Intracellular Compartments and Protein Transport Flashcards
What is a general function for each of the following organneles?
- While going to lysosomes, molecular materials pass through endosomes -> sorting ingested molecules or recycling them back to the plasma membrane
- Also cytoskeleton:
- holding organelles in place
- transport of molecules (via motor proteins)
Why and in what ways could these membrane enclosed organelles evolve?
- In cells of the size of bactetium, all functions are conducted by the plasma membrane -> BUT this works only for prokaryotes -> the moment we increase the size and complexity the plasma membrane won’t be able to keep up
1) Invagination of the plasma membrane (likely allows for segregation of chemical reactions)
- e.g. lysosomes, Golgi a., ER
2) “Eating” a smaller structure which provides the function the plasma membrane wouldn normally encompass
- e.g. mitochondria, chloroplast
Why would we mean by “protein sorting”?
= process of accurately dividing and delivering types of proteins to specific parts of of the cell
- Some organelles derived - proteins directly from the cytosol e.g. mitochondria, chloroplast
- Other organelles use indirect route via ER e.g. Golgi apparatus, lysosomes
- proteins and lipids enter th ER -> some retained, some send further e.g. GA
Is this true: “All proteins are derived in cytosol”?
Although that is the case for most, which are synthetized on ribosomes exposed to the cytosolic environment -> some specialized proteins (mitochondria and chloroplasts) are formed inside the organelles’ ribosomes
- The direction and goal of movement of each protein is governed by a “sorting signal” = a specific amino acid sequence
Each protein faces the same issue when being imported into the cell - i.e. if fully hydrophobic how shall it cross the hydrophobic core
What 3 ways are utilized here?
1) Nuclear pores = special penetrations of both inner and outer layer of the nuclear envelope
- function as selective gates for transport of macromolecules and diffusion of small molecules
2) Protein Translocators = specific molecules that can unfold proteins and let them pass through
- utilized when going from cytosol to ER, mitochondria or choloroplasts
3) Transport Vesicles = “bags of proteins” that transport its content from one membrane compartment (e.g. ER) to another (e.g. Golgi a.) by endo- and exocytosis
How do we know what directs proteins to their final destination? What happens to it after?
Sorting signal is determined by sorting sequence = order and PROPERTIES of amino acids (since a lot of variety exists leading to the same organelle it may be that it is more determined by e.g. hydrophobic interactions, placement of charge, rather than the order alone
- Experiments
- removing the sorting sequence => becomes cytosolic protein
- adding sorting signal back on top of the protein => starts moving
What do you see in this picture - what are its functions?
Nuclear envelope is made up of 2 compartments
- inner nuclear membrane - containing binding sites for chromosomes and nuclear lamina = meshwork of filaments that stabilize/support the whole structure
- outer nuclear membrane - similar to ER
- perfurated by nuclear pores = gates for import and export of molecules
- some degree of selectivity due to proteins lining the pores -> soft tangled meshwork that allows only specific large molecules or small ones
How does the passage via nuclear pores work? What is being imported and exported?
Export: RNA molecules, Import: proteins from cytosol
- In order to pass large molecules need to display Nuclear localization signal which consists of 2 short sequences of multiple positively charged amino acids e.g. lysine, arginine
- this gets recognized by Nuclear import receptors -> together with the protein they penetrate the nuclear pore
-> once pore empty it will seal back again
- repeated sequences bind together forming a loosely packed gel
The import of nuclear proteins requires energy - how do we get it?
Provided by GTP hydrolysis mediated by a monomeric GTPase named Ran
NOTE: Unlike with crossing of mitochondria or chloroplast membrane - nuclear pore doesn’t require the protein to unfold = it passes in its final conformation
Some proteins that mitochondria and chloroplasts need for functioning are encoded by nucleus instead of the organelles themselves -> how do they get there?
Such proteins contain signal sequance at their N-terminus which allows them to target a specific organelle
- Enter at certain sites whether the membranes (outer and inner) come together -> proteins get unfolded -> pulled through the membranes by Chaperone proteins that also fold it back inside
- Once there the signal sequence gets removed
-> this might additionally reveal another signal sequence that would transfer the proteins to specific sites of e.g. inner or outer membrane
What other category of molecules needs to be imported to mitochondria/chloroplasts?
Lipids and phospholipids
- imported from the ER -> transported by lipid-carrying proteins that extract a phospholipid molecule from one membrane and place it in another
- usually at junctions where ER and mitochondria are close to each other
What are Peroxisomes?
= organelles found in eukaryotic cells that produce hydrogen peroxide
- break down of molecules e.g. toxins, alcohol, fatty acids
- synthesis of certain phospholipids e.g. those abundant in myelin sheath
How do peroxisomes receive proteins?
They are selectively transported from the cytosol
- contain 3 amino acid sequence which signals to receptor proteins -> transport towards peroxisomes -> uses translocator protein that aids the passage WITHOUT unfolding the protein
Some originate in ER - those are transported via vesicles
-> either fuse with the peroxisome membrane
-> or aids growing into a mature peroxisome
What is the main function of ER in terms of what has been discussed thus far?
Serves as an entry point for proteins destined to end up both in other organelles (Golgi, lysosomes, peroxisomes) as well as in ER
- will be ferried by transport vesicles
What are the 2 types of proteins transferred from cytosol to ER?
1) Water-soluabe proteins -> pass the whole ER membrane system and are released into the ER lumen
- destined to be secreted or serve the lumen
2) Prospective transmembrane proteins that get partly translocated and stay embedded in ER
- destined to reside in membrane
How do the proteins get through / How does this process differ from nucleus or mitochondria?
- Initially guided there via ER signal sequence = segment of 8 or more hydrophobic amino acids
- The proteins are passing through AS they are STILL SYNTHESIZED
- due to that ER has ribosomes placed on its membrane forming a site called rough ER (Membrane-bound ribosomes)
- Also no additional energy needed to push through
- NOTE: there are also “free ribosomes” that take care of all the proteins NOT being translocated through ER
How exactly do the soluable proteins get to ER lumen?
Two protein components help guide the way:
1) Signal-recognition particle (SRP) in cytosol binds to both ribosome and ER signal sequance that emerges from the ribosome (mRNA when translated binds to many ribosomes => polyribosome )
- this binding (SRP-ribosome) slows the synthesis process until…
2) SRP recepter on ER recognizes the SRP -> binding -> SRP gets released -> receptor passes ribosome to protein translocator in ER membrane -> synthesis and passing continues
What more can you say about the passing though ER and the signal sequence?
- Signal sequence (N-terminus) opens a channel in the translocater protein -> sequence stays bound to the channel while the rest of the chain is threaded thorugh the membrane
- Signal sequence gets removed by transmembrane signal peptidase -> released from the channel into lipid bilayer => degradation
=> Once C-terminus passed through the translocation channel protein will get released into the ER lumen
How does this process work in transmembrane proteins (in the simplest terms)?
Transmembrane protein with single membrane-spanning segment
- N terminus binds to the translocation protein -> the process continues until it gets halted at the Stop-transfer sequance -> N-terminus gets cleaved off while stop-transfer sequence stays in the bilayer forming alpha-helix structure
=> N-termines on the lumenal side and C terminus on the cytosolic
How is it with proteins that need to pass back and forth?
They use Internal signal sequance = Start-transfer sequance (instead of N-terminus)
-> initiates the translocation -> continues until stop-transfer sequance is reached
-> 2 hydrophobic seqaunces are then released into the bilayer where they form 2 membrane spanning alpha helices
- If it is multipass membrane protein there needs to be more start and stop signals in complex interplay
What kind of pathways take place in terms of “vesicular transport”?
- Secretory pathway = major outward route
- synthesis of proteins at the ER membrane -> enters ER -> within vesicles gets transported to Golgi organ (further modification)
-> either leads to cell surface
-> or branches off to endosomes and lysosomes
- synthesis of proteins at the ER membrane -> enters ER -> within vesicles gets transported to Golgi organ (further modification)
- Endocytic pathway = major inward route
- from plasma membrane -> endosomes and lysosomes
- meant for ingestion, degradation
What is important in vesicular transport?
- Vesicles take up only proteins that are meant to go with them e.g. some proteins may stay in Golgi apparatus
- Vesicles must transport it to the specific organelle NOT elsewhere
What do we mean by “coated vesicles”?
= vesicles tend to have a protein coat around their membrane when budding off -> it will be shed away to make fusion with the next organelle possible
- function: shaping of the membrane to form vesicles, capturing molecules inside the vesicles
How does the mechanism of forming a vesicle work in the best known “coat”?
The best known is Clathrin coat
- involved in both secretory and endocytic pathway
- Via adaptin clathrin molecules bind to cargo receptors
- molecules carry specific “transport signals” that are recognized by these receptors (in Golgi or plasma membrane)
-> create a basket-like coat -> dynamin narrows the membrane and pinches off the vesicle -> vesicle can move -> clathrin removed
