Organelles Flashcards
What are membrane-enclosed organelles?
Membrane-enclosed organelles are specialized structures within eukaryotic cells that have distinct functions and protein compositions.
How did membrane-enclosed organelles likely evolve?
They likely began with an expansion of the plasma membrane as cells grew larger.
Where are virtually all proteins encoded?
Nuclear genes.
What is the default destination for newly synthesized proteins?
The cytosol.
What are the three distinct mechanisms for protein import into organelles?
- Direct delivery from the cytosol
- Delivery to the ER, then transport via vesicles
- Active transport through nuclear pores.
What is the signal hypothesis?
The concept that proteins have intrinsic signals that govern their transport and localization in the cell.
What are targeting signals?
Specific sequences of amino acids that direct proteins to their organelle or location.
What is a signal sequence?
Continuous stretches of the polypeptide sequence made up of specific types of amino acids which can be located anywhere or in a specific region depending on the signal
Properties of amino acids is determined by their side chains (hydrophobic, positive/negative charge)
Signal sequence may be removed once the protein has been delivered as its not needed anymore
What is a nuclear localization signal (NLS)?
A signal that targets proteins to the nucleus, typically consisting of a high proportion of positively charged amino acids = Arg/Lys
Often short (less than 12 amino acids)
May be located anywhere in the protein and is NOT removed after the protein has been targeted
What is the function of nuclear pores?
- To act as gateways for the transport of molecules into and out of the nucleus.
- Small water soluble molecules diffuse freely through the pores
- Larger components including proteins & RNA are actively transported across the nuclear pore complex, in and out of the nucleus
What role does the small GTPase Ran play in nuclear import?
Ran controls the import of proteins into the nucleus by binding to receptors and facilitating cargo release
Exists in 2 conformations
- GDP-bound at the Cytosol
- GTP-bound at the nucleus
GDP-bound —> regulatory protein 1 (GAP), an activating protein, triggers GTP hydrolysis which results in quick conversion to the GDP bound state
GTP-bound —> regulatory protein 2, an exchange factor, promotes exchange of GDP for GTP which results in conversion to the GTP bound state
True or False: The nuclear localization signal is removed after the protein has been targeted.
False.
What happens to nuclear import receptors after they bind cargo proteins?
They are transported through nuclear pores and recycled back to the cytosol.
Fill in the blank: The energy for nuclear transport is provided by _______.
GTP hydrolysis.
What effect would a drug that blocks Ran’s ability to exchange GDP for GTP have on nuclear transport?
Nuclear import receptors would not release their cargo protein in the nucleus.
What is the significance of studying signal sequences through cut and paste experiments?
It helps determine if the nuclear localization signal is necessary and sufficient for nuclear import.
What are the general rules for protein targeting and import into organelles?
- Signal to target the protein to the correct subcellular destination
- Receptors that recognize these signals
- Transport machinery to cross membranes or be inserted into it
- Energy input e.g. ATP/GTP
What is the composition of nuclear localization signals?
Typically composed of less than 12 amino acids with basic/positive charged residues.
What cellular structures are involved in protein import into the nucleus?
- Nuclear import receptors —> binds to proteins with NLS in the cytosol
- Fibrils —> have direct receptor to pores. Receptors bind pore proteins and cargo proteins are moved into the nucleus
- Nuclear pores —> are large so folded proteins can be imported
What are the implications of raised calcium levels on nuclear import?
They stimulate NFAT transcription factors to enter the nucleus by exposing an NLS.
How does the Cytosol act as the default destination?
In the absence of any signal, newly synthesised proteins remain in the Cytosol instead of going to organelles
About 70% of proteins remain in the cytosol
Nuclear envelope
- composed of 2 membranes with an underlying lamina inside the nucleus which provides structure to the nucleus
- continuous with the ER (similar composition) = outer membrane
- contains proteins that bind to chromosomes and lamina = inner membrane
- protein network made of lamins = lamina
Typical NLS sequence
Pro,Lys,Lys,Lys,Arg,Lys,Val
Describe how Ran controls nuclear import
- Protein with NLS binds to the nuclear import receptor
- This moves the protein through the pores via interaction with the pore proteins
- Receptor and cargo protein is imported into the nucleus
- Inside the nucleus, Ran-GTP binds the receptor which displaces the cargo protein from the receptor
- Receptor bound to Ran-GTP is recycled back to the cytosol
- Ran hydrolyses GTP and dissociates from the receptor
- Cycle repeats and each cycle uses up GTP
