Cell bio: Post Transcriptional control: mRNA degradation, translational control, Post translational control: protein degradation control, protein activity control

mRNA degradation control

What do exonucleases do?

• cut nucleic acids from outside in

What does mRNA degradation always start with?

• removal of poly A tail

What protein complex removes poly A tail?

• CCR4/NOT

What occurs during 5’ to 3’ degradation?

• decapping degradation form 5’ end

What complex helps with decapping and what chops up mRNA?

• DCP complex help remove cap

• XRN1 chop mRNA

What occurs during 3’ to 5’ degradation?

• exosome degrades from 3’ to 5’

How does miRNA mediated gene silencing work?

1. miRNA processed in nucleus and is moved to the cytoplasm

2. dicer cleaves to make smaller miRNA

3. one strand of miRNA loaded onto RISC

4. RISC binds target RNA causing degradation by argonaute

5. translation reduced

miRNA mediated gene silencing works with what kind of RNA?

• endogenous RNA (made by cell)

How does siRNA mediated RNA interference work?

1. foreign double-stranded RNA enters cell

2. dicer cleaves to siRNA

3. one strand of siRNA loaded onto risc

4. risc bind to target RNA

5. argonaute cleaves target RNA

6. translation reduced

are precursors made in the nucleus during siRNA mediated RNA interference?

• no

What does the 2nd pathway of siRNA mediated RNA interference do?

• allows cell to destroy foreign mRNA before its completely transcribed

What protein does the siRNA bind to?

• RITS complex

What are the three phases of CRISPER?

• acquisition

• expression

• interference

Acquisition

• viral fragments inserted into crisper locus in genome

• creates genetic “memory” of infection

Expression

• crisper locus transcribed

• RNA from locus processed into crisper RNA (crRNA)

• crRNA bind CAS protein

Interference

• crRNA guides CAS to matching viral DNA

• viral DNA cleaved

• infection prevented

Translational control

What does translational control/regulation do?

• controls how efficiently an mRNA is translated into protein

What is faster and more energetically favorable, transcription or translational control?

• translational control

Where does translational control occur?

• initiation stage

Which two levels do hairpin structures (RNA base paired together into secondary structure) regulate translation?

• initiation

• elongation

What does hairpin do during initiation?

• hides ribosomes binding region

• prevents ribosome from binding

What does hairpin do during elongation?

• slows/stalls translation

How do RNA binding proteins (RBPS) reduce translation?

• RBP binds in 5’ UTR covering ribosome binding site

• ribosome can’t bind to mRNA

How do RBPs increase translation?

• RBPs bind to teh 5’ cap and 3’ poly A tail, loop RNA into circular structure (protects/stabilizes RNA), increases efficiency of translation

• RBPs bind to regulatory sequences in 3’ UTR, block enzyme that degrades mRNA (CCR4 NOT), increasing translation by protecting transcript from degradation

Post translational control

Protein degradation control

What are proteases?

• enzymes that cleave proteins

• cleavage can result in degradation of the enzyme or altered function

What are proteosomes?

• protein complex where proteins are broken down

What is ubiquitin?

• covalently attached unit that marks proteins for degradation

• protein ready for degradation when many copies (polyubiquitin) attach

What enzymes are required for ubiquitin mediated protein degradation?

• E1 ubiquitin activating enzyme

• E2 ubiquitin conjugating enzyme

• E3 ubiquitin ligase

E1 ubiquitin activating enzyme

• use ATP to activate ubiquitin

E2 ubiquitin conjugating enzyme

• receives ubiquitin from E1

E3 ubiquitin ligase

• recognizes target protein

• works with E2 to attach the ubiquitin molecule to target protein

What does deubiquitinating enzyme do?

• remove ubiquitin

• increase translation

Protein Activity control

Negative regulation

• prevents enzyme from function

Positive control

• stimulates enzyme function

Feedback inhibition

• end product reduces activity of enzyme

Feedback activation

• end product enhances activity of enzyme

What is allosteric regulation?

• binding of a regulator to a site other than the active site to alter the function of an enzyme

Competitive inhibition

• inhibitor binds to active site preventing the substrate from binding and decreasing enzyme activity

• concentration dependent

What can covalent modifications do?

• change if protein is on/off causing conformational changes

• change location of protein

• serve as a docking site for other proteins

Kinase

• adds phosphate (negative charge)

Phosphatase

• remove phosphate