Title: BV2 / BV2 - 2 / Modul 4 / Membrane transport and signalling

Q: What type of environment is the membrane?
A: Hydrophobic environment

Q: What is needed because of the hydrophobic membrane-environment?
A: We need systems that can transport molecules over the membrane

Q: Vilka typer av molekyler behöver hjälp för att ta sig över membranet?
A: Polära molekyler och joner

Q: Which compounds cannot pass the membrane?
A: Biological membranes are impermeable for most hydrophillic and/or charged molecules

Q: In which 2 ways can membrane transport occur?
A: Passive transport and active transport

Q: What can passive transport be categorized into?
A: 1. Diffusion
2. Facilitated diffusion

Q: What can active transport be categorized into?
A: 1. Primary
2. Secondary

Q: How can diffusion be described with key words?
A: - Linear relationship
- Mass action
- No saturation

Q: How are thermodynamics involved in diffusion?
A: Theromdynamics will attemt to euqulize the sides until u have an  equilibrium

Q: Is there a limit for diffusion?
A: There is no limit of molecules that can diffuse at the same time

Q: What drives diffusion?
A: Concentration gradient

Q: What are the general principles of facilitated diffusion?
A: - Saturation curve - looks like Michaelis-Menten curve
- Mass action

Q: How does the curev of facilitated diffusion change with concentration?
A: At low concentrations its the same linear relationship but the higher concentration there is u get a saturation curve similar to those of enzymes

Q: Is there a capacity limit for facilitated diffusion?
A: There is a capacity to exceed - tells us that there is something in the membrane that facilitates diffusion

Q: What happens in facilitated diffusion?
A: A protein binds the factor and encapsulates it in a complex that can cross the membrane

Q: What does channel proteins and pores not have?
A: As a channel or pore u don't have the facilities to facilitate transport in one direction or the other

Q: How does facilitated diffusion with transport proteins work, briefly?
A: The compound binds to the binding site -> a conformational change occurs
- It is reversible

Q: In which directions will a facilitated diffusion transport protein work?
A: Transporter will work in both directions

Q: How can facilitated diffusion be explained step-by-step?
A: 1. The carrier is open to the outside
2. X enters from outside and binds at a binding site
3. The outer gate closes and X becomes occluded, still attached to its binding site
4. The inner gate opens with X still attached
5. X exits and enters the inside of the cell
6. The outergate closes, occluding an empty binding site. This cycle can also flow in reverse order

Q: How can carriers be described?
A: Carriers are conduits that are gated by 2 "doors" that are never open at the same time

Q: How can one make transport proteins controllable?
A: Via:
- Conformational change the flux can be controlled

Q: What is the key with gated diffusion?
A: The key is preventing gradients from equilibrating

Q: What are pores always?
A: Always open

Q: Are channels always open?
A: Channels can open and close

Q: What can be said about a carriers cycle?
A: Carriers have a certain cycle, never a complete transfer
- Makes sure that only one type of molecule can pass this pore

Q: When is active transport used?
A: When compounds are being transported
against the gradient

Q: How can osmosis be described?
A: If the concentration of one ion is higher on outside than inside it will flow thermodynamically from outside to inside

Q: How many ions does the Na+K+ pump pump with each round?
A: In - 2 K+
Out - 3 Na+

Q: What does the sodium-potassium pump create?
A: Creates a net positive charge on the outside
- Charged gradient between the different sides of the membranes
- Also a pH gradient, will in turn drive other transport processes

Q: How does the sodium-potassium pump work?
A: Powered by ATP, it moves K+ and Na+ ions in opposite directions, each against its concentration gradient
In a signle cycle of the pump, 3 sodium ions are extruded from and 2 potassium ions are imported into the cell

Q: What is an example of primary active transport?
A: Uniport and cotransport - Pump-mediated

Q: What does active transport create?
A: Creates change- and concentration gradients

Q: What are examples of secondary active transport?
A: Antiport and symport - carrier-mediated

Q: What do antiports and symports do?
A: The couple the flow of one molecule against its concentration gradient with the flow of a 2nd molecule against its concentration gradient

Q: Where can secondary transport be performed?
A: Can be performed when there is a gradient from the primary form

Q: What is an ex of active transport?
A: Intestinal glucose uptake

Q: Where can we fins membrane transport in the body?
A: In the intestine - intestinal nutreint uptake

Q: What can be coupled to sodium gradients?
A: Aminoacids

Q: What do u need to pull out all the nutrients?
A: To pull out all the nutrients u need active transport to the epithelial cells

Q: What do molecules without transporters do?
A: Many molecules don't have transports, ex. signals
- Need to go into the cell/start a function i the cell
- Instead they have receptors on the cell

Q: Why do we need signaling between the cells?
A: - Every cell needs to know what to do
- Without signals every cell would die - there are signals telling it to survive
- Other signals are needed for proliferating

Q: What do signal molecules do together?
A: They work in combinations to regulate the behaviour of the cell

Q: Via which mechanism can cells receive signals from other cells?
A: - Conctact dependent - gap junctions/communicating junctions
- Paracrine signalling

Q: What determines which receptors a cell has?
A: Each cell type dusplays a set of receptors that enables it to respod to a corresponding set of signal molecules produced by other cells

Q: What do mitogens do?
A: They stimulate cell division by relieving intracellular negative controls that otherwise block progess through the cell cycle

Q: What do growth factors do?
A: Stimulate cell growth - an increase in cell mass - by promoting the synthesis of proteins and other macromolecules and by inhibiting their degradation

Q: What do survival factors do?
A: They promote cell survival by suppressing apoptosis

Q: Which types of cell signalling are there?
A: - Endocrine
- Paracrine
- Autocrine
- Synaptic
- Contact-dependent

Q: In which 2 ways can signal molecules be sensed?
A: - On the surface
- Inside the cell

Q: Which signalling molecules bind to receptors outside the cell?
A: Hydrophilic

Q: Which signalling molecules bind to receptors inside the cell?
A: Hydrophobic

Q: Where can receptors be found inside the cell?
A: In the nucleus or cytosol

Q: How are hydrophobic signals transported to the cell?
A: Since they are nearly insoluble in aqueous solutions they are transported in the bloodstream and other extracellular fluids after binding to carrier proteins, from which the dissociate before entering the target cell

Q: What do hydrophobis signal molecules do?
A: They go through the membarne - find a receptor - excerts its function/purpose

Q: Are all molecules that bind to nuclear receptors small or hydrophobic?
A: No, e.g. the active hydroxylated form of Vitamin D3 as well

Q: Briefly, how does a suface receptor relay the signal of a signalling molecule?
A: An exctracellular stimulus is relayed by the receptor - intracellular signalling side - intracellular signalling cascade - function

Q: Why are surface receptors useful?
A: - One activator on the outside can activae many things on the inside
- Amplification of the signal
- Dampening (have an OFF-switch)
- Integration of multiple signals

Q: What are the 3 phases/steps in signalling with surface receptors?
A: Reception -> Transduction -> Response

Q: What are the 3 largest classes of cell surface receptors?
A: - Ion-channel-linked Receptor
- G-protein-linked receptor (GPCR, 7-TM)
- Enzyme-linked receptor

Q: What are 7 TM receptors?
A: They have 7 alfa helixes that cross the membrane
- N-terminal extracellularly - C-terminal intracellularly
They are coupled by a type of protein family called G-proteins

Q: What are 3 examples of 2nd messengers?
A: - Ca2+
- cyclic AMP
- Inositol 3,4,5-phosphate

Q: What are examples of -crine signalling molecules that bind to G-receptors?
A: - Epinephrine
- Norepinephrine
- Dopamine
- Histamine

Q: What are the 2 classes of cell surface signalling molecules?
A: 1. Small molecules
2. Protein/peptide ligands

Q: What are examples of peptide ligands?
A: - Insulin
- Glukagon
- CCK
- Gastrin
- Opioids
- Vasopressin
- ACTH
- Growth hormone
- Somatostatin

Q: What are examples of GI-peptide ligands?
A: - Insulin
- Glukagon
- CCK
- Gastrin

Q: What are endocrine hormones?
A: The
hormones that travel the whole bloodstream

Q: Which cells don't have a receptor for glukagon?
A: - Adipose cells
- Muscle cells

Q: Which cells have a receptor for glukagon?
A: Liver cells

Q: What are our glukagon levels in a fasting state?
A: High glukagon levels

Q: What happens when we eat a meal?
A: Blood glucose levels shoot up and as a result insulin spikes and glukagon becomes lower

Q: What does glucose do regarding glukagon?
A: Glucose inhibits glukagon synthesis, so does insulin

Q: What happens in the absence of glucose, regarding glukagon?
A: In the absence of glucose u get a Ca2+ influx that releases glukagon

Q: What are some facts about glukagon?
A: Peptide hormon
28 amino acids
Proglukagon has 180 amino acids
Made in the ER - travels to the golgi - is realesed upon stimuli

Q: What type of hormone is glukagon?
A: Peptide hormone

Q: How many aminoacids does glukagon contain?
A: 28 aa

Q: How many amino acids does proglukagon have?
A: 180 aa

Q: Where is glukagon made in the cell?
A: It is synthesized in the RER, it then travels to the glogi and is then released upon stimuli

Q: What happens with proglukagon?
A: Since it's cleaved in pieces it's cleaved into glukagon 1 and 2 and has different functions from glukagon

Q: What can proglukagon give rise to?
A: Proglukagon can give rise to different hormones with different effects

Q: Which signals stimulate glukagon synthesis and release?
A: - Epinephrine
- Amino acids

Q: Vem genomförde isolering av peptidhormoner från gristarm vid KI?
A: Viktor Mutt

Q: Where is glukagon synthesized?
A: Glucagon is a peptide hormone of the pancreas

Q: How are precursor hormones cleaved?
A: Introns are removed and exons are merged

Q: How does a hormone go from DNA to a hormone?
A: DNA -> messenger RNA -> preprohormone -> prohormone -> hormone

Q: What can proglukagon become?
A: - Glucagon
- Mini-glucagon
- GRPP
- MPGF

Q: What does a hormone cosist of in the mRNA state?
A: Signal(exon)-Hormone(exon)-Copeptides(exon)

Q: What does a hormone cosist of in the preprohormone state?
A: NH_2-signal-hormone-copeptide(s)

Q: What does a hormone cosist of in the prohormone state?
A: Hormone-copeptide(s)

Q: What does a hormone cosist of in the hormone state?
A: Hormone + Copeptide(s)

Q: What cannot be included in the secondary transport-category?
A: A transporter thant only transports one type of molecule can not be included in the secondary transport-category

Q: How can GLP-1/2 and glucagon work?
A: They work to some aspects synergistic and in others antagonistic

Q: How does the glucagon signal cascade look like - overview?
A: - Glucagon receptor (7-TM receptor, G-protein coupled receptor)
- Heterotrimeric G protein -> Adenylate cyclase
- cAMP
- PKA

Q: How can the glucagon activation process be described step-by-step?
A: Ligand+receptor -> activated receptor -> activated G-protein -> activated andenylate cyclase -> increased cAMP -> PKA -> activated effectors

Q: What is the point of the glucagon signal cascade?
A: To tell the liver to release glucose

Q: How can the 7-TM-/GPCRs be described?
A: 1. The extracellular domain contains the binding site for a ligand (a hormone or neurotransmitter)
2. Seven transmembrane helices
3. The intracellular domain that interacts with G-proteins

Q: What is the alfa-subunit in G-proteins?
A: Is a GTPase

Q: What are G-proteins?
A: GTP-dependent regulatory proteins

Q: What happens upon binding to a G-protein?
A: Upon binding the G-protein is activated in its GTP hydrolysis function
- GTP becomes GDP
- The receptor mediates the exchange of a GDP to a fresh GTP
- The beta- and gamma-subunits can release from the alfa-subunit so the alfa-subunit will be free

Q: Which function is activated upon binding to a G-protein?
A: Upon binding the G-protein is activated in its GTP hydrolysis function

Q: How many alfa-subunits are there in humans?
A: 15

Q: How manybeta-subunits are there in humans?
A: 5

Q: How many gamma-subunits are there in humans?
A: 10

Q: What do the different alfa-, beta- and gamma-subunits do?
A: They lead to many different adaptor modalities and downstream signaling outcome

Q: What can the beta- and gamma- G protein subunits do?
A: The beta and gamma can have a life on there own and do stuff/perform reactions

Q: What are the rules of beta- and gamma-subunits and what do they depend on?
A: There are rules of which beta and gamma dimerize and with alfa they bind to - depending on tragets downstream

Q: How can the glucagon signal cascade with heterotrimeric G-proteins be decsribed?
A: 1. Unoccupied receptor does not interact with G-proteins (G_s-protein with bound GDP)
2. Occupied receptor changes shape and interacts with alfa-subunit of G_s-protein. G_s-protein releases GDP and binds GTP
3. alfa-subunit of G_s-protein dissociates frombeta-gamma and activates andenylyl cyclase
4. When hormone is no longer present, the receptor reverts to its resting state. GTP on the alfa-subunit is hydrolyzed to GDP, alfa rejoins beta-gamma, and adenylyl cyclase is deactivated

Q: What does the amplification of andelyate cyclase give rise to?
A: One active AC can produce many cAMPs

Q: What's the 2nd messenger of andenylate cyclase?
A: cyclic-AMP

Q: What do cAMP bind to?
A: PKA

Q: What does PKA consist of?
A: 2 regulatory- and 2 catalytic subunits

Q: What type of activation occurs in PKA from cAMP binding?
A: Allosteric activation:
- cAMP binds to the 2 regulatory subunits -> conformational change -> releases the 2 catalytic -> they phosphorylate proteins

Q: Which amino acids are being phosphorylated by PKA in the glucagon signal cascade?
A: Serine and threonine

Q: What does PKA have downstream?
A: PKA itself have many different downstream targets - Activates and deactivates enzymes

Q: What is an example of covalent modification?
A: PKA -> phosphorylation

Q: What can the glucagon signal cascade turn on?
A: Gluconeogenesis enzymes

Q: What does glucagon do when fasting?
A: The glucagon signal cascade first activates glycogen release
- It can get ramped up by resynthesize of enzymes of glukoneogenesis

Q: How does the epinephrine signal cascade differ from that of glucagon?
A: It's a different GPCR but the same cascade

Q: How is glycogen breakdown stimulated in cells not expressing the glucagon receptor?
A: Muscle cells, adipose tissue, and many others do not express the glucagon receptor
- In the muscle, glycogen breakdown is stimulated by nor-/epinephrine instead

Q: Why are we not using glycogen stored in muscle when in a fasting state?
A: U are not using muscle glukogen coz that is destined for muscle activity

Q: When are we using mucsle-glycogen?
A: When one excretes adrenaline glycogen in muscles will be used for muscle activity

Q: Which type of receptor does epinephrine bind to?
A: Beta-Adrenergic receptor

Q: What happens with glucagon/glycogen breakdown when we start eating glucose again?
A: It needs to be switched off
- A protein coming arresting and blocking the receptors binding sites for G-proteins
- Proteins stimulate GTP hydrolysis, leading to it being hydrolyzed(beta-subunit can join)
- Glycogen synthase is phosphorylated and makes it inactive, dephosphorylated is active

Q: In which 3 steps can the glucagon signal cascade be shut off?
A: 1. Inactivation of GPCR
2. Inactivation of G-protein
3. Switch off glycogen breakdown

Q: Via what are there alternative GPCR pathways?
A: Via phospholipase-C/Diacylglycerol

Q: How can the alternative GPCR pathways be compared to the "normal"?
A: Same principles apply, but different second messengers used, different downstream kinase

Q: What is an example of signalling that occursvia enzyme-coupled receptors?
A: Insulin-signalling

Q: How can enzyme-couples receptors work?
A: Either the receptor is itself an enzyme (e.g. kinase) or associated with an enzyme

Q: What is an example of an enzyme where the receptor itself is the enzyme?
A: Kinase

Q: Via which receptor does insulin-signalling occur?
A: Receptor tyrosine kinases

Q: What are the 6 classes of enzyme-linked receptors tat have thus been identified?
A: 1. Receptor tyrosine kinases
2. Tyrosine-kinase-associated receptors
3. Receptorlike tyrosine phosphates
4. Receptor serine/threonine
5. Receptor guanylyl cyclases
6. Histidine-kinase-associated receptors

Q: What do receptor tyrosine kinases do?
A: Phosphorylate specific tyrosines on a small set of intracellular signalling proteins

Q: What do tyrosine-kinase-associated receptors do?
A: Associate with intracellular proteins that have tyrosine kinase activity

Q: What do receptorlike tyrosine phosphatases do?
A: Remove phosphate groups from tyrosines of specific intracellular signalling proteins

Q: What are receptorlike tyrosine phosphatases called receptorlike?
A: Because the presumptive ligands have not yet been identified, and so their receptor function has not been directly demonstrated

Q: What do receptor serine/threonine kinases do?
A: Phosphorylates specific serines or threonines on associated latent generegulatory proteins

Q: What do receptor guanylyl cyclases do?
A: Directly catalyze the production of cyclic GMP in the cytosol

Q: What do histidine-kinase-associated receptors do?
A: Activate a "2-component" signalling pathway in which the kinase phosphorylates itself on histidine and then immediately transfers the phosphate to a second intracellular signalling protein

Q: How does activation of receptor tyrosine kinases occur?
A: Activation typically by dimerization

Q: What plays role in crosstalk/integration of signaling cascades?
A: - Amplification of initial signal
- Dampening (have an OFF-switch)
- Integration of multiple signals