Lecture Transcriptional Control in Prokaryotes and Eukaryotes
Regulating Gene Expression Proteins are not required by all cells at all times Eukaryotes – 4 ways...
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Regulating Gene Expression• Proteins are not
required by all cells at all times
• Eukaryotes – 4 ways– Transcriptional (as
mRNA is being synthesized)
– Post-transcriptional (as mRNA is being processed)
– Translational (as proteins are made)
– Post-translational (after protein has been made)
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Transcriptional Regulation• Activating gene transcription • DNA Acetylation • DNA wrapped around histones keep gene promoters inactive• Activator molecule is used (2 ways)
1. Signals a protein remodelling complex which loosen the histones exposing promoter
2. Signals an enzyme that adds an acetyl group to histones exposing promoter region (acetylation)
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Transcriptional Regulation• Inhibiting gene transcription• DNA methylation (Silencing)– Methyl groups are added to the cytosine bases in the
promoter of a gene (transcription initiation complex) – Inhibits transcription – silencing– Genes are placed “on hold” until they are needed
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Post Transcriptional Regulation• RBP (RNA binding proteins) • Used in:
– Pre-mRNA processing– Alternative splicing– Polyadenylation (to 3’ end)
• Rate of mRNA degradation– Masking proteins used to degrade mRNA – Translation does not occur
• Hormones – Casein – milk protein in mammary gland– When casein is needed, prolactin is produced
extending lifespan of casein mRNA– Translation continues to occur
• MicroRNA– Produced by DICER protein – Block protein production – Studied for being early cancer detection
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Translational Regulation• Occurs during protein
synthesis by a ribosome• Polyadenylation
– Changes in length of poly(A) tail
– Enzymes add or delete adenines
– Increases or decreases time required to translate mRNA into protein
• Deadenylase– Removal of poly (A) tail
(polyadenylation)• Exonuclease
– Degrade mRNA after removal of poly (A) tail
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Post-Translational Regulation• Proteolysis
– Removes sections of protein to make it active or inactive
• Inactivating– Removal of N-methionine
• Chemical modification– Chemical groups are added
or deleted – Puts the protein “on hold”– Phosphorylation
• Ubiquitination– Proteins tagged with
ubiquitin are degraded via proteasome
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Cancer• Lack regulatory mechanisms • Mutations in genetic code
(mutagens)– Probability increases over
lifetime– Radiation, smoking, chemicals
• Mutations are passed on to daughter cells – Can lead to a mass of
undifferentiated cells (tumor)– Benign and malignant
• Oncogenes– Mutated genes that once served
to stimulate cell growth– Cause undifferentiated cell
division
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Genetic Mutations• Positive and negative – Natural selection/
evolution – Cancer –death
• Small-Scale – Single base pair
• Large-Scale – Multiple base
pairs/whole genes
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Small-Scale Mutations• Four groups – Missense, nonsense, silent, frameshift• Lactose, sickle cell anemia
– SNPs – single nucleotide polymorphisms• Caused by point mutations
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Missense mutation• Change of a single base pair or group of base pairs• Results in the code for a different amino acid • Protein will have different sequence and structure
and may be non-functional or function differently
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Nonsense mutation• Change in single base pair or group of base pairs • Results in premature stop codon • Protein will not be able to function
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Silent Mutation• Change in one or more base pairs• Does not affect functioning of a gene• Mutated DNA sequence codes for same amino acid • Protein is not altered
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Frameshift mutation• One or more nucleotides are inserted/deleted from a DNA
sequence• Reading frame of codons shifts resulting in multiple missense
and/or nonsense effects• Any deletion or insertion of base pairs in multiples of 3 does
not cause frameshift
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Large-scale mutations • Multiple nucleotides,
entire genes, whole regions of chromosomes
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Large-scale mutations • Amplification – gene
duplication– Entire genes are copied to
multiple regions of chromosomes
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Large-scale mutations • Large-scale deletions – Entire coding regions of DNA are removed • Muscular Dystrophy
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Large-scale mutations • Chromosomal translocation– Entire genes or groups of genes are moved from one
chromosome to another – Enhance, disrupt expression of gene
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Large-scale mutations • Inversion– Portion of a DNA molecule reverses its direction in the
genome– No direct result but reversal could occur in the middle of
a coding sequence compromising the gene
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Causes of genetic mutations• Spontaneous mutations– Inaccurate DNA replication
• Induced mutations – Caused by environmental agent – mutagen – Directly alter DNA – entering cell nucleus – Chemicals, radiation
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Chemical Mutagens • Nitrous Acid
– Modify individual nucleotides– Nucleotides resemble other
base pairs– Confuses replication machinery
– inaccurate copying • Ethidium bromide
– Used to dye DNA/RNA– insert itself into DNA
• Aflatoxin produced by aspergillus (fungi) – found in peanut butter, corn– Low levels approved by FDA – Causes mutation of p53 gene
(acts as tumor suppressor)– Cancer causing?
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Radiation - Low energy • UV B rays • Non-homologous end joining– Bonds form between adjacent nucleotides along DNA
strand – Form kinks in backbone – Skin cancer
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Radiation – high energy • Ionizing radiation – x-ray, gamma rays • Strip molecules of electrons • Break bonds within DNA– Delete portions of chromosomes
• Development of tumors
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Large-scale mutations • Trinucleotide repeat
expansion• Increases number of
repeats in genetic code • CAG CAG CAG CAG CAG
CAG CAG CAG • Occurs during DNA
repair/replication• “loop out” structures may
form due to repetitive nature of DNA
• Increase in expansion could cause disease or increase severity of disease– Neuromuscular/
neurodegenerative disorders
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Transposable Elements (TE)• Roughly half the Human
Genome is made up TE’s• Result in mutation • DNA Transposons– Jumping genes “cut and
paste” mechanism – Move from one location of
the genome to another– Encode protein transposase
which is required for insertion and excision
– Terminal inverted repeats (9-40) base pairs long
– Less than 2% of human genome
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Transposable Elements (TE)• Retrotransposons (RNA transposons)
• move through action of RNA intermediates • Produce RNA transcripts • Reverse transcriptase enzymes reverse RNA back to DNA and inserted • Give rise to variation in organism• Evolution of species 1. LINE – long interspersed transposable elements
• 6 kilobases long 2. SINE – short interspersed transposable elements (not in humans)
• Few hundred bases long
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Genomes and Gene organization• Human Body– 22 autosomal chromosomes– 1 pair of each sex chromosome (XX, YY)