Protein Sorting and Cell-Cell Communication

Covers content from lecture 27 - lecture 29

What is post-translation?

when proteins are fully translated on free-cytoplasmic ribosomes and can be sent to the nucleus, mitochondrion, chloroplasts, or peroxiosomes

True or False: Proteins need additional instruction to send proteins to a specific location, and proteins will remain in the cytosol if there isn’t additional information.

true

What is co-translation?

when synthesis begins in the cytosol, signal is presented, and the synthesis is completed inside of the rough ER

What proteins are made co-translationally?

• golgi

• rough or smooth ER

• transmembrane proteins in plasma membrane

• secretory proteins

• lysosomes

• endosomes

True or False: The presence or absence of a signal determines the protein’s fate (where it ends up).

true

What determines if a molecule needs energy to enter the nucleus?

the size of a molecule; small molecules (20 kDa) can diffuse across while larger molecules (60 kDa) needs to be transported across and requires GTP

What keeps cytoplasmic proteins out of the nucleus?

only the appropriate proteins have the right signal

What are signals?

sequences of amino acids that present a 3-D shape

What is the signal needed to allow molecules into the nucleus?

a nuclear localization signal (NLS)

True or False: NLSs can be artificially added to a molecule, and the molecule will enter a nucleus. NLSs can be removed from a molecule, and the molecule will not be transported into the nucleus.

true

What is important to know about the chemical structure of an NLS?

NLSs are rich in basic (positive) amino acids

What keeps nuclear proteins out of the cytoplasm?

only the appropriate proteins have the right signal—meaning the proteins contain a nuclear export signal (NES)

True or False: NES are rich in leucine.

true

True or False: Nuclear pore complexes don’t recognize NLSs.

true

What is an importin?

a type of protein that imports a cargo protein into the nucleus

What is important about importins?

it’s given a free pass across the nuclear pore complex along with anything attached to the it, and it’s able to recognize the NLS

True or False: After the importin takes the protein to the nucleus, the importin is sent back out into the cytoplasm.

true

What molecule recognizes NESs?

exportins

What recognizes exportins?

the nuclear pore complex

True or False: GTP is hydrolyzed to allow the import/export cycle.

true

What happens when GTP is hydrolyzed?

the exportin lets go of the NES

What is special about the NES?

it has a nuclear localization signal (NLS) as well

True or False: A protein can have both an NES and an NLS.

true

What is the mitochondrial matrix?

the innermost space of a mitochondria

What are the components of the human mitochondrial genome?

• 16.5 kb of DNA total

• 13 protein-coding genes

• 22 tRNA genes

• 2 rRNA genes

• contains an ori

• lacks ribosomal protein genes, DNA Pols, etc.

What do the protein-coding genes code for?

mRNA and components of the ETC

True or False: Most of the mitochondrial proteins are not coded for by genes on the mitochondrial genome.

true

Where are most mitochondrial proteins transcribed and translated?

nucleus and cytoplasm (respectively)

What is the mitochondrial pre-sequence/transient sequence?

a stretch of amino acids that tells a cell post-translationally to be transported into the mitochondria

True or False: An electrochemical gradient exists in the mitochondria.

true

What is important to know about the electrochemical gradient in the mitochondria?

• higher concentration of positive charges in the cytoplasm and the inter membrane space

• the matrix is more negative relative to the inner membrane space

What are transient sequences rich in?

basic amino acids

What surrounds the transient sequence to keep the protein unfolded?

chaperone proteins

True or False: TOM and TIM make up the channel that the proteins go through, and the transient sequence is compelled to go through the channel to the more negative environment in the matrix.

true

What cleaves the transient sequence?

transient peptidase

After the transient sequence is cleaved, what happens?

the protein is pulled the rest of the way in

True or False: Matrix chaperones are added to fold the proteins in the correct way.

true

If the matrix chaperones cannot fold the proteins correctly, what happens?

the protein is put into a semi-folded position and put into a chaperone complex

True or False: Not every protein needs the extra help with the chaperonin complex.

true

True or False: There is no mitochondrial export signal.

true

True or False: A RER protein import is exposed right away and recognized right away.

true

What are signal sequences/signal peptides mostly?

hydrophobic amino acids at the N-terminus

What recognizes the ER signal sequence?

signal recognition particles

True or False: SRP pauses translation in the ribosome and the ribosome, mRNA, and the emerging protein goes with the SRP and finds it’s way to the nearest ER.

true

True or False: SRP docks to a SRP receptor and goes through a channel.

true

True or False: GTP is used to have the protein let go of the ribosome and have the ribosome channel open.

true

When SRP is let go, what happens?

the emerging protein enters the cell

What cuts off the ER signal sequence?

signal peptidase

True or False: The emerging protein just goes in the ER due to the inertia by the elongation process of translation.

true

When is a ribosome done importing a protein?

• when the ribosome reaches a stop codon

• ribosomal subunits dissociate and float away

• channel slams shut when the ribosome lifts off to keep anything from leaking out

True or False: The outer membrane of the nuclear complex is part of the endomembrane system, but the inner membrane is not.

true

What is the perinuclear space?

the space in between the outer and inner membrane

True or False: If a molecule is in the perinuclear space, it cannot get into the inner membrane space without an NLS.

true

What organelles are in the endomembrane system?

• outer membrane of nuclear envelope

• lysosomes and endosomes

• rough and smooth ER

• golgi

What are the various types of cell-to-cell communication?

• direct intracellular

• contact-dependent

• autocrine

• paracrine

• endocrine

What is direct intracellular signaling?

signals pass through a cell junction from the cytosol of one cell to adjacent cells

True or False: Openings are usually created by intermediate filaments in direct intracellular signaling.

true

What is contact-dependent signaling?

signal molecules are attached to the cell and an appropriate cell with the correct receptor has to attach to the other cell; membrane-bound signals bind to receptors on adjacent cells

What is autocrine signaling?

cells release signals that affect themselves and nearby cells (local communication)

What is paracrine signaling?

cells release signals that affect nearby target cells

What is endocrine signaling?

cells release signals that travel long distances and affects target cells

What do all signaling types have in common?

• a signaling molecule

• receptor

• appropriate action is taken

What are the three stages of a response to a signal?

1. receptor activation

2. signal transductance

3. cellular response

What is receptor activation?

the binding of a signaling molecule causes a conformational change in a receptor that activates its function

What is signal transduction?

the activated receptor stimulates a series of proteins that form a signal transduction pathway

What is the cellular response?

the signal transduction pathway affects the function and/or amounts of cellular proteins—producing a cellular response

What is a ligand?

a signaling molecule that binds to the receptor —changing the receptor

True or False: Every receptor has its own specifc Kd (dissociation constant).

true

What is Kd?

the concentration of ligand at which 50% of the cell’s receptors for that ligand are bound

True or False: There’s an inverse relationship between Kd and affinity.

true

True or False: Higher affinity, the less ligand needed.

true

True or False: Ligand above Kd is likely to lead to a greater cellular response than a ligand below Kd because less ligand around produces less of a cellular response.

true

True or False: Ligand binding generally affects the conformation of the receptor—triggering an effect.

true

Why can steroids slide through the plasma membrane?

• small molecule

• nonpolar/hydrophobic

• receptors are not needed

True or False: Some receptors have intracellular binding domains.

true

For an intracellular signal localized in the nucleus, what would the receptors need?

an NLS

True or False: Some NLSs can be developed by the amino acid by being folded a certain way.

true

How do mRNAs get out of the nucleus?

• mRNAs are capped

• proteins with an NES bind to the mRNA and both structures are transported out of the nucleus

What are three examples of extracellular binding domains?

• ligand gated ion channels

• GPCR

• enzyme-linked receptors

What occurs in ligand gated ion channels?

the ligand binds to receptor molecules—leading to a conformational change in the effector domain; gate is opened and ions flow through

Why might cells be held in G0?

there aren’t enough growth factors around

True or False: The more steps you have, the more possibilities you have to regulate something and amplify the signal.

true

What is the process of receptor activation in the EGF pathway?

each of the phosphorylated tyrosines can lead to a different effect

What is the process of relay between the receptor and protein kinase cascade in the EGF pathway?

• a single signaling molecule can lead to different responses if different tyrosines get phosphorylated

• message is passed by changes in affinity

What is the protein kinase cascade in the EGF pathway?

• phosphorylation of proteins that will eventually lead to transcription

What is the general process of the EGF pathway?

1. receptor activation

2. relay between the receptor and protein kinase cascade

3. protein kinase cascade

4. activation of cellular response

5. cellular response

What are the structures of a G protein?

alpha, beta, and gamma subunits

True or False: The a-subunit of a G protein has GDP attached.

true

What is the GPCR process?

1. A signaling molecule binds to a GPCR—causing it to bind to a G protein and the GPCR changes shape.

2. G protein sticks to GPCR, and the G protein swaps GDP for GTP. Subunits dissociate (a and beta-gamma dimer dissociate) and carry out their own signal transduction pathway.

3. Signaling molecule dissociates from the receptor, and the a subunit hydrolyzes GTP into GDP and Pi. The a subunit and the B/gamma dimer reassociates.

What is the function of adenylyl cyclase?

breaks off two phosphate groups (the pyrophosphate)

What is the function of phosphodiesterase?

breaks the phosphodiester bond in nucleotides and nucleic acids

What is the process of epinephrine, GPCR, and second messengers?

1. The binding of epinephrine activates a GPCR. This causes the G protein to bind GTP—promoting the dissociation of the alpha subunit from the beta/gamma dimer.

2. The binding of the alpha subunit to adenylyl cyclase promotes the synthesis of cAMP from ATP.

3. cAMP binds to the regulatory subunits of PKA—release the catalytic subunits of PKA.

4. The catalytic subunits of PKA use ATP to phosphorylate specific cellular proteins—causing a cellular response.

What activates adenylyl cyclase?

the activated alpha subunit

True or False: Adenylyl cyclase will not form cAMP without being activated.

true

True or False: cAMP is a second messenger and can activate PKA.

true

What are the two subunits of PKA?

regulatory and catalytic subunits

True or False: cAMP binds to the regulatory subunit—exposing the catalytic subunit.

true

True or False: The catalytic subunit can phosphorylate other proteins.

true

What does activated PKA do in the epinephrine pathway?

• phosphorylates inactive phosphorylase kinase in order to activate the kinase

  • enzyme breaks glucose off of glycogen

• phosphorylates active glycogen synthetase—causing the kinase to be inactive

  • glycogen synthesis is inhibited

True or False: Phosphorylation can activate or inactivate proteins.

true