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Chapter 28 Regulation of Gene Expression - Coggle Diagram
Chapter 28 Regulation of Gene Expression
基本知識
Trends in Understanding Gene Regulation
Past focus has been on understanding transcription initiation.
There is increasing elucidation of posttranscriptional and translational regulation.
Mechanisms can be elaborate and interdependent, especially in development.
Regulation relies on precise protein-DNA and protein-protein contacts.
The Vocabulary of Gene Regulation
Housekeeping gene
under constitutive expression
constantly expressed in approximately all cells
Regulated gene
Levels of the gene product rise and fall with the needs of the organism.
Such genes are inducible.
• able to be turned on
Such genes are also repressible.
• able to be turned off
Ways to Regulate Protein Concentration in a Cell
Synthesis of primary RNA transcript
How to process this RNA into mRNA
Posttranscriptional modifications of mRNA
Degradation of mRNA
Protein synthesis
Posttranslational modification of protein
Targeting and transport of the protein
Degradation of the protein
DNA elements that control transcription
RNA Polymerase Binding to Promoters Is a Major Target of Regulation
RNA polymerases bind to promoter sequences near the starting point of transcription initiation.
The RNA pol-promoter interaction greatly influences the rate of transcription initiation.
Regulatory proteins (transcription factors) work to enhance or inhibit this interaction between RNA pol and the promoter DNA.
A Consensus Sequence Is Found in Many E. Coli Promoters
Most bacterial promoters include the conserved –10 and –35 regions that interact with the σfactor of RNA polymerase.
Some promoters also include the upstream element that interacts with the αsubunit of RNA polymerase.
Mechanisms to Regulate Transcription in Bacteria
Use of σ factors
recognize different classes of promoters
allows coordinated expression of different sets of genes
Binding other proteins (transcription factors) to promoters
recognize promoters of specific genes
may bind small signaling molecules
may undergo posttranslational modifications
protein’s affinity toward DNA is altered by ligand binding or posttranslational modifications
allows expression of specific genes in response to signals in the environment
Regulation by Specificity Factors Such as σSubunits of RNA Pol
-Specificity factors alter RNA polymerase’s affinity for certain promoters.
Heat Shock Induces Transcription of New Products to Protect Cell
Occurs when bacteria are subject to heat stress
RNA Pol replaces σ70 with σ32
Causes RNA Pol to bind to different set of promoters
Small-Molecule Effectors Can
Regulate Activators and Repressors
Repressors reduce RNA Pol-promoter interactions or block the polymerase.
Effectors can bind to repressor and induce a conformational change.
Activators Improve Contacts Between
RNA Polymerase and the Promoter
Binding sites in DNA for activators are called enhancers.
In bacteria, enhancers are usually adjacent to the promoter.
In eukaryotes, enhancers may be very distant from the promoter.
Negative Regulation
Positive Regulation
DNA Looping Allows Eukaryotic Enhancers to Be Far from Promoters
Activators can influence transcription at promoters thousands of bp away.
How? Via formation of DNA loops
Looping can be facilitated by architectural regulator proteins.
Co-activators may mediate binding by binding to both activator and RNA polymerase.
Many Bacterial Genes Are Transcribed And Regulated Together in an Operon
An operon is a cluster of genes sharing a promoter and regulatory sequences.
First example: the lac operon
Protein factors that control transcription
Activation of Bacterial Translation
by Small RNA Molecules
The ribosome-binding Shine−Dalgarno sequence is sequestered into a stem-loop structure in the mRNA.
In the presence of protein Hfq, small regulatory RNA DsrA binds to the mRNA.
The binding of DsrA opens up the stem-loop and allows mRNA binding to the ribosome.
DsrA RNA promotes translation.
Inhibition of Bacterial Translation
by Small RNA Molecules
The ribosome-binding Shine−Dalgarno sequence is sequestered into a stem-loop structure in the mRNA.
In the presence of protein Hfq, small regulatory RNA OxyS binds to the mRNA.
The binding of OxyS blocks the ribosome binding site in mRNA.
OxyS RNA inhibits translation.
Regulation of Bacterial mRNA Function in Trans by sRNA
Cis Regulation by Riboswitches
Trans-Acting sRNAs Facilitate
Translation of σS mRNA
σS mRNA is present a low levels but is not translated due to a hairpin structure that inhibits its binding to ribosomes.
sRNAs (small RNAs) bind to this mRNA and inhibit formation of the hairpin.
sRNA-mRNA interactions are facilitated by a chaperone protein called Hfq.
Riboswitches Are a Developing
Area of Research
Riboswitches have been found to respond to many coenzymes, metabolites, and so on.
They are also found in eukaryotic introns and seem to regulate splicing.
Some riboswitches are unique to bacteria and are therefore a target for antibiotics.
Some RNAs Participate in Regulation
“Cis” regulation: a molecule affects its own function
“Trans” regualtion: a molecule is affected by another separate molecule
Lac operon as a model for regulation
The lac Operon Reveals Many
Principles of Gene Regulation
Work of Jacob and Monod − 1960
Shows how three genes for metabolism of lactose are regulated together as an operon:
β-galactosidase (lacZ):cleaves lactose to yield glucose and galactose
lactose permease (galactoside permease; lacY):transports lactose into cell
thiogalactoside transacetylase (lacA)
They rely on negative regulation via a repressor.
Lactose Metabolism in E. Coli
When glucose is abundant and lactose is lacking, cells make only very low levels of enzymes for lactose metabolism.
If glucose is scarce and cells are fed lactose, the cells can use it as their energy source.
The cells suddenly express the genes for the enzymes for lactose metabolism.
Lactose Metabolism in E. Coli
Inhibiting the Transcription of the
lac Operon via a Repressor Protein
The Lac repressor binds primarily to the operator O1.
The repressor also binds to one of two secondary operators, with the DNA looped between this secondary operator and O1
A gene called lacI encodes a repressor called the Lac repressor.
Structure of the lac Operon
Lac Repressor Bound to O1 and O3 with DNA Looped Between
The lac Operon Is Induced by Allolactose
How Lac Repressor Binds to DNA
Lac repressor is a tetramer.
The O2 sequence reflects the symmetry of the repressor.
There are approximately 20 repressors per cell.
The lac Operon Is Governed by More Than Repressor Binding
The availability of glucose governs expression of lactose-digesting genes via catabolite repression.
When glucose is present, lactose genes are turned off.
It is mediated by cAMP and cAMP receptor protein(CRP or CAP for catabolite activator protein).
When Glucose Is Absent, lac Operon Transcription Is Stimulated by CRP-cAMP
When Lactose Is Absent Little to No Transcription Occurs
When Lactose Is Present, Transcription Depends On Glucose Level
Two Requirements for Strongest Induction of the lac Operon
Lactose must be present to form allolactose to bind to the repressor and cause it to dissociate from the operator.(reducing repression )
[Glucose] must be low so that cAMP can increase, bind to CRP, and the complex can. bind near the promoter(causing activation)
Combined Effects of Glucose and Lactose on the lac Operon
inhibition(When lactose is low, repressor is bound)
permitting transcription(When lactose is high, repressor dissociates)
transcription is dampened(When glucose is high, CRP is not bound and)
activation(When glucose is low, cAMP is high and CRP is bound)
Protein factors that control transcription
Binding of Proteins to DNA Often
Involves Hydrogen Bonding
Gln/Asn can form a specific H-bond with adenine’s N-6 and H-7 H’s.
Arg can form specific H-bonds with the cytosine-guanine base pair.
The major groove is the right size for the α helix and has exposed H-bonding groups.
Protein-DNA Binding Motifs
A few protein arrangements are used in thousands of different regulatory proteins
and are hence called motifs.
helix-turn-helix(used by Lac repressor)
zinc finger
leucine zipper
and so on
The Helix-Turn-Helix Motif Is Common in DNA-Binding Proteins
Helix-Turn-Helix Motif
The Zinc Finger Motif Is Common in
Eukaryotic Transcription Factors
~30 aa
“Finger” portion is a peptide loop cross-linked by Zn2+
Interact with DNA or RNA
-Binding is weak, so several zinc fingers often act in tandem.
Binding can range from sequence specific to random.
Zn2+usually coordinated by 4 Cys, or 2 Cys, 2 His
Zinc Fingers
The Leucine Zipper Motif
Dimer of two amphipathic αhelices plus a DNA-binding domain
Each helix is hydrophobic on one side and hydrophilic on the other.
The hydrophobic side is the contact between the two monomers.
Approximately every seventh residue in helices is Leu (L).
Helices form a coiled coil.
The DNA-binding domain has basic residues (Lys (K), Arg (R)) to interact with polyanionic DNA.
Eukaryotic Gene Regulation
Amino Acid Biosynthesis Regulated
The trp Operon
SOS Response
Synthesis of Ribosomal Proteins and
rRNA is Controlled at Translation
When bacteria need more protein (as in cell growth), they make more ribosomes.
Ribosmal protein (r-protein) operons are regulated via translational feedback (next slide).
Translational Feedback Mechanism
Each operon for an r-protein encodes a translational repressor.
Repressor has greater affinity for rRNA than for mRNA.
rRNA Synthesis Is Also Regulated
by Amino Acid Availability
The stringent response occurs when aa concentrations are low.
Lack of aa produces uncharged tRNA.
Uncharged tRNA binds to ribosomal A site.
rRNA synthesis triggers a cascade that begins with binding stringent factor protein (RelA) to ribosome.
Stringent Factor Catalyzes Formation of
an Unusual Guanosine-Based Messenger
Stringent factor catalyzes formation of nucleotide guanosine tetraphosphate (ppGpp).
Binding of ppGpp to RNA polymerase reduces rRNA synthesis.
Stringent Response in E. Coli
其他
Three Features of Transcriptionally
Active Chromatin
Euchromatin = less-condensed chromatin, distinguished from transcriptionally inactive heterochromatin
Chromatin remodeling of
transcriptionally active genes:
nucleosomes repositioned
histone variants
covalent modifications to nucleosomes
Nucleosomes Can Be Restructured by Specific Protein Complexes
Features of Eukaryotic Gene Regulation
Access of eukaryotic promoters to RNA polymerase is hindered by chromatin structure.
Positive regulation mechanisms predominate and are required for even a basal level of gene expression.
Eukaryotic gene expression requires a complicated set of proteins.
Histone Modification Alters Transcription
Phase Variation – Regulation of Flagellin Genes
Covalent Modification of Histones
Methylation
Phosphorylation
Acetylation
Ubiquitination
Sumoylation
Occur mostly in the N-terminal domain of the histones found near the exterior of the nucleosome particle
Regulation by Gene Recombination
Processes can remove promoters relative to the coding sequence or can put genes into multiple orientations to alter the expression.
Example: Flagellin genes in Salmonella
In one orientation fljB is expressed with a repressor for fljC gene.
In another orientation, only fljC is expressed.
The process is called phase variation.
Nucleosome Remolding