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Responding to the Environment (Eukaryotic Transcriptional Regulation (Cell…
Responding to the Environment
Modulating Transcription
Prokaryotics & Enrivonment
Growth
Nutrient-rich environment
Containing amino acids and carbohydrates
Favourable temperature
Gene regulation helps with environmental responses
Levels of Regulation
Gene expression
Functional product of gene is made, modified, activated
Transcriptional Control
Transcription of DNA to mRNA
Amount of mRNA in cell
Transcription activation
Proteins bind to region near promoter, increase binding of RNA polymerase
Gene is transcribed
Turned on
Slowest regulation level
Cell starts from scratch
Most efficient regulation level
No energy or resource waste
Translational Control
Translation of mRNA to proteins
Eukaryotes
Binding ribosome to 5' end/5' CAP of mRNA
Prokaryotes
Ribosome binds to Shine-Dalgarno sequence
Quick speed = little protein produces
Post-Translational control
Modifications and activation of produced proteins
Polypeptide chain to be folded into functional 3-dimensional structure
Fastest regulation level
Stockpiles of inactive protein
Prokaryotic Transcriptional Regulation
Environmental Cues & Gene Expression
E.Coli
Metabolize all glucose before switching to lactose
Transcriptional level regulation
Trancriptional responses to environment
Increase in beta-galactosidase & lactose permease proteins
Not detectable when using glucose
Expressed when glucose fully depletes
Gene expression cues
Environmental cues shift digestive proteins being utilized
Lactose permease
Transport of lactose into bacterial cells
Operon model
Groups of functionally related genes are organized into transcriptional units along chromosome
Operator
On-off switch
Inhibit or allow transcription
Lac Operon - Transcriptional regulation
Prokaryotes
Regulates B-galactosidase and lactose permease expression
Regulatory sequences of transcription
Promoter that binds RNA polymerase complex
Operator (lacO)
Binding site for repressor protein
Expressed by lacI coding sequences
Lactose metabolism
lacY gene
Codes for lactose permease transport protein
Import of lactose
lacZ gene
Codes of B-galactoseidase
Cleave disaccharide lactose into glucose and galactose
lacI
Controls gene expression of lacY and lacZ
Codes for repressor protein
Bind to operator and inhibit transcription
Lac Operon
Positive Regulation
Environmental Cues
Absence of glucose
Increase in intracellular cAMP
Lac operon
Promotes production of B-galactosidase and lactose permease
cAMP glucose levels
High levels
Low levels of intracellular cAMP
Low levels
High levels of cAMP
Lactose operon
Protein
CRP/CAP
Bound by cAMP CRP-cAMP or CAP-cAMP binding site
cAMP signals
Bind to CRP protein as allosteric activator
CRP-cAMP complex bind to DNA furthering transcription in presence of lactose
High extracellular glucose concentration
Enzyme inhibition
Low levels of cAMP
CRP-cAMP will not bind to lac operon
Negative Regulation
Lactose operon
Negatively regulated transcription
Repressor protein, binds to operator region of operon, turning off transcription
Transcription genes required for lactose metabolism are turned off
Repressor protein, in lacI expressed in low levels
Binds to lacO
Repressor proteins
Lac operon repressor, expressed by lacI gene
inhibits expression of B-galactosidase and lactose permease proteins
Tetrameric protein
4 subunits bind to lac operon DNA
Twisted into loop
RNA polymerase not able to bind to lac operon promoter
1 more item...
Allosentical Inhibit
Glucose deletes, utilize lactose
Lactose is inducer molecule
Lactose transcription inducer
Lactose binds to repressor protein
Repressor now can't attach to operator sequences in lac operon DNA
Eukaryotic Transcriptional Regulation
Cell Differentiation
Stem Cells
Zygotes have identical stem cells
Same genome, gene regulation differences lead to altered proteomes, having differences in cellular functions
Transcription Machinery
Transcription factors
Proteins that bind to DNA sequenes
Interact with DNA double helix
Control transcription and gene regulation
Determine what cell will specialize as
Controls active genes on chromosome
Chromatin Remodeling
DNA Compaction
Gene is controlled by own promoters and enhancers
Compacting DNA into tightly wound chromatin fibers
Fit all DNA in nucleus, easy to move around
Tightly wound heterochromatin not expressed
Need to unwind DNA to transcribe
Chromatin remodeling
Activator protein (transcription factor) binds to accessible enhancer site
CpG Methylation
CG sites
Cytosine and Guanine follow each other on DNA
DNA methyltransferase
Enzymes that add methyl group
Developmental Changes
Human Globin Gene Expression
Blood cell progenitors (stem cells)
Differentiate into red blood cells with hemoglobin
Adults
2 A-globins, B-globins
Have Y-globins, wound up tight, unexpressed
Unable to transcribe
Fetus
2 A-globins, Y-globins
Y-globin binds oxygen more strongly than B-globin
Have B-globins, wound up tight, unexpressed
Unable to transcribe
Turning Signal Off
Measuring Gene Expression
In Situ
"In it's original place"
Assess gene expression levels of few genes
Spatial differences in expression
Compare throughout developmental stages
Microarray
Examine expression of thousands of genes at once
Base-pair interactions
Binding of nucleic acid complementary strands
DNA molecules ac as probes for gene expression
Transcriptome
Turning Off Expression
RNA Stability
Removing mRNA
Stop gene expression
mRNA should be degraded
Regulate mRNA through stability of polyA tail
miRNA - translation inhibition
Form hairpin loops, from complimentary base-pairing
Becomes part of RNA-induced silencing complex (RISC)
Bind to mRNA
Inhibit translation
siRNA - mRNA degradiation
Associated with RISC
Induce cleavage of mRNA
Protein Stability
Protein degradation
Proteasomes degrade unneeded/damaged proteins
Proteasomes break peptide bonds
Regulate protein concentrations
Proteins are removed from cell by marking them for enzymatic degradation
Selective deragation
Length of time protein functions in cell, can be limited
Ubiquitin-proteasome pathway
Transcription factors
Cell-cycle regulators
Misfolded proteins
Applied Lecture
Lactose Intolerance
Lactase hydrolyzes lactose into monosaccharides
Drop in lactase enzymes after weaning
Gene stays on
Ancestors with domesticated dairy animals
MCM6 gene mutation
Born without lactase gene
Can't detect lactose
Results
Excessive lactose attracts water molecules
Water not properly absorbed into bloodstream
Intestinal bacteria on lactose
Congenital
LCT gene on chromosome 2 codes for lactase enzyme
Galactosemia
Galactose in blood
Born without enzymes needed for galactose processing
GALT, GALE, GALK1 & chromosome 9 genes
Process galactose into glucose
Result
Toxic accumulation of galactose
Organ and tissue damage
Treatment
Omitting galactose from diet
Treatment
Omitting lactose from diet
Probiotics
microorganism helps with digestion
Modifies microbiome of colon
Applied Lecture
Epigenetics & Gene Regulation
Genome is inheriteed
Epigenome can be altered
Chemical DNA modifications in genome, change gene activity, DNA sequence unaltered
Epigenome
Modification of histone tails
DNA methylation
Chromatin remodeling
DNA packaged around nucleosomes
Cancer
Abnormal epigenomes
Cell's DNA sequence mutations
Agouti
Mice have pigment change
Black or yellow
Genetically identical
Normal mice
Brown
Gene kept off from DNA methylation
Obese
Yellow
Genes not methylated
Maternal Diet
Maternal methy source
Brown -> yellow
DNA methylation
Brown -> yellow
Gene expression
Yellow -> brown
Twins
differential methylation
Results
Patterns of epigenetic modifications and gene expression are different with age
Difference in environment can produce long-term epigenetic differences
Epigenetic drift
Studies
Environment can affect phenotype
Changes can occur from embryo to adulthood
Heritable
Reversible
Obesity, heart disease, etc. affect it
External Factors
Lifesyle
Diet
Habits