Membraneless organelles Flashcards
What are membraneless organelles also called?
condensates
RNP granules
What are membraneless organelles?
No phospholipid boundary
protein & RNA
concentrate specific components
4 examples of condensates?
P bodies - control cell fate
Purinosomes - purine biosynthesis
Nucleoli - ribosome biogenesis
histone locus bodies - mRNA processing
What do membraneless organelles behave like?
liquid droplets
e.g. P granules drip around nucleus - move to only 1 side to determine cell fate
How do droplets coalesce?
join together
grow in size over tiem
dynamic reorganisation shown by FRAP
Why do molecules form droplets?
Reaction crucible
Sequestration
Organisational hub
What is LLPS (liquid-liquid phase separation)?
Reversible process
protein molecules concentrates in droplets
goes from low entropy to high entropy
How can polymer physics be used to explain LLPS?
energy of homotypic and heterotypic reactions
When X>0 - phase separation is favoured
What parameter can be used to compare energy between the heterotypic and homotypic interactions?
Chi parameter
What is the critical concentration?
above Ccritical - proteins form droplets
below - droplets disperse & dissolve
can modulate Ccritical by posttrans mods, temperature, & ionic strength
How do scaffolds help to drive membraneless organelle formation?
recruits client proteins/RNAs
What is IDR?
a functional protein region WITHOUT a unique structure
What are SLiMs?
short stretches of PROTEIN SEQUENCES that mediate protein-protein INTERACTIONS
What are LARKs?
Short stretches of PROTEIN SEQUENCES that can BIND weakly to EACH OTHER by forming pair of kinked B-sheets
What is multivalent?
comprising multiple binding sites for a ligand/protein partner
What is Ribonucleoprotein (RNP)?
complex of RNA and RNA-binding protein
How is LLPS regualted?
multivalent interactions between proteins and RNA = drive LLPS
allows molecules to mix together & associate at a high enough concentration to give scaffold structure
What determines recruitment to the scaffold
more multivalent = more strongly recruited
What are the features of proteins that drive membraneless organisation?
Stereospecific interactions between well-folded domains
Interactions of SLiMs and well-folded domains
Specific interactions between local structures in IDRs
Promiscuous interactions between IDRs
How can phase separation be promoted in cells?
- Change in conc, temp, pH, ionic strength
- Increasing valency
- Decreasing solubility - higher propensity to oligomerise or interact with another protein
What are post-trans mods that tune LLPS?
Lys acetylation Lys ribosylation Arg Methylation Arg citrullination Phosphorylation
Why is RNA a key player in LLPS?
act as scaffold for LLPS
RNA modifications alter structure
increased RNA length can promote LLPS
What is membraneless organelle formation driven by?
protein OR RNA
increase either protein-protein or RNA-RNA interactions
cause phase separation
How do molecules in cells also undergo transitions to the solid state?
pathological aggregates
amyloid fibrils
types of plaques
What liquid-to-solid transitions are associated with disease?
Motor Neurone Disease
Dementia
formation of amyloid fibres
What do stress granules contain?
mRNA - pre-initiation complexes
translation initiation factors
RNA-binding proteins & non-RNA-binding proteins
What do stress granules do?
control utilisation of mRNA during stress
implicated in diseases e.g. cancer, neurodegeneration
How are stress granules assembled?
40S 60S ribosome association with mRNA
puromycin promotes SG formation by stalling transcription
60S subunit falls off
mRNA-40S complex associates with RNA binding proteins that nucleate SG formation
How is stress granule assembly regulated?
form pre-initiation complex by recognition by eIF-4F complex and joining to eIF2 complex
Why does inhibiting translation promote SG formation?
inhibits the mTOR pathway
What happens to nuclear proteins in dementia patients?
proteins leave nucleus
become phosphorylated = do not go back into nucleus
proteins form amyloid fibrils
What is a key RBP involved in neurodegenerative disease?
TDP43
How does phase separation play a role in neurodegeneration?
RNP granule dynamicity
Trapping of cellular factors
How does phase separation play a role in cancer?
cellular signalling transcription
How does phase separation play a role in infectious diseases?
virus replication
cellular stress response
innate immune response
dormancy of bacteria/fungi
What is the key protein that assembles rotavirus factories?
NSP5
How does phase-separation affect SARS-CoV-2?
Nucleocapsid protein (N) undergoes phase separation with RNA Small molecules that bind N modulate droplet properties
