W7L3

Question 1

Cells recognize and respond to
environmental cues via receptors

Answer 1

Cells communicate and
coordinate activities by
sending and receiving
signals

▪ Each cell expresses a
variety of receptor
proteins that allow it to
respond to particular
signals.

▪ Signals originate from:
 other cells
 environment

Question 2

Environmental signals

Answer 2

1. Sensory signalling
   Detection of external signals by sensory
   receptors for
   - Light (rhodopsins)
   - Smell (olfactory GPCRs)
   - Taste (GPCRs, ion channels, transporters)
   - Sound, balance and touch
   (mechanoreceptors)
2. Cellular environment
   Cells can detect
   - Nutrients
   - Protons, pH
   - Nucleic acids
   - Osmotic pressure (hypertonic or hypotonic solution)
   - Fluid sheer stress
   - Xenobiotics (chemicals from outside of the body, such as drugs and toxins)

Question 3

Endogenous Extracellular Signalling Molecules

Answer 3

1. Secreted from the signalling cell into the extracellular space
   - e.g. hormones, neurotransmitters
2. Released by passive diffusion through channels in PM
   - e.g. ATP, prostaglandins
3. Liberation of PM-embedded ligands by matrix metalloproteinases
   - e.g. growth factors, cytokines
4. Exposed to the extracellular space, but remain tightly bound to the surface of the signaling cell
   - is a transmembrane protein, can bind to receptors on other cells
   - e.g. ephrins
5. Extracellular matrix proteins
   - e.g. integrins

Question 4

Modes of intercellular signalling

Answer 4

1. Endocrine signalling
2. Paracrine signalling
3. Synaptic signalling (a specialized form of paracrine signalling)
4. Contact-dependent signalling (aka juxtacrine signalling)
5. Autocrine signalling
6. Protease-dependent signalling

Question 5

Endocrine signalling

Answer 5

Endocrine cells release hormones into the bloodstream or lymphatic system - transported throughout body (signalling over long distances)

Hormones act on target cells that express a receptor for the ligand

Examples
- Insulin released from the pancreas promotes glucose uptake throughout the body
- Parathyroid hormone (PTH) is secreted from the parathyroid gland and acts on PTH receptors in bone, kidney, and intestine

Hormone signalling is relatively slow compared to other modes

Question 6

Paracrine signalling

Answer 6

Para = near

Secreted signalling molecules act as local mediators that impact target cells in the immediate environment

Examples
- platelet-derived growth factor (PDGF)
— wound healing
- locally released parathyroid hormone
related peptide (PTHRP) activates PTH1
receptors in bone and regulates
mineralization
— PTH (parathyroid hormone) is a hormone; PTHRP is a paracrine factor

Question 7

Synaptic signalling (a specialized form of paracrine signalling)

Answer 7

Regulates neuronal communication

Allows for adaptive changes required for behavioural responses or reflexes

• neurons (nerve cells) extend long axons
• axons contact each other at synapses
• neurons can also form synapses with
  muscle cells
• rapid mode of signalling
• typically stored in intracellular vesicles
  and released into synapses

neurotransmitters include acetylcholine,
catecholamines (dopamine, adrenaline,
noradrenaline), serotonin, endorphins,
GABA, endocannabinoids, corticotropin
releasing factor…

• Can be normal synaptic signalling (presynaptic to postsynaptic)
• Backwards signalling is less common (postsynaptic to presynaptic signal, i.e. cannabinoids)

Question 8

Contact-dependent signalling (aka juxtacrine signalling)

Answer 8

Physical contact between adjacent cells (or between cell and ECM)

“Ligand” protruding from one cell activates receptor in plasma membrane of neighbouring cell
- ephrin receptors
- adhesion GPCRs

Signalling molecules remain bound to surface of signalling cell

Activate target cells that come into contact with signalling cell

Important in developmental biology

Question 9

Autocrine signalling

Answer 9

Auto = self

Cell releases a signalling molecule that acts back upon receptors expressed on its own surface to control its activity

Example:
- Cytokine interleukin-1 in immune cells
- Vascular endothelial growth factor (VEGF) in cancer cells
- Neurotransmitters (e.g., noradrenaline, 5HT) which bind to presynaptic autoreceptors

If you block autoreceptors, this will increase the amount of nt

Question 10

Protease-dependent signalling

Answer 10

Proteolytic activation of precursors

Examples:

• Proteinase-activated receptors: tethered ligand on GPCR aminoterminus is exposed by proteolysis
• Release of membrane-anchored ligand from extracellular surface by metalloproteinase activity
• Conversion of pro-hormone to active form by proteinase activity intracellularly (e.g., endorphins) or
  extracellularly (e.g., angiotensin II)

Question 11

Endogenous ligands can play multiple
signaling roles

Answer 11

Some endogenous ligands can play multiple
signaling roles

i.e. noradrenaline released from adrenal glands travel in blood system and can act as a hormone, a neurotransmitter in the synapse or an autocrine factor by activating pre synaptic receptors

From the perspective of the receiving cell,
what’s the difference between hormonal,
paracrine and autocrine signaling?

Question 12

How do cells recognize and respond to signals
in their environment?

Answer 12

Cells surface receptors transduce signals into cells

Make a Ligand– receptor complex

Transduce: to convert (something, such as
energy or a message) into another form
–> signal transduction

Question 13

Affinity

Answer 13

how tightly a drug binds to its receptor

K_D = Koff / Kon

how fast does it go in and come out of the binding site

something that goes in quickly and comes out slowly has high affinity. The lower the K_D, the higher the affinity

Question 14

Specificity/selectivity

Answer 14

a receptor binds preferentially to ligands that fit well
into its binding site

Question 15

Saturability

Answer 15

ligand binding is limited by the number of receptors

Question 16

Reversibility

Answer 16

the ability to bind to and dissociate from a receptor. Most ligands bind reversibly

Question 17

Competition

Answer 17

a binding site can only accommodate one ligand at a time. If two or more ligands are present they will compete with each other. i.e. agonist vs drug

Question 18

Agonist

Answer 18

an endogenous ligand or drug that binds to a receptor and promotes activation

Question 19

Antagonist

Answer 19

a ligand that binds to a receptor but does not activate it

Question 20

Cellular determinants of receptor signaling

Answer 20

The ways in which different cells responds to a particular agonist may vary:

1.Receptor expression level
- availability of receptors will determine cellular response
- cells that lack receptors for a particular signal cannot respond
- cells with greater receptor density may respond at lower agonist
concentrations and/or activate minor pathways

2. Receptor variants
   - Ligand may binding to distinct receptor subtypes in different cells
3. Intracellular signaling components
   - same receptor may activate different pathways in different cells
   - depends on intracellular factors available to integrate and interpret
   receptor signal
   - e.g. complement of kinases, ion channels, phosphatases, substrates

Question 21

Signal turnoff

Answer 21

Endogenous agonists may be
deactivated metabolically
- hydrolysis of acetylcholine by
cholinesterases
- breakdown of peptide and protein
hormones by proteases

Endogenous agonists may be
reabsorbed from extracellular space via
specialized uptake proteins (e.g.,
transporters for dopamine, adrenaline,
noradrenaline, serotonin,
prostaglandins)

Drugs that mimic endogenous agonists can be metabolized by liver enzymes and/or excreted, e.g., via the urine

Receptor activation can also trigger intracellular processes that limit signaling, such as G protein uncoupling and receptor internalization

Question 22

Molecular Switches –GTP binding and hydrolysis

Answer 22

short term biochemical changes that a cell can undergo to adjust its activity

Heterotrimeric G proteins and small Ras-like G proteins
belong to a superfamily of GTPases where activation and deactivation correspond to GTP binding and hydrolysis, respectively.

For most G proteins, these changes occur on a time scale of seconds to minutes (i.e., slow relative to many
biochemical processes)

Question 23

Regulation of G protein activation state by other
proteins

Answer 23

▪ GEFs (guanine nucleotide exchange factors) promote
GDP dissociation and thus facilitate activation by GTP
- cells contain 10X amount of GTP as GDP

▪ GAPs (GTPase accelerating proteins) promote the
hydrolysis of GTP, thereby deactivating G proteins

▪ GDIs (guanine nucleotide dissociation inhibitors) inhibit GDP dissociation and thus impede activation

Question 24

Molecular switches - protein phosphorylation/dephosphorylation

Answer 24

Protein phosphorylation
* ubiquitous strategy
* reversible covalent modification,
with proteins dephosphorylated by
phosphatases
* kinases undergo conformational
changes in response to diverse
inputs, which then regulates kinase
catalytic function (phosphorylation)

ATP cleaved to ADP; phosphate
released covalently attached to a
protein
- Phosphorylation does not
always mean activation

Phosphorylation via kinases. Dephosphorylation via phosphatases

Question 25

General Characteristics of Cell Signalling

Answer 25

Exogenous or extracellular agonist can be called 1st or primary messengers
- they alter the intracellular concentrations of other signalling molecules called secondary messengers

Primary messengers
- Receptors bind agonists and regulate
intracellular signalling proceses
- Alternatively, receptors may be or
control ion channels, leading to
altered levels of intracellular ions

Secondary messenger is ion or molecule that is changed in the cell in response to primary messenger

2 nd messengers include
-Inositol trisphosphate (IP3)
-Diacylglycerol (DAG)
-Cyclic nucleotides (e.g. cAMP, cGMP)
-Calcium ions
— called secondary messengers, but calcium can either come into the cell directly via ion channels or can be released from intracellular stores in response to second messengers. Calcium is an intracellular signalling molecule of primary importance

Intracellular signalling proteins alter
activity of target proteins resulting in
changes of cell behaviour [response]

Question 26

Signaling
cascades

Answer 26

At each step in a signaling
cascade, the “product” of one
step becomes the activator or
substrate of the next.

Amplification may occur, e.g.,
if a kinase phosphorylates
multiple substrate molecules

Question 27

Receptor reserve (aka spare receptors)

Answer 27

Have multiple receptors being activated. But only 1 receptor produces enough product (or only 1 receptor is needed) to activate the subsequent molecules in the path.

What is the purpose of the other receptors? It is quite redundant. If there is too much upstream signalling or if it is excess (more than required for downstream signalling), then this is called spare receptors or receptor reserve.

Receptor reserve is important in potency
- if you have receptor reserve, you have more receptors than needed to activate the intracellular signalling system. This will impact the concentration dependence of the activating ligand.
- A smaller amount of activating ligand will be able to saturate the system if you have receptor reserve at the level of the plasma membrane

Question 28

The strength of a receptor signal will depend on…

Answer 28

1. Amplification, e.g., via activities of
   - downstream kinases
   - downstream enzymes
2. Attenuation
   - Phosphatases, counter-movement of ions, 2nd messenger breakdown
   - GTPase activating proteins to deactivate G-proteins that have been activated by receptors
3. Availability of scaffolding proteins, e.g., AKAPs, PDZ proteins
   - Bring together components of signaling cascades into close proximity; increases efficiency of signalling
   - Increased local concentrations of soluble factors
   - Temporal focusing of the signal via negative regulators
4. Tachyphylaxis/desensitization (cell can limit signalling)
   - Acute – ion channel conformation, receptor phosphorylation
   - Substrate depletion
   - Receptor internalization (take away receptor so it cannot reach ligands that will activate it) , degradation (increase this rate)
   - Receptor mRNA downregulation