TOPIC B CELL TALK: Communication networks inside and outside cell Flashcards
What is paracrine signalling?
- Local signalling
What is endocrine signalling?
- Long distance–> hormone related
- Released from tissue that is FAR from target cell
What are the three different types of effector proteinsproduced from signalling pathways and what are their actions?
- Metabolic enzyme (altered metabolism)
- Cytoskeletal protein (altered shape or movement)
- Transcriptional regulator( altered gene expression)
What are the two features of signalling?
- SPECIFICTY `
2. AMPLIFICATION
What is specificity?
- Signal molecule that fits binding site on complementary receptor (others don’t fit)
What is amplification?
- Enzymes activate OTHER enzymes
- no. of affected molecules INCREASES geometrically in enzyme cascade
What occurs in negative feedback?
- Build up of product turns off original activator
What is positive feedback?
- build up of product amplifies pathway
What type of signal transduction is FAST acting?
- Altered protein function (proteins already synthesised and ready to go)
e. g. insulin INCREASING GLUT receptors in membrane
Which type of signal transduction is SLOW acting?
- Altered protein synthesis (and expression)
e. g. If cell wants to synthesise more receptors
What are the three different classes of hormone?
- Polypeptide/protein hormones e.g. Insulin
- Amine –> Adrenaline
- Steroid–> Estrogens/testosterone (passes through cell membrane freely)
What 4 things is cell communication required for?
- Regulation of development
- Organization into tissues
- Control of growth, death, and division
- Coordination of diverse cellular activities
What is the biggest class of hormones?
- Polypeptide/protein
e.g. insulin/glucagon
leptin, GH, TRH LH Growth factors
Why are polypeptide/portein hormones stored in secretory vesicles for up to 1 day vand what form are they in?
- Because otherwise it could be degraded
- Stored as a PROHORMONE (inactive form)
What is the lifetime of protein hormones circulating in the blood freely?
- Minutes before they are degraded
What is the secretion of polypeptide/protein hormones regulated by?
- Other hormones
- metabolites
- CNS
What is the formation of mature insulin from immature insulin?
- Preproinsulin—(signal sequence chopped off)—-> Proinsulin—(c peptide chopped off)—-> Mature insulin
What chains does insulin have and what are they joined together by?
- Alpha and beta chain
- Joined together by disulfide bonds
What is insulin synthesised by?
- The beta cells of pancreas as PROHORMONE
What are peptide.amine hormones derived from and what are they ?
- Tyrosine
- Adrenaline and NA
- T3 and T4
Where are NA and A secreted from?
- Adrenal medulla
If NA and A are secreted freely in the blood, how long will they last for?
- Seconds (so short lifetime)
What are the 3 different types of cell surface receptors?
- Ion-channel coupled receptors
- G-protein coupled receptors (activated by GTP binding)
- Enzyme coupled receptors (variable) –> Receptors itself is enzyme
- can have catalytic domains OR associated with enzyme close to membrane
Where does the signalling molecule bind to in nuclear receptors?
- Ligand binding domain (distinct from other domains)
In nuclear receptor pathways, what MAY NEED TO occur before gene transcription can happen?
-may need to have coavticator proteins binding (can become part of transcription activating domain)
What happens in nuclear receptor pathways when the ligand binds?
- there is conformational change q
What other important domain is part of the intracellular receptor?
- DNA-binding domain
What are the two pathways that can be active once G protein coupled receptor is activated?
`1. Cyclic AMP pathway (second messengers)
2. Inositol phospholipid pathway
What features are common to all G protein coupled receptors? -
- Cell surface receptors
- 7 transmembrane segments
- signalling molecule can activate MULTIPLE GcPRS
- ALL use G proteins to relay a signal
What is the structure of a G protein? (and subunits)
- Trimeric G protein complex
- Has alpha, beta and gamma subunits - in some types may only have 1 or 3 subunits
What can the alpha subunit bind on a G protein?
- GDP (inactive form)
- GRP active form
What do the phospholipid tails on the alpha and beta subunits of the G prtoein do?
- Anchor subunits into membrane
What are the 7 steps of the activation of a G protein? (Last step: when is switch turned off?)
- Ligand binds to G protein coupled receptor
- RECEPTOR changes conformation, interacts with inactive GDP bound G protein
- Causes G protein to EJECT GDP and replace with GTP
- This exchange causes conformational change
- G protein: binding GTP releases receptor
- It interacts with one or more effectors —> active protein–> delivers message
- Switch is turned off WHEN G PROTEIN HYDROLYSES its own bound GTP–> GDP (after time)
What happens in G protein coupled receptors when GTP is bound to the alpha unit?
- It DISSOCIATES from Beta or gamma units so they can activate another complex/pathway)
What is the role of a second messenger?
- Internal messenger that activates secondary pathways in cell e.g. cAMP–> activates PKA and self transcription
What occurs in the universal (not individual cell type) Inostiol Phospholipid pathway?
Signal molecules comes in from EXTRACELLULAR source and activates receptor–> induces conformational change–> activates G protein complex (beta and gamma subunits)—> Activates protein (phospholipase C–this cleaves phospholipid)—> Activated phospholipase C acts on INOSITOL PHOSPHOLIPID (IP2) in membrane and cleaves it from DAG (diacylglycerol) in membranre—> this FORMS IP3 (secondary messenger)
- IP3 binds to ligand gated ion channel–> Casues Ca2+ efflux from ER —> Ca2+ activates PROTEIN KINASE C (PKC) —> then activates other processes (cellular) like gene trancription and muscular contraction
What does PKC (protein kinase C) need to be activated in the insosital phospholipid pahtway?
Needs BOTH DAG (diacylglycerol) and and Ca2+ to be activated
What occurs in the cyclic AMP pathway of G protein coupled receptors?
Membrane bound recpetor: Adenylyl Cyclase converts ATP–> cyclic AMP
- Only alpha subunit involved
- cAMP then activated in signalling molecule in cell (second messenger)
- Inactive PKA (4 subunits–2 catalytic and 2 regulatory)
- cAMP binds to regulatory subunits of PKA—> these regulatory subunits then released from catalytic domains —> activated PKA
What is an example of the cAMP pathway and what occurs?
ADRENALINE INDUCING GLYCOGEN BREAKDOWN
- Adrenaline binds and activated GPCR (sdrenergic) activates activated adenylyl cyclase and alpha subunit of protein
- ATP —> cAMP –> active PKA –> activates enzymes specific to glycogen breakdown
- then via ATP it forms active glycogen phosphorylase
In activation of gene trasncription via adrenaline through cAMP pathway, what is the pathway from active PKA?
- Active PKA moves into nucleus (via pore) and binds to transcription regulaotrs–> leads to gene trasncription
Does active PKA have different cellular responsed based on cell type?
- YES!!
What are G protein coupled receptors as drug targets?-
Adrenergic receptors (beta blockers) –> to decrease HR and contractility)
What is a treatment of cardiac aryhtmia and angina?
- Inhibiting Beta adrengergic recpetor (GPCR) action
What does the drug inderal (proprananol) do?
- Acts as a Beta 1 agonist
- competitively INHIBITS GPCR binding to NA–> slows heart contraction
What leads to cortisol secretion and what type of receptor pathay does this use?
ACTH
- ACTH is a GPCR
What is cushing syndromec and what is it associated with?
- Chronic exposure to EXCESS glucocorticoids (steroids)
- decreased growth in kids
- Increased weight, hpertension, diabetes
- osteoperosis, muscle weakness
What are potential causes in cushings syndrome?
- Genetic Mutations in signalling pathway
What can lead to changes in the level of cortisol secreted?
- Mutations in any step of the pathway (cAMP signalling)
- Pathway is overactive (cancer also has this overactive effect)
- GPCR mutation
What are the two types of activating mutations that occur in cushings syndrome?
- Loss of function mutation (turns off protein) and gain of function (protein may be overactive)
What occurs in mutation in regulatory protein?-
- May not be able to turn off signalling pathway
What occurs with a mutation in an enzyme that needs to be activated
Pathway may be blocked
What are the key molecules in the phosphatidylnosol pathway?
- GPCR–> G protein–> inositol phospholipid –> inositol 1,4,5 triphosphate–> PKC
What are the key signalling molecules in the cAMP pathway?
- GPCR activation –>G protein activation–> Adenylate cyclase activation–> ATP–> cAMP–> PKA activation
Which two factors are responsible for directly activating Protein Kinase C (PKC)?
- Diacylglycerol (DAG) and Ca2+
What activates protein kinase A (PKA)
- Cyclic AMP (cAMP)
What does a loss of function mutation lead to?
- Leads to INHIBITION of signalling pathway (symptoms like hormone deficiency)
What does a gain of function mutation lead to?
Overactive pathway (symptoms similar to excess hormone or signalling molecule)
Where can a loss or gain of function mutation occur?
- ANY of the proteins involved in in GPCR signalling OR second messenger pathways
What is cushings syndrome a result of ?
INCREASED cortisol secretion (adrenal cortex) OVERTIME
How is cortisol normally produced and secreted?
- CRH (corticotropin releasing hormone) in hypothalamus goes into hypophyseal system into ANTERIOR pituitary and acts on a GPCR to allow ACTH to be produced –> released into blood stream –> adrenal gland ACTH stimulates adrenal cortex cells to release and produce CORTISOL!
What type of mutation occurs in cushings syndrome and which parts can be affected?
- Gain of function mutation
- overactive pathway
- Alpha subunit of trimeric G protein
- PKA
- GCPR
What are two gene mutation that lead to overactive /overexpressed receptor or G proteins or the catalytic subunit of PKA?
- Primary Bilateral Macronodular Adrenal Hyperplasia (PBMAH)
- Adrenal adenoma (tumors on adrenals gland)
What are the gene mutations associated with primary pigmented Nodular Adrenocortical Disease (PPNAD)?
-Mutations in the catalytic or regulatory sub-units of PKA
Which type of mutation can central hypothyroidism be and what does it lead to?
- Loss of function mutation
- mutation in Thyrotropin Releasing Hormone Receptor (TRHR) in anterior pituitary
- Mutation causes inositol pathway to be INHIBITED
- so thyrotropic cells can’t be stimulated from the TRH released from hypothalamus
- SO NO production of TSH and no secretion of thyroid hormones
What is the correct order for the release of thyroid hormones?
- TRH released from HYPOTHALAMUS
- TSH released from anterior pituitary
- T 3 and T4 released from thyroid
A loss of function mutation in cAMP PDE would result in?
- Increased cAMP signalling due to inability to regulate duration of cAMP activation
What does GEF (guanine nucleotide exchange factor) do?
- Activates GTPase
- Stimulates the release of GDP
- GTP quickly binds–> GTPase activated–> GTP exchanged for GDP
What does GAP do?
- Gtpase Activsting Protein
- hydrolyses GTP–> GDP
- GDP remains tightly bound
Which 3 are the most commonly phosphorylated as part of the enzyme coupled receptpr pathways?
- Serine
- Threonine
- tyrosine
Which part of the RTK is the part that is catalytic and can phosphorylate other molecules when activated?
- Tyrosine kianse domain
What does the insulin rercepotr exist in membranes as?
Dimeric form
In general, what to RTK’s lead to when activated?
- cell growth
- cell division
- proliferation/cell survival
What do RTK’s prevent/inhibit?
- Apoptosis
How do receptor tyrosine kinases work?
- Receptor in membrane is monomer
- Ligand binds—> forms DIMER with a receptor of SAME family –> they can activate EACH OTHER (autophosphorylation) –> both receptors are activated
- Then receptors can activate multiple pathways in the cell
What do the phosphorylation points in the tail of the receptor (RTK pathway) provide?
- Binding sites for different adapter molecules
Which domains do proteins that are binding to ECF receptors have to have?
- SH2
SH3
PTB
PH
What does SH2 recognise?
- Phosphorylates tyrosine residues
What does SH3 do?
- Can bind to proline rich regions
What does PTB recognise?
- (Phospho Tyrosine Binding domain)
- Recognises Phosphorylated tyrosine residues
What does PH recognise?
- Hyperphosphorylated inositide molecules
What are adaptor proteins?
- Act like bridges from one protein kinase to another
- can form a bridge between molecules that don’t have their own SH2 or PTB domains e.g. Grb2
What are scaffold proteins and what do they allow/reduce?
- bring together multiple interacting signals/proteins so they can activate each other
- CAUSES RAPID ACTIVATION
- REDUCES other signal molecules from other pathways from interfering
In cancer, what part of the RTK are looked at?
- The phosphorylation sites
- Usually in cancer the phosphorylation sites can be overactive
What does PI3K do and what is it activated by?
- Recruited and activated by RTKs
- Phosphorylates inositol phospholipids in plasma membrane
- Becomes the docking site for intracellular signalling proteins (Akt) and activates them
What is Akt also known as and what does it do?
- PKB (protein kinase B)
- Promotes growth and survival (inhibits BAD–> BAD encourages apoptosis)
What is BcI2? (pro or anti apoptotic)
- Pro apaptotic
What is Bad? (pro or anti apaptotic)
- anti apoptotic
What occurs in terms of Bad and BcI2 when Akt (PKB) is active ?
- It phsophorylates Bad
- Bad and BcI2 dissociate
- So BcI2 is CTIVATED AND APOPTOSIS IS INHIBITED (SURVIVAL SIGNALS)
- So Phosphorylation INIHBITS Bad
What do tyrosine associated kinase receptors associate with?
- Proteins that have tyrosine kinase activity (not the tyrosine kinases)
- e.g. CYTOKINE RECEPTORS
Which two families of proteins are important for tyrosine associated receptors?-
- Src (SARC) kinase family (associate with receptors)
- Janus Kinase family e.g. JAK 1, 2, TYK 2
(signalling family receptors–> growth hormone receptors
How are cytokine receptors formed?
- Pre assembled with JAK protein on tail (has kinase function)
- JAKs are Phosphoryltors –> they autophospohrylate each other –> JAKS phosphorylate cytokine receptor
- STAT 1 and 2 recruited for transcription
What do epidermal growth factors activate?
- Receptor Tyrosine Kinases
Which protein domains bind phosphorylated tyrosine residues?
- PTB and SH2 (depends on individual affinity for which one actually binds)
Janus Kinases activates which family of proteins?
- STATs (src is also important)
What do nuclear receptors allow for?
- Transcription to be regulated!
- Reproduction, homeostasis, integrated metabolism
What is the biggest class of hormones that bind to nuclear receptors?
- Steroids
but also vitamin D and thyroid-less large
What type of receptor does cortisol use?
- A nuclear receptor becasue it is lipophillic
What receptor can be targeted in cushings syndrome?
- Glucocorticoid receptor (GR)
What does the drug Mifepristore (RU486) do?
- It is a GR and PR (progesterone receptor) antagonist (prevents signalling and activation)
- Active in abortion
- trialled in depression
Where are steroid hormoens synthesised?
- Adrenal, Overies and testes
What can congenital adrenal hyperplasia result from?
- Defficiencies in 1-hydroxylase
- 11 beta hydroxylase
- or 17 alpha hydroxylase or C17-20 lyase
What are DNA binding sites called?
- Hormone Response Elements (HREs)
Steroid hormones bind to HREs as…..
HOMO DIMERS (GR, ER, PR, AR)
RXR-NRs (Retinoid X receptors) bind to HREs as:
- Heterodimers (e.g. RAR, PPAR )
What do the estorgen receptors exist as?
- Homodiemrs
- ER alhpa and ER beta receptors form the homodimer
What is ERalpha important for?
- Mediating estradiol effect (femaile reporoduction ,
mammary gland function, bone, CV system, brain
What is ER beta im,portant for?
- OVARY, bone, breast, brain ,
- different expression patterns
What occurs in a ER alpha deficiency?
- Resistant to estorgens (high estrogen blood levels)
- Continued growth
- Estrogen important for male and femalse bone formation
What is CML and which pathway is an issue here?
- Chronic Myelopid Leukemia (excess white blood cells)
- overflow of immature leukocytes into bloodstream
- OVERACTIVE INTRACELLULAR SIGNALLING PATHWAY (cell growth and survival)
- oncogenic tyrosine kinase (BCR-ABL)
What is the action of BCR-ABL in the overactive signalling pathway of CML?
- This oncogenic tyrosine kinase phosphorylates kinases in the cell proliferation pathway (Ras/MAPk, PI3K/Akt, JAK/STAT)
- This leads to uncontrolled cell growth
What is a possible treatment of CML in terms of the oncogene BCR-ABL?
- Gleevic (inhibitor)
What is the action of Gleevic and what is it used as
?
- binds to BCR-ABL binding site (so ATP can’t bind) and so no phosphorylation
- Used as a possible treatment for CML
What is the action of TKIs (Tyrosine kinase inhibitors) ?
- Competitive inhibition of ATP and upstream or downstream substrate
How do mAbs (monoclonal antibodies) work (in context of CML treatment)
- They bind competitively with receptors ligand binding site (so can’t be activated or dimerise) (inhibit cell surface receptors)
What is HER2, which family is it from, which protooncogene is it encoded by and what is a special feature of it?
- Human Epidermal Growth Factor receptor 2
- From the EGFR family
- Encoded by proto oncogene ERBB2
- Special feature: Can form heterodimers with other EGFR family members
- Associated with breast cancer (aggressive)
What does the HER2/EGFR do when active?
- Activates the MAPK and PI3K pathways
- these pathways lead to cell growth and proliferation
What is Heceptin and what action does it have?
- Monoclonal antibody
- Specific to HER2
- binds to HER2 so it can’t activate the intracellular signalling pathway
- Is a treatment for HER2 positive breast cancer
