Chapter 18- Regulation of gene expression You Must Know Functions of 3 parts of an operon Role of repressor genes on the operon Impact of DNA methylation and histone acetylation on gene expression Role of oncogenes, proto-oncogenes, and tumor suppressor genes

Bacteria (prokaryotes) Characteristics Peptidoglycan Membrane, cell wall Binary fission Conjugation Exchange of genetic material and information F plasmid, F+ donates to F- R plasmids- resistance

Bacteria Response to Environmental Change Regulate Transcription Bacteria are favored by Natural Selection Reproduces quickly Produce what it needs Regulates enzymatic production by feedback inhibition or gene regulation Gene expression by the operon model

Environmental Impacts on Bacteria In our “gut”bacteria must respond to what we eat Lacks tryptophan (aa in turkey that makes you sleepy) bacteria actives another pathway-feedback mechanism You pig out on turkey, bacteria will stop manufacturing tryptophan- feedback inhibition

Bacteria in Colon Respond to what the host eats Sounds like + and – feedback Low tryptophan, must manufacture High Tryptophan Stop

Bacterial Genes Clustered into units called OPERONS 3 parts Operator Controls RNA polymerase access to genes Promoter Where RNA polymerase attaches Genes of the operon DNA required for all enzymes produced buy the operon

Operon On switch for segment of DNA Positioned in promoter or between the promoter and enzyme coding genes Controls access point of RNA polymerase Operator, promoter and genes they control is operon

Anatomy of an Operon Operator Controls access of RNA polymerase Found within promoter or between promoter and protein coding genes of the operon

Operon Promoter RNA polymerase attaches

Genes of the Operon Entire stretch of DNA Codes for all enzymes produced by the operon

Regulatory Proteins Located some distance from the operon Produce repressor proteins to bind to the operator site RNA polymerase Blocked from the genes Operon is OFF

Repressor Is the protein OFF switch Prevents gene transcription Binds to the operator Blocks RNA polymerase Part of a separate regulatory gene Can be active or not Corepressor molecule works with repressor protein to switch off the operon

Repressible Anabolic Synthesize essential end products from raw materials Suspend production of end product when it’s present conserves resources On all the time, can be turned off

Repressible Operon

Bozeman Lac Operon

Inducible Operon(enzymes) Lactose Normally off but can be activated Catabolic Repressor protein is active Inducer binds to and inactivates the repressor protein RNA poly can access

Inducible Operon

2 Types of Operons Tryp Operon-Normally on Lac Operon Normally off Repressible Inducible Tryp Operon-Normally on Can be inhibited Anabolic for building organic molecules repressor protein is inactive. Produced molecule can act as a corepressor and binds to the repressor protein and actives it Binds to the operator site and shuts down the operator Lac Operon Normally off Can be activated Catabolic-breaking down macromolecules Repressor protein is active small molecule called inducer ginds to and inactivates the repressor Repressor out of the operator site, RNA polymerase can access the genes of the operon

How do you regulate operons? Feedback mechanisms inhibition Anabolic (biosynthetic) Shuts down synthesis of tryptophan Regulate the expression of the genes encoding the protein Control enzyme production during transcription mRNA encoding OPERON!!!

Positive Gene Regulation When glucose (preferred source of food for E. coli) is scarce, CAP is activated by cyclic AMP-changes shape and binds to it Activated CAP Attaches to the promoter of the lac operon Increases the affinity of RNA polymerase Accelerates transcription CAP detaches when glucose levels increase Transcription back to normal Positive Gene Regulation Needs a stimulatory protein such as catabolite activator protein (CAP) ACTIVATOR OF TRANSCRIPTION

What does this all mean? Survival!!! If glucose is not present, lactose is broken down Glucose is plentiful, CAP is inactive, enzyme activity slows down for other sugars

Eukaryotic Gene Expression Regulated at many stages

What is the fundamental packaging of DNA? Chromatin Structure Genes within highly packed heterochromatin are usually not expressed Nucleosomes DNA bound to histone Tighter the winding. Less access for transcription Chemical modifications and DNA Chromatin Methylation, acetylation

Eukaryotic Regulation of Gene Expression Regulation of Chromatin Structure Histone acetylation, DNA Methylation, Epigenetic inheritance Regulation of Transcriptional Initiation Control elements, general transcription factors, specific transcription factors, Enhancer Combinatorial control, Nuclear architecture and gene expression Post-Translation Regulation RNA splicing, mRNA degradation

Chromatin Modification Nucleus Histone Modification Histone Acetylation Acetyl groups are attached to positively charged lysines in histone tails Loosens chromatin Promotes initiation of transcription

Chromatin Modifications Nucleus DNA Methylation Addition of methyl groups (methylation) Condenses chromatin Reduces transcription Addition of phosphate (phosphorylation) next to methylated amino acid loosens chromatin

Histone Code Hypothesis Proposes that specific combinations of modifications, as well as the order in which they occur determines the chromatin configuration Influences transcription

Chromatin Modifications Epigenetic Inheritance Chromatin modifications do not alter DNA sequence Can be passed on to future generations No change in nucleotide sequence Saw this with the sepia eyes over-riding an X-linked trait

Transcription -Nucleus DNA Control elements in enhancers Bind specific transcription factors Bend the DNA Activators contact proteins at the promoter initiation transcription

RNA Processing In Nucleus Alternate RNA splicing primary transcript is edited, mis-sliced mRNA degradation Sequences in the 5’and 3’ end determines life span

Post-Translational Regulation-cytoplasm Alternative RNA splicing Protein processing and destruction Proteasomes recognize proteins tagged with ubiquitin Mutations making specific cell cycle proteins impervious to proteasomes can lead to cancer

Noncoding RNA in Gene Expression Play multiple roles 1.5% of the human genome codes for protein Regulation and expression by ncRNA (noncoding RNA)

Form and Function of MiRNA

MRNA impact by miRNA and RNAi miRNA, RNAi, siRNA Destroys double stranded foreign RNA (virus) Degrades mRNA Prevents translation piRNA (piwiRNA)-induces formation of heterochromatin and prevents parasitic DNA or transposons

Translation miRNA or siRNA can block the translation of specific mRNA’s Can target specific mRNA’s for destruciton

Who cares? Scientists can use siRNA (small interfering) too Cure disease with inappropriate gene expression

Impacts of differential gene expression on multicellular organisms Cell division Increase cell number Cell differentiation Specialization of cells Morphogenesis Organization into tissues and organs

Differential Gene expression Leads to different cell types in multicellular organism Cell division Number of cells Cell differentiation specialization Morphogenesis Tissues and organs

What Controls differentiation and expression? Cytoplasmic determinants Maternal substances in the egg Cell-cell signals Molecules, growth factors Determination Cell-cell signals and is irreversible Pattern formation Hox

Cytoplasmic Determinants Maternal substances in the egg that influence development Induction Determination Signals that change the target cells causing cellular development by embryo

Determination Series of observable differentiation of a cell Irreversible differentiation by cell-cell signals

Pattern Formation Body plan as set up by cytoplasmic determinants and inductive signals Hox gene Molecular cues that control pattern formation are called positional information

Hox gene

Cancer Genetic changes that affect the cell cycle Gene regulation goes bad

Cancer Uncontrolled mitosis by a mutation in a gene whose products normally inhibit cell division

Oncogenes Genes that cause Cancer Proto-oncogenes code for proteins for normal cell growth but can become oncogenes when a mutation occurs that increases the product of proto How does a proto-oncogene become an oncogene? 1. movement of DNA 2. amplification of proto-onco 3. point-mutation of proto-onco

Tumor suppressor genes Cells secrete products to inhibit cell division BRACA1 and 2 (breast cancer gene) mutations in those genes increase risk Functions of these genes: Repair damaged DNA Control adhesion of cells to eachother or extracellular matrix

Tumor Suppressor p53 gene Named for the 53,000 dalton molecular wieght of its molecular product Halts cell cycle-Guardian Angel of the Genome Binds cyclin-dependent kinases Turns on genes for DNA repair Activates suicide genes-APOPTOSIS

Multistep model of Cancer Development

What happens when there is interference with normal cell-signaling pathways? Ras protein- encoded by ras gene (rat sarcoma) Relay switch in membrane for chemical cascade of kinases that stimulate cell cycle Must be triggered by the RIGHT growth factor, hyperactive ras gene- increases cell division P53 gene Promotes synthesis of cycle-inhibiting proteins Knock out P53- activates RAS protein

BRCA-1 and 2 Tumor suppressor genes Blood test to check for mutation the long (q) arm of chromosome 17 at position 21. BRCA-2- chromosome 13