Eukaryotic Gene Regulation Flashcards
Eukaryotic gene regulation (Overall)
Eukaryotes have complex gene regulatory systems because have more complex and different cell types
Reason:
1. Different genes are required in different cell types – need ways to regulate
Where can Eukaryotic gene regulation occur
Eukaryotic gene regulation can occur at each stage
- Eukaryotic gene regulation can occur at multiple levels
Nucleus:
1. Chromatin packaging/unpacking
2. Transcription
3. Intron Processing
Cytoplasm:
1. Translation
2. RNA stability
3. Post-Translation Modifications
Movement through Euk cell
DNA –> Transcribed to Pre-RNA –> Splice RNA –> Goes out of nucleus –> Translated into proteins –> modified after protein is made = post translational modications
Layers of regulation in Eukaryotes
Eukaryotic gene regulation always involves multiple layers of regulation
- Have many steps –> Hard to disect them all
Where does gene expression occur
Gene expression (transcription) occurs in uncondensed regions of the DNA
- Uncondensed Eurochromatin = RNA polymerase can get in
- Packed Heterochromatin = RNA can’t get
Need tools to open DNA
Eukaryotic DNA
Eukaryotic DNA is looped around Histones –> Need to get to DNA to transcribe
Need tools to open DNA
Chromatin Remodeling
Chromatin remodeling is the result of Histone and DNA modifications
Acylation Vs. Methylation
Histone tail Acylation – Relaxes Chromatin (high expression)
- Opens chromatin
Histone Tail Methylation – Tighter compaction (low Expression)
- Closed
Nucleosome
8 Histone protein with Histone tail –> Can tag things onto the tail
- Can Acylate tail – opens histone structure
Genes involved in Chromatin Remodeling
A biazillion Different genes are involved in chromatin remodleing
Overall:
1. Writers
2. Readers
3. Earasers
Writers
Introduce modification on DNA and histone tails
- Put groups onto histones or DNA – Add modifications that tell readers what to do
Example – Histone Acytelases + Histone Methyltransferases + Histone Kinases + DNA methylases
Readers
Recognize modifications and recruit chromatin remodeling enzymes or recruit transcription factors
- Proteins to open DNA + recurit RNA polymerase
Earasers
Remove the modifications introduced by the writers
Example – Histone Deaxetylases + Histone Demethylases
After = can add new things
Types of transcription regulation in Eukaryotes
***Types of transcriptional regulation
- Negative regulation – Repressors + Co-repressors + Inducer molecules
- Positive regulation – Activators + inducers + inhibitors
***Sinilar to prokaryotes transcriptional regulations
Transcriptional Regulation (Euk)
- Multiple Activators – binds to enhancer sequneces
- Upstream activating seqeunces
- Insulators
Multiple Activators in Euk
Form of transcriptional regulation
Multiple activators are used in Eukaryotes gene regulatory systsems – Activators can bind to enhancer elements that are near (cis) or far (trans) from the promoter
Trans enhancer elements = upstream activating sequences
Cis vs. Trans
Cis – Near promoter
Trans – Far from promoter
Trans enhancer elements
Upstream activating sequences
Transcriptional Regulation Process (Euk)
Many Transcription Factors on the core promoter
Mediator binds – transcrioton activates from all over genome + no set active transcription
Transcriptional Activator protein – Binds to Upstream regulator sequnece
***Look at image
Insulators
Cis (near the promoter) regulator elements – recruits proteins that block the action of enhancer elements
Have enhancer upstream of promoter - where the activator would bind BUT have a protein that binds to the insulator –> prevents activator from binding to enhancer
Anser: Negative Regulation
Euk gene regulation + RNA processing
Eukaryotic can have gene regulation through mRNA processing
Example of Euk gene regulation through mRNA processing
Example – Drosphilla
Sex determination in flies:
XX:AA –> 1:1 – female – Sxl Is expressed
XY:AA –> 0.5:1 – male – sxl is not expressed
It is the ratio of X:A chromosomes that controls sex determination
- No X-inactivation in flies
Females = make more X protein = have extra after dimerization = protein can bind to SXL = get female development
SXL gene in Drosphilla
SXL Express = female development
SXL not expressed = male development
sxl = a splicing factor that creates the Tra protein that leads to female development
Answer: Positive regulation – Activator yellow binds and get transcription
Genes in Dropshilla sex determination
Tra gene = has 2 introns –> have exon with stop codon
If have splicing factor (have sxl) = splice exon out –> Get Tra protein = get female develeopment
No Sxl = both introns removed = get mRNA with 4 coding domaines but have stop codon = get protein but early stop codon = male
General Structure of Euk mRNA
Euk mRNA contains:
1 Open reaidng frame (sometimes with introns that need to be removed)
5’ cap (added after transcription)
3’ Poly A tail (part of transcription termination sequence)
Regulatory information in the 5’ and 3’ UTRs
Answer: B –> Need both together to regulate translation
Poly A and 5’ cap + amont of protein made
Eukaryotic gene regulation through mRNA translation and stbability
Options: RNA + Splicing + PolyA tail
- Once mRNA is exported out of nucleus –> the 5’ cap binds to the PolyA tail – see which protein is avalable
- If less complex is formed = translate less
- Can target RNA for degredation = not translated
- Poly A tail –> recurits proteins + CAP can recurit proteins –> regulate translation by giving different proteins on the PolY A tail so can’t leave nucleus
Preventing mRNA from leaving nucleus
Poly A tail –> recurits proteins + CAP can recurit proteins –> regulate translation by giving different proteins on the PolY A tail so can’t leave nucleus
Gebe regulation through RNA switches
Riboswitch
Riboswitch
A secondary structure in RNAs that occur when a ligand binds that result in a change in protein Synthesis
- Influence ability of RNA to be translated
- Form a secondary structure – bind to ligand causing another secondary structure = blocks ribosome = ribosome can’t do translation
***Typically in the 5’ UTR region
Example of regulation through RNA swicth
Thiamine (B12) biosynthesis
Thiamine binds to the mRNA preventing translation of an enzyme needed to synthesize thiamine
- Reduce thiamine = opens = get translation
Answer: Negitive Feedback loop –> Get thiamine and stop thiamine biosynthesis
Ligand in Riboswitches
With riboswicthed Ligand is often the product of gene expression
Thermoswicth
A regulatory segment of mRNA that forms under certain temperatures
Example:
Low temp – Structure is more stable –> ribsome can’t get through = reduce development
High Temp –> secondary structure falls apart = get proteins = plant can develope
***Variation of riboswitch
What can Riboswitches do?
- Block transcription by creating transcription termination sequences
- Recruit factors that cleave RNA – secondary structure can be bound by endonuclease
- Recruit splicing factors to alter intron processes
- Block Ribosome binding site preventing translation
Low temp = not tranlsated = male
High temp = translated = female
RNAi
Gene regulation through ncRNAs
dsRNA in cells
dsRNA are targeted for degradation
- Regulate gene expression by getting rid of RNAs
RNAi processes
Short non-coding RNAs that are complimentary to speciefic mRNA can lead to the degradation of those mRNA targets and therefore lower protein expression
- RNA product of ncRNA is complementary to blue = bind = dsRNA = targeted for degradation
- Antisense RNA – Cuts off siRNA or miRNA – important regultor factors
siRNA or miRNA
21-25 nucleotides long
Important regulatore factors
Antisense RNA
Cuts off siRNA or miRNA – important regultor factors
RNAi in cells
RNAi = natural cell process – get rid of blue by modulating amount of Red RNA
RNAi in research
RNAi has been adopted by researchers to lower the gene expression by lowering mRNA levels
Image – getting rid of green RNA by imposing complementary RNA –> binds to mRNA – want to block to target for degradation
RNAi process
Synthetic gene sepcific siRNA –> siRNA delivered into cell –> siRNA enters into RISC and unzips –> complementary pairing –> target mRNA –> Target mRNA cleavage and degradation
RNAi in Drosphilla
RNAi in drosphilla = Knock-down
Have activator downstream of tissue specific promoter –> binds to upstream activator sequence that is engineered to express RNA version of gene –> Make complementary gene
- Fly makes GAL4 in liver – have siRNA only in the liver –> degrade protein only in lover – easy to study if regulate level in certain places but not everywhere
have siRNA that is complementary to gene (YFG) + have a tissue specific promoter + have Upstream activating sequnece –> Tissue specific promoter activator binds to upstream activating sequence – get expression of gene –> mRNA for YFG is targeted for degration leading to less YFP protein
RNAi in Dropshila process
Fly makes GAL4 in liver – have siRNA only in the liver –> degrade protein only in lover – easy to study if regulate level in certain places but not everywhere
Gene regualtion through ncRNA
- RNAi
- lncRNAs
Gene regulation through lncRNAs
- Transcriptional Regulation – binds to DNA blocking transcription
- Chromatin Modifications
- Signaling – Add signal to protein
- mRNA sponge – soak up RNAs
- Scafold – brings proteins
- Translational regulation
Gene regualation through post translational modifications
Modifcations to proteins affects their activity
Example – Phsophorylation of an intiation factor blocks translation in the ribosome
- Translation factor = phosphorylation = no translation
- No Phosphoryltion = translation
Example Post-translational modifications
Ubiquitin – targets modified proteins for degradation
- Target protein for degradation by tagging with ubiquitin
Gene regulation tool kit
Gene regulation + Imagination
If you can imagine it then biology has tried it – and iff it works then it will be there
***Use your tool kit of regulatory mechanisms to imagine a bazillion ways that gene expression may be regulated
