3.8 The control of gene expression

Cells are able to control their metabolic activities by regulating the transcription and translation of their genome. Although the cells within an organism carry the same coded genetic information , they translate only part of it. In multicellular organisms, this control of translation enables cells to have specialised functions, forming tissues and organs. There are many factors that control the expression of genes and, thus, the phenotype of organisms. Some are external, environmental factors, others are internal factors. The expression of genes is not as simple as once thought, with epigenetic regulation of transcription being increasingly recognised as important. Humans are learning how to control the expression of genes by altering the epigenome, and how to alter genomes and proteomes of organisms. This has many medical and technological applications. Consideration of cellular control mechanisms underpins the content of this section. Students who have studied it should develop an understanding of the ways in which organisms and cells control their activities. This should lead to an appreciation of common ailments resulting from a breakdown of these control mechanisms and the use of DNA technology in the diagnosis and treatment of human diseases.

What are mutations?

•Mutations are changes in the base sequence of DNA that can occur during DNA replication or due to external factors.

•These changes can affect the structure and function of proteins and, in some cases, the phenotype of an organism.

When do mutation occur?

Usually during DNA replication

Substitution Mutations

A single base is replaced by another.

What does substitution mutation lead to? (3 types of mutation)

•Silent mutation: No change in the amino acid due to the degenerate nature of the genetic code.

•Missense mutation: A different amino acid is produced, potentially altering the protein's structure and function.

•Nonsense mutation: The mutation introduces a stop codon, resulting in a truncated, non-functional protein.

Insertion mutations

One or more nucleotide bases are added to the sequence.

Deletion mutation

One or more nucleotide bases are removed from the sequence.

What is the result of insertion and deletion mutations?

Both cause a frameshift mutation, altering the reading frame of the genetic code, which typically leads to a completely different and non-functional protein.

Duplication Mutations

One or more bases are repeated, leading to a frameshift and abnormal protein production.

Inversion Mutations

•A segment of DNA is removed, reversed, and reinserted into the same location.

•This alters the order of bases, potentially affecting the amino acids coded.

Translocation Mutations

•A segment of DNA from one chromosome is removed and inserted into another chromosome.

•Often results in major changes to the organism's phenotype and can disrupt gene regulation.

What is the causes of mutations?

Spontaneous Mutations

Induced Mutations

Spontaneous Mutations

Random change in the DNA due to errors in replication that occur without known cause, so its natural.

Induced Mutations

•Radiation: UV or ionising radiation can break DNA strands or alter bases.

•Chemicals: Mutagens like benzene or asbestos can cause base changes or DNA damage.

•Viruses: Some viruses insert their genetic material into the host DNA, causing mutations.

What are the 3 effects of mutations?

-Neutral Mutations

-Harmful Mutations

-Beneficial Mutations

What is Neutral Mutations and when does it occur?

•No change to the organism's phenotype or survival.

-Occurs when:

•The mutation is in a non-coding region of DNA (e.g., introns).

•The mutation is a silent mutation, where the codon changes but the amino acid remains the same due to the degenerate nature of the genetic code.

•The protein's tertiary structure is altered, but its function is unaffected.

What are Beneficial Mutations

•Provide an advantage to the organism, improving survival or reproduction.

(Example: The evolution of trichromatic vision in humans allows better differentiation of colours, advantageous in identifying food and avoiding danger.)

-Beneficial mutations are a key driver of natural selection and evolution.

What are Harmful Mutations

Result in a disadvantage to the organism, often leading to disease or reduced survival.

What increases mutations rate?

Mutagenic agents e.g. UV light

Not all such mutations result in a change to the encoded amino acid, Explain why?

•Due to the degenerate nature of the genetic code, not all such mutations result in a change to the encoded amino acid.

•Some gene mutations change the nature of all base triplets downstream from the mutation, i.e result in a frame shift.

What are Stem Cells?

Stem cells are undifferentiated cells capable of

-:Self-renewal: Continuously dividing to produce more stem cells.

-Differentiation: Giving rise to specialised cell types.

What are totipotent cells?

Totipotent cells are a type of stem cell that can differentiate into:

•Any cell type found in the body (e.g., muscle, nerve).

•Extra-embryonic cells, such as those forming the placenta.

Where are they found?

Found in the early embryo, so they are limited timed.

How do totipotent cells become specialised?

•Initially unspecialised: Totipotent cells do not have specific functions when first formed.

•Then totipotent cells translate only part of their DNA, resulting in cell specialisation (Selective gene expression)

Where are Pluripotent Stem Cells found and what is its function?

•Found in the later stages of the embryo.

•Can differentiate into many types of specialised cells, but not extra-embryonic cells.

Why are Pluripotent Stem Cells able to treat diseases such as leukaemia

Because Pluripotent stem cells can divide in unlimited numbers

Where are Multipotent Stem Cells found and what is its function?

•Found in adult tissues (e.g., bone marrow).

•Can differentiate into a limited range of cell types
(E.g: Haematopoietic stem cells give rise to different blood cells.)

Where are Unipotent Stem Cells found and what is its function?

-Found in specific tissues.

-Can differentiate into only one type of cell.

-For example:

Unipotent cells, like cardiac progenitor cells, can only differentiate into one cell type, such as cardiomyocytes. Signals like FGF and BMP activate genes (e.g., Nkx2.5, GATA4) to guide this process, aiding limited heart tissue repair.

Induced Pluripotent Stem Cells (iPS)

-iPS cells are artificially created pluripotent cells generated from unipotent cells (e.g., skin cells).

-This process involves reprogramming genes to make the unipotent cells behave like pluripotent stem cells.

Where are Induced pluripotent stem cells (iPS cells) produced from

Induced pluripotent stem cells (iPS cells) can be produced from adult somatic cells using appropriate protein transcription factors.

Advantages of iPS cells:

-Avoids ethical concerns surrounding embryonic stem cells.

-Can be derived from the patient's own cells, reducing the risk of rejection.

Ethical Issues and Challenges

-Use of embryonic stem cells raises ethical concerns due to the destruction of embryos.

-iPS cells offer a more ethically acceptable alternative but are still under research to ensure their safety and efficacy.

What is Tissue Culture

-Technique used to grow plants or plant tissues in controlled conditions from explants, which are small pieces of plant tissue.

-This process is used for mass-producing plants or for preserving rare species.

Evaluate the use of stem cells in treating human disorders (Benefits)

-Regenerative Medicine: Stem cells can replace damaged tissues, e.g., producing insulin-secreting cells for diabetes or dopamine-producing neurons for Parkinson's disease.

-Tissue and Organ Repair: Embryonic stem cells (pluripotent) can form any tissue type, offering treatments for conditions like spinal cord injuries or heart disease.

-Drug Testing: Stem
cells allow drug safety and efficacy testing on specific tissues, reducing the need for animal models.

-Bone Marrow Transplants: Hematopoietic stem cells (multipotent) are already used to treat blood disorders like leukemia.

Evaluate the use of stem cells in treating human disorders (Limitations)

-Rejection Risk: Transplanted cells may provoke immune responses, requiring immunosuppressants.

-Limited Success: Current therapies, like using induced pluripotent stem cells (iPSCs), are still experimental and not fully reliable.

-Tumor Formation: Stem cells can proliferate uncontrollably, leading to tumor risks.

-Ethical Concerns: Harvesting embryonic stem cells destroys embryos, sparking debates about the moral status of early life forms.

Stages of Producing Tissue Cultures

1.) Selection of Explants:

-An explant is a small piece of tissue taken from a parent plant, such as a leaf, stem, or root.The explant is typically taken from a plant with desirable traits (e.g., disease resistance or high yield).

2.) Sterilisation:

-The explant is sterilised using substances like ethanol or sodium hypochlorite to remove microorganisms.

3.) Placement in Growth Medium:

-The sterilised explant is placed in a sterile nutrient medium containing:

-Glucose: for energy.
-Agar: as a solid support.
-Minerals -Essential nutrients for plant growth.
-Plant hormones (e.g., auxins and cytokinins) to stimulate cell division and differentiation.

4.) Callus Formation:

-The explant cells divide by mitosis, forming a callus, which is a mass of undifferentiated cells.

Regeneration of Shoots and Roots:

-By altering the concentration of hormones, the callus is encouraged to form:

-Shoots (using cytokinins).
-Roots (using auxins).

Transfer to Soil:
Once small plants (plantlets) have developed, they are transferred to sterile soil or compost in controlled conditions to continue growing.

What is tissue culturing used for?

1.) Cloning Plants:
-To produce genetically identical plants with desirable traits such as disease resistance or high yield.

2.) Preservation of Rare Species:
-Used to conserve endangered plants by creating large populations.

3.) Crop Improvement:
-Allows for rapid multiplication of genetically modified plants.

4.) Pathogen-Free Plants:
-Produces plants free from viruses and bacteria for agriculture.

Advantages of Tissue Culture

-Rapid multiplication of plants in a short time.

-Can be carried out all year round, regardless of season.

-Allows for the production of genetically uniform plants.

Disadvantages of Tissue Culture

-Requires specialised equipment and skills, making it costly.

-Risk of contamination, which can lead to loss of cultures.

-Produces genetically identical plants, making them vulnerable to diseases and pests.

What is the function of oestrogen?

The hormone oestrogen plays a key role in regulating transcription by influencing molecules called transcription factors.

What are specific transcription factors and how do they regulate the transcription of target genes in eukaryotic cells, and what role does their movement from the cytoplasm to the nucleus play in this process?

-Specific transcription factors are proteins that regulate gene expression by binding to DNA and controlling the transcription of target genes (conversion of DNA to mRNA).

-These factors are often inactive or sequestered in the cytoplasm. In response to a signal (e.g., hormones, growth factors), they move into the nucleus, where they bind to specific DNA sequences near their target genes.

-This binding can either stimulate transcription by enhancing RNA polymerase activity or inhibit transcription by blocking gene expression.

Explain the role of the steroid hormone, oestrogen, in initiating transcription.

1.) Diffusion of Oestrogen:

-Oestrogen is lipid-soluble, allowing it to freely diffuse through the phospholipid bilayer of the cell membrane.

2.) Binding to Receptor:

-Once inside the cell, oestrogen binds to a specific receptor molecule on a transcription factor in the cytoplasm.

3.) Activation of Transcription Factor:

-The binding of oestrogen causes a conformational change in the transcription factor, particularly in its DNA-binding site.

-This change enables the transcription factor to bind to DNA.

4.) Entry into the Nucleus:

-The activated transcription factor moves into the nucleus via nuclear pores.

5.) Stimulation of Transcription:

-Once inside the nucleus, the transcription factor binds to a specific region of DNA at the promoter site.

-This promotes the attachment of RNA polymerase, stimulating the transcription of the gene.

-Oestrogen controls transcription by activating transcription factors.

Importance of Oestrogen Regulation

-Tissue-specific transcription: Allows certain genes to be expressed in specific tissues, such as those in the reproductive system.

-Response to hormonal signals: Enables the body to adapt to changes in hormone levels.

What are Epigenetics?

-Epigenetics refers to heritable changes in gene function that occur without altering the DNA base sequence.

How are they influenced?

-These changes are influenced by environmental factors such as diet, stress, and exposure to toxins, which affect how genes are expressed.

What is affected?

-DNA sequence does not change, only the expression of genes is affected.

DNA Methylation: What is it? Where does it occur? What is the effect of it?

-Methyl groups (-CH3) are added to cytosine bases in DNA.

-This process usually occurs at CpG sites (cytosine followed by guanine).

-Methylation makes DNA more tightly packed, preventing transcription factors from binding.

-Effect: Genes become silenced or switched off.

What are histones?

-Histones are proteins around which DNA is wrapped.

What is acetylation and its effect

-Acetylation: Adding acetyl groups (-COCH3) to histones makes DNA less tightly packed, allowing transcription.

What does increased acetylation cause

Increased acetylation activates genes (switches them on).

What is Deacylation and its effect

Deacetylation: Removing acetyl groups makes DNA tightly packed, preventing transcription.

What does decreased acetylation cause

Decreased acetylation silences genes (switches them off).

Non-Coding RNA (ncRNA)

Some non-coding RNA molecules can interfere with gene transcription by binding to DNA or histones, influencing gene expression.

What are some examples of environmental influences that influence the mechanisms of epigenetic control

-Diet: Nutrients such as folate can influence DNA methylation patterns.

-Stress: Prolonged stress can lead to changes in histone acetylation.

-Toxins: Chemicals like cigarette smoke can affect epigenetic markers, potentially increasing disease risk.

How is epigenetics theorised to be inherited

-Epigenetic changes can sometimes be passed on to offspring.

(Example: Studies have shown that malnourished mothers may pass on changes in gene expression to their children, influencing metabolism.)

-However, during gamete formation, most epigenetic markers are removed through a process called epigenetic reprogramming, though some changes may escape this and be inherited.

How do diseases influence epigenetics e.g. cancer and Autoimmune disorders

-Cancer: Abnormal DNA methylation can switch off tumour suppressor genes, contributing to cancer development.

-Autoimmune Disorders: Misregulation of epigenetic markers can lead to overactive immune responses.

What are cancers and what do they result in?

Cancer occurs when there is uncontrolled cell division, resulting in the formation of a tumour.

What are the two type of tumours

Benign and malignant

What are the key characteristics of benign tumours

• Grow slowly and are usually non-invasive.

• Can cause mechanical damage by pressing against tissues or blood vessels.

• Do not spread to other parts of the body.

What are the key characteristics of malignant tumours

• Grow rapidly and are invasive, damaging neighbouring tissues.

• Can spread to other parts of the body via metastasis (through the bloodstream or lymphatic system).

• More difficult to treat than benign tumours.

Key Differences Between Benign and Malignant Tumours

How to prevent cancer

• Avoid exposure to known carcinogens (e.g., tobacco smoke, UV radiation).

• Maintain a healthy lifestyle, including a balanced diet and regular exercise.

• Regular screenings to detect tumours early.

How to treat cancer

• Surgery: Removal of the tumour.

• Chemotherapy: Targets rapidly dividing cells.

• Radiotherapy: Uses radiation to kill cancerous cells.

• Targeted Therapies: Inhibit specific molecules involved in tumour growth.

What are the 3 Genes Involved in Cancer Development

Oncogenes, tumor suppressor genes, Proto-oncogenes.

Proto-oncogenes

1. Stimulate normal cell division by producing proteins that help cells progress through the cell cycle.

2. When mutated, they become oncogenes, causing uncontrolled cell division.

Oncogenes

1. Formed from mutated proto-oncogenes.

2. They permanently:

   • Activate cell surface receptors.

   • Code for growth factors that overstimulate cell division.

Tumour Suppressor Genes

1. Normally regulate cell division by slowing it or causing apoptosis (programmed cell death) when damage is detected.

2. If these genes are mutated or silenced, the cell cycle becomes unregulated, leading to tumour formation.

What are the two roles of epigenetics in cancer

Abnormal Methylation

Oestrogen and Breast Cancer

Abnormal Methylation

1. Hyper-methylation (increased methylation) of tumour suppressor genes, such as BRCA1, can silence these genes, leading to cancer.

2. Hypo-methylation (decreased methylation) of oncogenes can activate them, causing uncontrolled cell division.

Oestrogen and Breast Cancer

• Increased oestrogen levels in post-menopausal women (due to accumulation in adipose tissue) can contribute to breast cancer.

• Oestrogen binds to transcription factors, activating genes that promote cell division and leading to tumour formation.

RNA interference (RNAi)

RNA interference (RNAi) is a natural process in cells that regulates gene expression by degrading mRNA, preventing it from being translated into a protein. (Translation of the mRNA produced from target genes can be inhibited by RNA interference)

What two RNA are used for this process?

Small interfering RNA (siRNA) and MicroRNA (miRNA).

How does siRNA Works

1. Formation of siRNA:

   • Double-stranded RNA (dsRNA) is cleaved into small interfering RNA (siRNA) by an enzyme called Dicer.

2. Binding to the RISC Complex:

   • One strand of siRNA (the guide strand) binds to a protein complex called RISC (RNA-induced silencing complex).

   • The complementary strand is discarded.

3. Targeting mRNA:

   • The siRNA within the RISC complex binds to complementary sequences on mRNA molecules.

   • This binding forms double-stranded mRNA, which is recognised as abnormal by the cell.

4. Degradation of mRNA:

   • Enzymes within the RISC complex degrade the double-stranded mRNA.

   • This prevents translation, effectively silencing the gene.

How does miRNA Works

1. Formation of miRNA:

   • MicroRNA (miRNA) is transcribed as a longer RNA molecule and then processed into shorter strands by enzymes like Dicer.

2. miRNA in RISC:

   • miRNA binds to the RISC complex similarly to siRNA.

   • However, miRNA is not fully complementary to its target mRNA, so it often inhibits translation rather than degrading the mRNA.

Key Differences Between siRNA and miRNA

Proteome

The proteome is the complete set of proteins produced by a cell, tissue, or organism at a given time.

Genome

The genome is the complete set of DNA in an organism, including all genes and non-coding sequences. It contains the instructions for making proteins and regulating cellular functions.

Compare genome of prokaryotes to eukaryotes

• In prokaryotes, the genome is mostly coding DNA, meaning most of it directly corresponds to proteins.

• In eukaryotes, the genome includes a large amount of non-coding DNA, which plays roles in gene regulation, alternative splicing, and other complex processes.

What determines the proteome

The genome determines the proteome

Which is more complex and why?

Due to factors like alternative splicing and post-translational modifications, the proteome is often more complex than the genome itself—especially in eukaryotes.

What Are Genome Projects?

Genome sequencing projects determine the entire DNA sequence of an organism. This includes reading the genetic code for both coding and non-coding regions.

Genome project in simple organisms

• In simple organisms, determining the genome allows scientists to predict the proteins (proteome) the organism can produce.

In complex organisms

• In complex organisms, such as humans, the presence of non-coding DNA and regulatory genes makes it harder to determine the full proteome.

How is Genome Projects used in medical applications

• Identification of potential antigens for vaccine development.

• Detection of mutations linked to diseases (e.g., Huntington’s disease).

• Creation of personalised medicine tailored to an individual’s genome.

How is Genome Projects used in Evolutionary applications

• Comparison of genomes across species to determine evolutionary relationships

How is Genome Projects used in Evolutionary applications

• Prediction of amino acid sequences in proteins, enabling the design of synthetic molecules or biological systems.

How is Genome Projects used in Genetic Screening

• Screening for mutated sequences, genetic carriers, and disorders such as cystic fibrosis or sickle cell anaemia.

• Pre-implantation genetic diagnosis (PGD) in embryos during IVF treatment.

What is the Human Genome Project (HGP)

An international scientific endeavour to sequence the entire human genome.

What was their achievments

• Successfully determined the sequence of human DNA.

• Provided insight into genetic diseases and potential therapies.

What was the uses of the HGP

• Early detection of genetic diseases before symptoms appear.

• Development of targeted treatments based on a person’s genetic makeup.

What was the Ethical Issues of the HGP

• Risk of genetic discrimination in employment or insurance.

• Concerns over ownership and misuse of genetic information.

• Questions about whether genetic testing could lead to eugenics.

Explain why determining the proteome is more complex in eukaryotic organisms than in prokaryotes due to non-coding DNA, alternative splicing, and post-translational modifications.

1. Non-Coding DNA:

   • Much of the human genome consists of introns, regulatory regions, or repetitive sequences that do not code for proteins.

2. Alternative Splicing:

   • A single gene can produce multiple proteins due to the process of alternative splicing, where exons are rearranged during mRNA processing.

3. Post-Translational Modifications:

   • Proteins can undergo changes after synthesis (e.g., phosphorylation, glycosylation), which cannot be predicted from the DNA sequence alone.

Explain how you could use Genomic Information to Determine Protein Sequences

1. Reading the Genome:

   • By sequencing the DNA of an organism, scientists can identify the genes that code for proteins.

   • The sequence of bases in a gene is transcribed into mRNA and then translated into a sequence of amino acids.

2. Determining the Proteome:

   • In simpler organisms (e.g., bacteria), identifying the proteome (all the proteins produced) is straightforward as most of the genome is coding DNA.

   • In complex organisms like humans, non-coding DNA and regulatory sequences complicate the process.

3. Predicting Amino Acid Sequences:

   • Advances in bioinformatics enable the identification of coding sequences (exons) in the genome.

   • Once the coding sequence is known, the genetic code is used to predict the sequence of amino acids in the protein.

4. Applications:

   • Identifying proteins involved in disease pathways for the development of targeted therapies.

   • Understanding protein function to improve agriculture, biotechnology, and medicine.

What is the challenges to determine protein sequences in complex organisms

1. Non-Coding DNA:

   • Much of the human genome consists of introns, regulatory regions, or repetitive sequences that do not code for proteins.

2. Alternative Splicing:

   • A single gene can produce multiple proteins due to the process of alternative splicing, where exons are rearranged during mRNA processing.

3. Post-Translational Modifications:

   • Proteins can undergo changes after synthesis (e.g., phosphorylation, glycosylation), which cannot be predicted from the DNA sequence alone.

What is Non-Coding DNA?

• Non-coding DNA refers to regions of DNA that do not code for proteins.

What is the 3 functions of Non-Coding DNA?

• Regulatory Roles:

  • Non-coding DNA contains promoters, enhancers, and silencers that control when and how genes are transcribed.

  • These regions influence which genes are switched on or off in different cell types or under different conditions.

• RNA Molecules:

  • Some non-coding regions are transcribed into functional RNA molecules, such as tRNA, rRNA, and microRNA (miRNA), which regulate gene expression or assist in protein synthesis.

• Structural DNA:

  • Non-coding regions contribute to the structure of chromosomes, such as telomeres (protective caps at the ends of chromosomes) and centromeres (important during cell division).

What does regulatory genes code for? Why are they important?

• Regulatory genes code for proteins such as transcription factors, which control the activity of other genes by binding to DNA.

• These genes determine whether a specific gene is transcribed into mRNA, affecting protein production.

What is the Challenges in Translating Genomes into Proteomes

1. Selective Gene Expression:

   • Not all genes are active in every cell at all times.

   • Genes are selectively expressed depending on the cell type, developmental stage, or environmental conditions.

2. Complexity in Eukaryotes:

   • In higher organisms, a significant proportion of the genome is non-coding DNA, making it difficult to predict the proteome (all proteins a genome can code for).

3. Alternative Splicing:

   • A single gene can produce multiple proteins through alternative splicing, where different combinations of exons are joined.

What is the aim of sequencing projects

• Sequencing projects aim to determine the complete sequence of nucleotides in an organism’s genome.

Genome sequencing in simpler organisms

• In simpler organisms (e.g., bacteria), genome sequencing helps determine the sequences of proteins derived from their genetic code.

• This is useful for identifying potential antigens for use in vaccine production.

Genome sequencing in complex organisms

• In more complex organisms, the presence of non-coding DNA and regulatory genes means that understanding the genome cannot directly predict the proteome.

• The proteome refers to all the proteins a genome can code for. However, not all proteins are produced in every cell due to selective gene expression.