Vessicle trafficking: Endo and Exocytosis Flashcards by Joel Glotfelty

The transfer of proteins is post-translationally mediated by specific signal sequences for all organelles except

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Protein transfer to all organelles except ER is post-translational and follows synthesis on

Free ribosomes

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Protein transfer to all organelles except ER is post-translational and follows synthesis on free ribosomes and is mediated by organelle-specific

“signal” amino acid sequences

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Unique amino acid sequences target proteins to different

Organelles

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Proteins enter and leave the nucleus through

-mRNA also exits the nucleus through these structure

Nuclear pores

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Made up of an outer octagonal ring of protein and an inner central pore

Nuclear pores

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The nuclear pore is comprised of about

30 proteins

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The nuclear pore mediates traffic in and out of the

Nucleus

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Small molecules can diffuse through nuclear pores, but a transport system is needed for

Larger molecules

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Nuclear protein “signal sequence” is recognized in the cytoplsm by

Importins

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Proteins bound to importins are transferred through the nuclear pore and are then released from importins in the nucleus after binding to

RanGTP

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Induces the dissociation of the cargo protein by binding to the β subunit of importin

RanGTP

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To export from the nucleus, RanGTP induces binding of a cargo protein to

-transports protein to cytoplasm

Exportin

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The RanGTP/importin β complex is transported back to the cytoplasm, where RanGTP is converted to RanGDP by

RanGTPase activating protein (RanGAP)

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The RanGTP/importin β complex is transported back to the cytoplasm, where RanGTP is converted to RanGDP by RanGTPase activating protein (RanGAP), resulting in the release of

Importin β

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Used at many steps in intracellular trafficking

Monomeric GTP binding proteins

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The monomeric GTP binding proteins used at many steps in intracellular trafficking are inactive in their

GDP forms

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GTP binding proteins are involved in multiple aspects of cell trafficking including

) release of nuclear proteins (Ran)
) Transport vesicle formation (Arf, Sar-1)
) Transport vesicle recognition (Rab)

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Post-translational transport to peroxisomes uses an amino acid “signal” sequence” and is abolished in

Zellweger syndrome

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Proteins for peroxisomes are synthesized by free cytosolic ribosomes and then transported into

-abundant in the liver

Peroxisomes

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Phospholipids and membrane proteins are also transported to peroxisomes by the

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The major protein of the peroxisome

-decomposes hydrogen peroxide into water

Catalase

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A tetramer of apocatalase molecules assembled within the peroxisome

Catalase

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Added to each monomer to prevent it from moving back into the cytosol across the peroxisomal membrane

Heme

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A lethal condition caused by the defective assembly of peroxisomes due to the lack of transport of enzyme proteins (but not membrane proteins) into the peroxisome

Zellweger syndrome

Cells in patients with Zellweger syndrome contain empty

Peroxisomes

Transfer from the ER to the golgi occurs

Co-translationally

Makes up 50% of the total cell membrane

Associate with mRNA for secretory pathway proteins and begin translation in cytoplasm

Cytoplasmic ribosomes

While being synthesized on cytoplasmic ribosomes, the N-terminus of secretory pathway proteins binds the

Signal Recognition Particle (SRP)

There are three major steps during SRP-mediated protein translocation across the ER. The first step is that the binding of SRP to signal peptide causes a

Pause in translation

There are three major steps during SRP-mediated protein translocation across the ER. The second step is that the SRP bound ribosome attatches to the

SRP receptor in ER membrane

There are three major steps during SRP-mediated protein translocation across the ER. The third step is the dissociation of the

SRP and SRP receptor

What are some of the protein processing steps that begin during translocation through the ER?

1. ) signal peptide is removed 2. ) hydroxylation 3. ) disulfide bond formation 4. ) chaperone interaction 5. ) glycosylation

No proteins are exported from the ER unless they are

Properly folded

A critical processing element for the protein to be able to exit the ER

Glycosylation

A pre-formed oligosaccharide with 9 mannoses is added to the protein co-translationally from membrane lipid donor to

Specific residues (usually Asn)

The oligosaccharide is then modified by

Compartment-specific enzymes

We can deduce how far a protein has progressed in its synthesis by the extend of its

Modifications

The ability to trace sugar processing through cell compartments was critical in showing that transport through these compartments post-ER was via

Vesicles

Used for transport in both biosynthetic/secretory and endocytic pathways

Vesicles

What are the three generic steps for vesicle transport?

1. ) Form vesicle 2. ) Select cargo 3. ) Have address

What is the main advantage to vesicular transport?

Transporting cargo without crossing membranes

Vesicles in vesicular transport fuse with the membrane of the organelle is is delivering cargo to. Thus, there is no need for

Membrane crossing

To form the vesicle, you need protein "coats" which deform the

Membrane

The modifications and glycosylation of the protein will enable the vesicle to

Select its cargo

Allow vesicles to identify specific targets

SNARES

Coat proteins, cargo to be transported, and address molecules all need to be assembled during

Vesicle formation

Vesicle cargo is recruited by binding specific receptors that recognize specific signals on the

Cargo proteins

What are the three different protein coats that are used to deform the membrane during vesicle formation?

1. ) Clathrin 2. ) Coatamer (COP) I 3. ) Coatamer (COP) II

The formation of clathrin-coated vesicles (used at several transport steps) looks like direct binding between clathrin and the plasma mebrane, but is it actually?

Regulated by monomeric GTP binding proteins that cycle between active and inactive states

Assembly of COP-I and COP-II coats

What are the GTP proteins involved in the first steps of vesicle formation for 1. ) COP-I 2. ) COP-II

1. ) ARF | 2. ) Sar-1

The formation of the multi-subunit coatamer coated vesicles is initiated by binding to the donor membrane of a GTP binding protein that has become embedded in the

Plasma membrane

When GTP is bound, the lipid tail of Sar-1 is exposed. Sar-1 then insertes its tail into the

Plasma membrane

What happens once Sar-1 has used its tail to attach to the plasma membrane?

COP-II subunits bind the membrane

Specific cargo receptors can then bind to these

COP-II subunits

Used for transport from the ER to the cis-Golgi

COP-II

Used for retrograde transport, i.e. from trans-golgi to cis-golgi or from cis-golgi to ER

COP-I

Used fro transport from the cell surface to the early endosome, from the trans-golgi network to the late endosome, and from the late endosome to the trans golgi network

Clathrin

Mediate vesicle-target recognition and fusion

v- and t-SNAREs

Function as a lock and key. If there is a correct fit, they will form a coiled coil until the membranes fuse

v- and t-SNAREs

Located on vesicles

v-SNAREs (vesicle SNAREs)

Located on target compartments and bind v-SNAREs

t-SNAREs (target SNAREs)

Can also function in vesicle targeting by binding "effectors" when GTP is bound

Rab proteins

Cleave SNAREs and prevent vesicle fusion and neurotransmitter release

Botulinum and Tetanus Toxins

Predicted that different complementary vesicle (v-) and target (t-) SNARE membrane proteins are used for recognition in different transport steps

SNARE hypothesis

The process of cutting the budding vesicle from the membrane

Scission

Once scission has occurred, there is an uncoating event that exposes the

v-SNAREs

Before the vesicle is close enough for the v- and t-SNAREs to interact, there is a tethering even, whereby the vesicle is tethered to the new membrane, a process that likely involves

Rab

What are the 7 steps of the formation and fusion of vesicles?

1. ) Initiation 2. ) Budding 3. ) Scission 4. ) Uncoating 5. ) Tethering 6. ) Docking 7. ) Fusion

If transport from the ER to the golgi is mis-routed, then there is retrieval of the vesicle by COP-I via

KDEL sequence on escaped proteins

Sugar modifications that began in the ER continue in the Golgi by way of

Cisternal-specific Enzymes

The sorting of proteins to lysosomes, cell membrane,and secretory vesicles occurs in the

Trans Golgi Network (TGN)

Marked by the addition of phosphate to a mannose sugar residue

Lysosomal proteins

This modification is recognized by specific M-6-P receptors in the

Trans Golgi Network

Once the TGN recognizes that proteins are marked for the lysosome, the proteins are packaged into clathrin vesicles and delivered to the

Late endosome

The acidic pH of the lysosome then causes the ligand and receptor to

Dissociate

In which disease does M-6-P modification of mannose on lysosomal proteins not occur because the phosphotransferase required is mutated?

Human I-cell disease

In human I-cell disease, what happens to all of the proteins that were supposed to be lysosomal proteins?

They are secreted w/ secretory proteins

Enter constitutive or regulatory secretory pathways

Secretory proteins

The default secretory pathway with no sorting information

Constitutive secretory pathway

Proteins are greatly concentrated during

Transport

During transport, prohormones are

Processed

Entry into the cell occurs via

Phagocytosis and endocytosis

The phagosome hijacks host vesicles and becomes like the RER in -blocks delivery to lysosome and can replicate

Legionairre's disease

Acts as a sorting station

Early endosome

What are two types of cells that perform phagocytosis?

Neutrophils and macrophages

Can be followed by recycling of receptor, receptor | degradation in lysosome, or transcytosis to bypass tight junctions following sorting in the early endosome

Endocytosis

The process where vesicles are sent from the early endosome to the cell membrane for incorporation -Used to deliver antibodies to neonates

Transcytosis

Following internalization, surface receptors an cargo are sorted in the

Early endosome

The late endosome matures into the

Lysosome

Carries cholesterol through circulation as a lipoprotein complex

LDL

Cytoplasmic proteins that bind both clathrin and receptors for vesicle cargo as clathrin vesicles form

Adaptins

A mutation in the receptor tail can interfere with receptor binding to

Adaptin

Cargo bound to its receptor will not be incorporated into vesicles unless the receptor tail is bound to

Adaptin

Mutation in the LDL receptor tail can prevent cholesterol clearance, which can lead to

Hypercholesterolemia

Utilize endocytic and biosynthetic pathways for propogation

Enveloped viruses

The envelop fuses to the endosome at

Acidic pH

Occurs when receptors are delivered to the lysosome to be degraded instead of being recycled -Ex: EGF receptor

Receptor Down Regultion