Gene Expression and Genetic Regulation

Gene Expression

Lecture 21, Part 1
Assignments, Quizzes, and Exams

  • Smart Book Homework – Chapters 20 and 21

  • Quizzes – Quiz 13 & 14

  • Exams:

    • Exam #4 – Thursday, May 1, 222 Johnson Hall, online with Respondus Lockdown Browser

    • Exam #4 Review Session – Tuesday, April 29, 7:00-9:00 PM, Nutrien 135

    • Final Exam – Monday, May 12, 9:40-11:40 AM, 222 Johnson Hall, Online with Respondus Lockdown Browser

    • Final Exam Review Session – Sunday, May 11, 3:00-5:00 PM, Nutrien 135

    • Group Learning Sessions – Monday-Thursday, 5:00-9:00 PM, TILT Great Hall

Concept Check

  • Control of gene expression in eukaryotes (where, how).

  • Components of an operon (POG = promoter, operator, genes).

  • Function of a repressor.

  • Difference between inducible and repressible operons (Lac, Trp).

  • How Lac and Trp operons work (substrate, on/off, product).

Eukaryotes vs. Prokaryotes

  • Prokaryotes are single-celled (bacteria).

  • Eukaryotes can be single-celled (yeast).

  • Most eukaryotes are multi-cellular, leading to more complex transcription control.

Gene Expression

  • Necessary proteins must be synthesized at the proper time for a cell to function correctly.

  • Gene expression: turning on a gene to produce RNA and protein.

  • Mechanism needed to control when a gene is expressed, how much protein is made, and when to stop production.

Gene Control

  • Multicellular organisms develop through:

    • Increase in the number of cells (growth).

    • Cells becoming specialized (differentiation).

  • Both growth and differentiation are regulated.

  • Each cell has some genes switched on and some switched off to perform its unique role.

Mechanisms of Gene Control

  • Genes regulation

  • Epigenetics.

  • Heterochromatin

  • Transcriptional control.

  • Inducers and Repressors.

  • Post-Transcriptional modifications

  • mRNA regulation

  • Translational control.

  • Protein modification.

  • Transcriptional control is the most common type of regulation.

Regulation in Eukaryotes

  • Regulation of gene expression can occur at all stages.

    • DNA uncoiling and loosening from nucleosomes to bind transcription factors (epigenetic level). This involves modifications to histone proteins and DNA, influencing chromatin structure and accessibility for transcriptional machinery.

    • RNA transcription (transcriptional level). This includes the binding of transcription factors to promoter and enhancer regions to initiate or repress transcription.

    • RNA processing and export to the cytoplasm (post-transcriptional level). Modifications such as splicing, capping, and polyadenylation occur to produce mature mRNA. Nuclear export mechanisms ensure the mRNA reaches the cytoplasm for translation.

    • RNA translation into protein (translational level). Ribosomes bind to mRNA and, with the help of tRNA, synthesize proteins. Regulatory proteins and RNA structures can influence the efficiency of translation.

    • After protein synthesis (post-translational level). Proteins undergo folding, modification, and assembly. They can be activated or inactivated through phosphorylation, glycosylation, ubiquitination, and other modifications.

Enhancers

  • Enhancers: Regions in eukaryotic genes that increase or enhance transcription. These can act over long distances and often involve DNA looping to bring distally bound transcription factors into proximity with the promoter.

  • Enhancers are not necessarily close to the genes they enhance.

  • Location: Upstream, within the coding region, downstream, or thousands of nucleotides away.

  • Enhancers are DNA sequences; transcription factors are proteins that bind to DNA sequences. These transcription factors can be activators or co-activators that stabilize the transcriptional complex.

Control Types

  • Negative: Stops transcription.

  • Positive: Promotes transcription.

  • Inductive: Induces (causes) transcription. Often involves a signal molecule that inactivates a repressor.

  • Repressive: Inhibits (prevents) transcription. Often involves a signal molecule that activates a repressor.

Bacterial Genes and Operons

  • Operon: Cluster of functionally-related genes controlled by a shared operator. This allows for coordinated expression of genes involved in the same metabolic pathway.

  • Promoter: Binding site on DNA where RNA polymerase attaches. The promoter contains specific sequences like the TATA box that help position RNA polymerase for transcription initiation.

  • Operator: Binding site on DNA where the repressor attaches. The operator is typically located downstream of the promoter.

  • Repressor: Protein that binds to the operator to decrease transcription. The repressor can block RNA polymerase from moving forward or prevent the binding of necessary transcription factors.

Prokaryotes

  • Gene expression in prokaryotes ensures that a cell’s resources are not wasted making unneeded proteins. This is crucial for adapting to changing environmental conditions.

  • In bacteria, structural proteins with related functions are encoded together in an operon and transcribed together. This arrangement allows for efficient and coordinated gene expression.

  • POG:

    • Promoter - DNA sequence

    • Operator - DNA sequence

    • Gene - DNA sequence

Regulation - Inducible vs. Repressible

  • Inducible: Metabolite present = Operon (Genes) On. The presence of the metabolite signals the need for the gene products.

  • Repressible: Metabolite present = Operon (Genes) Off. The presence of the metabolite indicates that the gene products are not needed.

  • Positive: Operon ON if protein is present. An activator protein is required for transcription.

  • Negative: Operon OFF if protein is present. A repressor protein blocks transcription.

Operons - Prokaryotic

  • Operon - group of related genes

  • Polycistronic RNA - polygene. A single mRNA molecule that contains coding sequences for multiple genes.

  • Bacterial control of gene expression

  • Operon: cluster of related genes with on/off switch

  • Three Parts:

    1. Promoter - where RNA polymerase attaches

    2. Operator "on/off", controls access of RNA poly

    3. Genes code for related enzymes in a pathway

Repressors

  • Lactose present = need enzymes (proteins) to digest = binds to repressor and removes = transcription occurs. The lac repressor undergoes a conformational change when lactose is present, reducing its affinity for the operator.

  • Tryptophan present = do not need proteins (to synthesize this) = binds to repressor and keeps in place = transcription does not occur. The trp repressor binds to tryptophan, enhancing its affinity for the operator.

Genome Size Comparison

  • Comparative genome sizes of organisms:

    • Homo sapiens (human): 3.2 billion3.2 \text{ billion}, ilda 25,000 genes, 1 gene /100,000 bases, 46 chromosomes

    • Mus musculus (mouse): 2.6 billion2.6 \text{ billion}, ilda 25,000 genes, 1 gene /100,000 bases, 40 chromosomes

    • Drosophila melanogaster (fruit fly): 137 million137 \text{ million}, 13,000 genes, 1 gene/9,000 bases, 8 chromosomes

    • Arabidopsis thaliana (plant): 100 million100 \text{ million}, 25,000 genes, 1 gene / 4000 bases, 10 chromosomes

    • Caenorhabditis elegans (roundworm): 97 million97 \text{ million}, 19,000 genes, 1 gene / 5000 bases, 12 chromosomes

    • Saccharomyces cerevisiae (yeast): 12.1 million12.1 \text{ million}, 6000 genes, 1 gene / 2000 bases, 32 chromosomes

    • Escherichia coli (bacteria): 4.6 million4.6 \text{ million}, 3200 genes, 1 gene/1400 bases, 1 chromosome

    • H. influenzae (bacteria): 1.8 million1.8 \text{ million}, 1700 genes, 1 gene /1000 bases, 1 chromosome

Genome - Prokaryote vs. Eukaryote

  • Eukaryotes:

    • Much greater size of genome

    • DNA packaged in chromatin fibers. This packaging involves histone proteins and DNA methylation, affecting gene accessibility.

      • Regulates access to DNA by RNA polymerase

    • Cell specialization. Different cell types express different sets of genes, allowing for specialized functions.

    • Need to turn on & off large numbers of genes. This regulation is essential for development and homeostasis.

    • Most of DNA does not code for protein- 97% "junk DNA" in humans. This includes introns, regulatory sequences, and repetitive DNA elements.

  • Prokaryotes:

    • Small size of genome

    • Circular molecule of naked DNA- DNA is readily available to RNA polymerase

    • Control of transcription by regulatory proteins- Operon system

    • Most of DNA codes for protein or RNA- No introns, small amount of non-coding DNA

      • Regulatory sequences: promoters, operators

lac Operon

  • lac operon = inducible

  • When lactose (a sugar) is present, the repressor is released so that transcription of genes involved in digestion of the lactose can occur. Allolactose, a derivative of lactose, binds to the repressor, causing it to detach from the operator.

  • So, presence of the substrate (lactose) removes the repressor and induces transcription.

trp Operon

  • trp operon = repressible

  • When tryptophan (an amino acid) is present the repressor stays bound to the DNA to prevent transcription of genes that are involved in making tryptophan. Tryptophan acts as a corepressor, binding to the repressor protein, which then binds to the operator.

  • So, the presence of substrate (tryptophan) causes the repressor to stay bound to the DNA (operator) to repress (inhibit) transcription.

lac/trp Comparison

  • lac: substrate induces. lactose

  • trp: product represses. tryptophan

Summary - Operon

  • An operon is a functioning unit of genomic DNA that contains a group of genes controlled by a single promoter.

  • These genes share information needed to create the tools for a particular task, so they share a promoter ensuring they’ll all be transcribed together.

  • The lac, or lactose, operon contains genes coding for proteins in charge of transporting lactose into the cytosol and digesting it. Enzymes like beta-galactosidase, permease, and transacetylase are crucial for lactose metabolism.

  • The trp, or tryptophan, operon contains the genes coding for proteins in charge of making tryptophan. These enzymes catalyze the synthesis of tryptophan from chorismate.

Genetic Regulation

Lecture 21, Part 2
Concept Check

  • Review bacterial operons (components and function).

  • Describe the function of DNA methylation.

  • Explain how genes are regulated (turned on and off) during development.

  • Describe the different types of potency and what this means.

  • Explain why a zygote is considered totipotent while a blastocyst is not.

  • Explain the function of stem cells.

Review

  • Operons: Clusters of bacterial genes

    • P = promoter (where RNA polymerase binds)- Before (upstream of) the genes

    • O = operator (where the repressor binds)- After the promoter, so that it can stop RNA polymerase from transcribing

    • G = genes- Group of genes involved in a specific pathway

    • Remember that bacterial mRNA can be polycistronic

mRNA Processing

  • In eukaryotic cells, pre-mRNAs undergo three main processing steps:

    • Capping at the 5' end. A modified guanine nucleotide is added to protect the mRNA from degradation and enhance translation.

    • Addition of a poly(A) tail at the 3' end. A string of adenine nucleotides is added to increase mRNA stability and promote export from the nucleus.

    • Splicing to remove introns. Non-coding regions (introns) are removed, and coding regions (exons) are joined together.

Methylation

  • DNA methylation

    • Methyl Group - Functional Group - CH3

    • Genes Off- Methylation of DNA blocks transcription factors- no transcription- genes turned off

      • attachment of methyl groups (-CH3) to cytosine- C = cytosine. This modification is typically found at CpG dinucleotides.

      • nearly permanent inactivation of genes- ex. inactivated mammalian X chromosome = Barr body. This process is called X-inactivation and ensures that females, with two X chromosomes, do not have twice as many X-linked gene products as males.

Development

  • Gene Expression during Development

    • During zygote development:- genes expressed only when specific proteins are needed

      • cell differentiation:- development of different cells with specialized functions

        • forms tissues and organs

      • morphogenesis: development of form in an organism. This involves cell migration, shape changes, and programmed cell death (apoptosis).

Expression

  • Genes -20% expressed/cell

  • Eukaryotic Gene Expression

    • Contain many different types of cells

    • On average, a typical human cell expresses only 20% of its genes w/highly differentiated cells expressing less! This allows for cell-specific functions and responses to stimuli.

    • Although they all contain the same genome, their specific gene expression allows these cells to be unique in function.

Environment

  • Environmental Influences

    • Phenotype is not determined only by genes. The environment- the conditions surrounding an organism - also affect phenotype.

    • For example, the sex of sea turtles depends on the temperature of the environment in which the egg develops.

      • Eggs in warmer nests result in females, and eggs in cooler nests result in males. This is known as temperature-dependent sex determination (TSD).

    • Likewise, genes and environment interact with human traits.

      • Genes influence height, but do not completely control it.

      • Take two identical twins and raise them in separate environments, one that has good nutrition and health care and the other that does not. Their height and size can vary greatly. Epigenetic modifications can also be influenced by environmental factors, leading to differences in gene expression.

Cells

  • The process of cellular differentiation is regulated by transcription factors and growth factors, and results in expression or inhibition of various genes between the cell types, thereby resulting in varying proteins expressed in different cell types. Signaling pathways like the Wnt, TGF-beta, and Notch pathways also play crucial roles.

  • Somatic cells are diploid cells that make up most of the human body, such as the skin and muscle. They are produced through mitosis.

  • Germ cells are any line of cells that give rise to gametes - eggs and sperm - and thus are continuous through the generations. They undergo meiosis.

  • Stem cells have the ability to divide for indefinite periods and to give rise to specialized cells. This self-renewal capacity is critical for tissue maintenance and repair.

Potency

  • Totipotent, Pluripotent & Multipotent- What is the Difference!!!

    • Embryonic Stem Cells- found in embryo

      1. Totipotent - whole- zygote. Can give rise to all embryonic and extraembryonic tissues.

      2. Pluripotent - several- found in inner cell mass

        • inner- cell mass- blastocyst. Can give rise to all cells of the body, but not extraembryonic tissues.

      3. Multipotent - a few in this case- e.g. Mesenchymal- Stem Cell. Can differentiate into a limited number of cell types within a particular lineage.

      • Adult Stem Cells- found in many organs

        • specialization potential is limited to one or more cell lines

          • Bone

          • Cartilage

          • Connective tissue

      • At this point = cells differentiated

Stem Cells

  • Stem cells are undifferentiated biological cells found in multicellular organisms, that can differentiate into specialized cells or can divide to produce more stem cells. This plasticity makes them valuable in regenerative medicine.

  • In mammals, there are two broad types of stem cells: embryonic stem cells, which are isolated from the inner cell mass of blastocysts, and adult stem cells, which are found in various tissues. Each type has distinct properties and applications.

  • In adult organisms, stem cells act as a repair system for the body by replenishing adult tissues. They help maintain tissue homeostasis and repair damage from injury or disease.

  • In a developing embryo, stem cells can differentiate into all of the specialized cells of the body. This process is tightly regulated by signaling pathways and transcription factors.

  • The pathway that is taken to produced specialized cells includes: the embryonic cells develop from totipotent cells, to pluripotent cells which undergo differentiation and become more specialized.

Stem Cells Types

  • Totipotent Stem Cell

    • These cells have unlimited capability, and have the ability to form extraembryonic membranes and tissues, the embryo itself, and all postembryonic tissues and organs.

      • An example is an embryo

  • Pluripotent Stem Cell

    • These cells are capable of giving rise to most, but not all, tissues of an organism.

      • An example is inner mass cells

  • Multipotent Stem Cell

    • These cells are committed to give rise to cells that have a specific function. An example is blood stem cells

Types of Stem Cells:

  • The first embryonic cells that arise from the division of the zygote are the ultimate stem cells; these stems cells are described as totipotent because they have the potential to differentiate into any of the cells needed to enable an organism to grow and develop.

  • The embryonic cells that develop from totipotent stem cells and are precursors to the fundamental tissue layers of the embryo are classified as pluripotent.

  • These cells then become slightly more specialized and are referred to as multipotent cells.

Transcription Factors

  • The primary mechanism by which genes are turned “on” or “off” is through transcription factors.

  • A transcription factor is a protein that binds to specific genes on the DNA molecule and promotes or inhibits their transcription. They can recruit co-activators or co-repressors to the DNA, influencing gene