of cell types that must be established, mentioning
how differential gene expression is the basis of cellular differentiation.
B) Describe how cell specific transcription factors binding to enhancers in the
control regions of genes, regulate the turn on of different genes in different cells.
(N.B. you can use the lac operon in bacteria as an example of gene regulatory
mechanisms but make sure you know that the lac operon operates in bacteria and
is not involved in the differentiation of cells in a complex multicellular organism.)
C) Use the example of muscle differentiation and the experiment used to find the
regulator MyoD to illustrate the importance of gene regulation during
differentiation.
Key terms to be familiar with: differential gene expression, gene regulation, DNA,
gene, mRNA, transcription, translation, regulatory sequences, promoter, enhancer,
transcriptional machinery, transcription factors and cell specific transcription factors,
activators, repressors, myoD, determination, cascade of events, master regulatory genes,