Cell Bio Test #3: Intracellular Transport 1

Cytosol

metabolic pathways, protein synthesis

nucleus

contains nuclear genome, DNA/RNA synthesis

mito

ATP synthesis via oxidative phosphorylation

chloroplasts

ATP synthesis and carbon fixation via photosynthesis

Golgi apparatus

proteins and lipid modification and sorting

endoplasmic reticulum (ER)

mem synthesis, protein distribution
main site of lipid synthesis
Rough: ribosomes attached
Smooth: Ca2+ sequestration

lysosomes

intracell. degradation

endosomes

sorting of endocytosed material

peroxisomes

contain enzymes that produce H2O2, proteins don't need to unfold to enter perox.
contains protein transolcator that aids in transport
oxidation of toxic molecules

Endomembrane system

ER, Golgi app, peroxisomes, endosomes, lysosomes

What are the 3 mechanisms of protein transport for an imported water-soluble protein into organelles?

All of these processes require energy
2) needs to be unfolded

1) Transport through nuc. pore: folded proteins enter via nuc pores
2) Transport across mem: unfolded proteins enter organelles via protein translocators (must unfold to get across mem)
3) Transport by vesicles: folded proteins move via transport vesicles that fuse w/ other mem (pinches off mem to form vesicle that fuses w/ another part of a mem)

nuclear pores

selective gates but allow free passage of small, water-soluble mol
penetrates outer and inner mem

What do signal sequences do?

direct proteins to the correct organelle

What are ss comprised of?

a stretch of 3-60 aa

The final destinations have the same or different signal seq?
-If there is no signal sequence on the protein, where does it stay?

they have diff ss;


it stays in the cytosol

structure of nucleus

Folded proteins w/ nuclear localization siganls (NLS) are bound by what type of receptors?
-What crosses the nuclear envelope via the nuclear pore?

bound by nuclear import receptors;


Nuclear import receptors and cargo(prospective nuclear protein) to be able for the protein to get delivered to the nucleus

What does Ran-GTP do?

it binds to the nuclear import receptor in the nucleus and then returns it to the cyto.

once in nuc, Ran-GTP binds to receptor and exits via nuc pore into cyto, ran hydrolyzes the bound GTP. Ran-GDP dissociates from receptor

What two things mediate the activity of Ran?

Ran-GAP (GTPase activating protein) and RAN-GEF(guanine exchange factor)

How are unfolded proteins able to enter the mito and chloroplasts?

via protein translocators

What two things help drive import into the mito?

Chaperons and mem potential

Peroxisomes use what two things to remove protons?
What do perox do?
What are perox involved in the formation of?

O2 and H2O2;
break down F.A, toxins, alch

plasmalogen: phospholipid that is needed for neuron function

How do peroxisomes import proteins?

via peroxins that use ATP as energy source

Rough ER

When translating proteins that have an ER ss, ribosomes that attach to it, are targeted where?
The free ribosomes that have no SS attached, go where?

to the ER


they remain free in the cytosol

ER protein targeting depends on what?
ER proteins are threading through what?

the signal recognition particle (SRP) and the SRP receptor;
through protein translocator (to get into the ER)

SRP and SRP receptor bind and hydrolyze what?

GTP

After the ss is cleaved from the proteins, what happens to the soluble proteins?

the mature soluble protein is released into the ER lumen

What determines the arrangement of the ER mem protein?

they hydrophobic start transfer and stop transfer sequences

What determines the orientation of protein insertion?
Before the start transfer, what is N terminus?
After the start transfer, N terminus is what?

the location of positively charged aa;
cytosolic;

lumenal

The outer nuclear mem is continuous w/ what?

b/c of this what does outer nuc mem have on it?

the ER.


ribosomes

Many organelles are held in their locations by what?

by their attachment to the cytoskeleton

When is the ss removed from finished protein?

once it has been sorted

Deleting a ss from an ER protein does what?

converts it to a cytosolic protein

inner nuc mem

contains proteins that act as binding site for chromo, and proteins that provide anchorage for nuc lamina

ribosomes are bound to which surface of ER mem?

cytosolic surface

nuclear pore complex

forms a gate in which selected macromolecules and larger complexes enter or exit the nucleus

What is the ss that directs a protein from the cytosol into the nucleus?

nuclear localization signal.

What energy drives nuclear transport?

the energy supplied by GTP hydrolysis

nuclear import receptors interact w/ what?

cytosolic fibrils that extend from the rim of the poe

What happens after a nuclear import receptor picks up a prospective nuclear protein in the cytosol and then enters the nucleus?

Then the nuclear import receptor encounters GTPase (Ran), which carriers a mol of GTP. This ran GTP binds to the import receptor=nuclear protein is released in the nucleus.
-Receptor is still carrying Ran-GTP, so goes back to cytosol, and some protein triggers Ran to hydrolyze the GTP that is bound to it. Ran-GDP falls off the receptor which makes it free to bind another protein that needs to go to nuc.

What do chaperons helps to do inside the organelles?

They help to pull the protein across the 2 mem and fold it once inside

Transport initiation outside the outer mem

The precursor protein is recognized by a receptor in the outer mem. This receptor is associated w/ a protein translocator. Then the receptor, precursor protein, and translocator diffuse across outer mem till it reaches 2nd translocator in inner mem. The 2 translocators transport protein across both mem, unfolding protein in process. ss cleaved off by peptidase and chaperons help to refold these

What organelle serves as an entry point for proteins that are destined for other organelles, as well as this organelles itself?

ER

Proteins destined for where all enter the ER first from the cytosol?
What happens once these are inside the ER lumen?

golgi apparatus, endosomes, lysosomes;

proteins will be transported via vesicles from organelle to organelle or to plasma mem. They will NOT re-enter cytosol

What 2 types of proteins are transferred from the cytosol to the ER?

water-soluble proteins: completely transported across ER and then enter ER lumen
prospective transmem proteins: partially transported and then become embedded in it

ER ss is?

hydrophobic

Why is it necessary for the ribosome protein to be attached to the ER mem?




What do all of theses ribosomes make up ?

b/c most proteins that enter the ER are threaded across the ER mem before the polypeptide chain has been completely synthesized. So the ribosome is synthesizing the protein while it is going through ER mem;

Rough ER

mem-bound proteins




Free ribosomes

attached to the cytosolic face of ER
-make proteins that are being translocated in ER

unattached to any mem
make proteins that are encoded by nuclear DNA

Why do mem bound ribosomes need no additional energy required for transport?

b/c the elongation of each polypep provides the thrust needed to push it through the ER mem

As an mRNA molecule is being translated, what happens? What is this called?

many ribosomes bind to it, forming a polyribosome

Soluble proteins that are made on the ER are released where?
Some of these remain embedded in the ER mem as what?

in ER lumen;


as transmem proteins

SRP is present where?
What does it bind to?

cytosol;
both ribosome and ER ss

SRP receptor is embedded where?

ER mem;
it recognizes SRP

SRP binds to ribosomes w/ ER ss, protein synthesis slows until SRP binds to SRP receptor. After binding, What happens?

the SRP is released. Receptor gives ribosome to protein translocator in ER mem, protein synthesis resumes, polypep chain is threaded across channel in translocator, translocator transfers growing polypep across lipid bilayer

Where is ss located on soluble proteins?

at N-terminus

In addition to direction protein to correct organelle, what does the ss also do?

it functions to open the channel in the protein translocator, ss remains bound to channel while chain is threaded through. After, the ss is released from channel and degraded in lipid bilayer

When is the protein released into the ER lumen?

after the C-terminus of a soluble protein has passes through the translocation cahnnel

N- terminal ss OR internal signal sequence (start-transfer)
stop transfer seq

initiates translocation;
additional seq of hydrophobic aa

When the stop transfer seq remains in bilayer after N-term is cleaved what happen?

the stop transfer seq forms an a-helical mem spanning segment that anchors protein to mem

What is special about the internal signal sequence (start-transfer seq)

it is NEVER removed from the polypep) unlike the N-term that is cleaved off in some mol

single-pass transmembrane protein

N-terminal ER ss initiates transfer,
contains 2nd hydrophobic seq that acts as stop-transfer seq. After goes through channel, N-terminal is cleaved off and the transmem protein is anchored in mem. Then protein syntheisis on cytosolic side finishes

double-pass transmembrane protein

internal sequence (start transfer) acts as start signal and helps to anchor to final protein in mem. Eventaully, when stop-transfer enters translocation channel, coth seq are discharged into bilayer. NEITHER seq are cleaved. polypep chain stays anchored in mem

Proteins go from ER-->G.A-->?
How is this carried out?

to other organelles. This is carried out by vesicular transport. Routes of comm between interior of cell and around it

Secretory pathway

starts w/ synthesis of proteins in ER mem, ER, G.A, side branch leads off thorugh endosomes and lysosomes

Endocytic pathway

responsible for ingestion/degradation
-moves mol from plasma mem through endosomes to lysosomes