- heterotrimeric protein (alpha, beta gamma) - activated/deactivated by many GEFs/GAPs

Introduction to Signal Transduction Flashcards by Chandler Geary

How can signals be sent? (x4)

1) Endocrine: secreted into blood acting on distant target (hormones)
2) Paracrine ( acting on nearby cell, neurotransmitter)
3) autocrine ( secreted ligand acting on same cell (growth factor)
4) cell to cell signaling: plasma membrane attached proteins can signal to each other to affect dilation etc

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what is signal transduction?

process of converting extracellular signals into intracellular responses

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how do cells respond to signals? (2 ways)

1) fast: by changing activity of existing enzymes

2) slow: change levels of expression of enzymes vis gene regulation

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what does signal amplification mean?

-at each step of signal transduction pathway, the number of activated participants increases

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What does amplification cascade mean? Example of something that has it?

ex= hormone signaling pathways

- one epinephrine molecule leads to 1000s of cAMP final product. each step in process results in amplification

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rate of responses to extracellular signals?

1) rapid:(seconds-minutes) phosphoryaltion, ion/glucose/aa transport
2) delayed: (hours) trxn/translation
3) prolonged: (hours-days) requires trxn of cell cycle genes, cell growth/proliferation

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How are signals conveyed in the cell?

1) receptor mediated

2) non-receptor mediated

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what is receptor mediated signaling?

membrane receptors on cells respond to responsed to extracellular signals that bind to cell surface
allows cells to respond to hormones etc

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How do steroids signal?

-use nuclear receptors that are trxn factors capable of responding directly to steroid binding

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Non receptor mediated signaling?

cells sense contacts w/ the extracellular matrix or other cells by proteins etc
cells turn on internal signals in response to situations such as poorly folded proteins

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G-protein coupled receptors (GPCR)

largest protein superfamily
heptahelical (7 transmembrane)
couples to one or two of the 20 known heterotrimeric G-proteins at one time to activate alpha subunit

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What is a G-protein?

heterotrimeric protein (alpha, beta gamma)

- activated/deactivated by many GEFs/GAPs

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Alpha subunit of G-protein?

-GTP binding trimer with Beta and gamma

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GEF & GAP?

1) GEF (guanine nucleotide exchange factor) activate reactions by exchanged GDP–>GTP
2) GAP (GTPase activating protein) cause hydrolysis of GTP–>GDP therefore terminating signaling

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GPCR/G-protein steps?

GPCR bound by ligand, activated (G-protein bound GDP inactive state)
ligand binding/ conformational changes causes GDP–>GTP hydrolysis by GEFs activating G protein
G-protein alpha subunit binds GPCR, dissociation from beta gamma subunits
a. Alpha subunit & some pieces of beta gamma cause downstream signaling
Downstream signaling activates kinases which dephosphorylate GPCR causing it to dissociate from G-protein alpha subunit
RGS (regulator of G protein signals) are protein kinases that cause hydrolysis of GTPGDP therefore deactivating the G protein
a. GTP hydrolysis is the internal clock, controls G protein signaling
GPCR ONLY ACTIVE WHEN BOUND TO GPROTEIN no enzymatic activity on own

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G alpha protein subunit?

are G proteins
21 different versions encoded in human genome
activated by different GPCRs, regulate variety of diff effector proteins that synthesize second messengers

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cAMP and G alpha proteins?

cAMP is a second messenger
has stimulatory G alpha subunit activates it
an inhibitory Galphai inhibits production

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G(alpha)s

alpha–> adenylate cyclase (AC)–> cAMP–> protein kinase A (PKA)

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G(alpha)i

alpha inhibits AC-> lower cAMP production

-actiavtes RhoGEFs & K+ channels

G(Alph)q

1) couples to phospholipase CB (PLCB) to hydrolyze PIP2 & produce inositol triphosphate (IPS) and diacylglycerol (DAG)
2) IP3 mobilizes Ca2+
3) DAG and Ca2+ activate PKC

-can activate RhoGEfs & K+ channels

G(alpha) 12/13

-couples to certain RhoGEfs to activate RhoA, PlC

Typical G(alpha)s signaling?

1) binds AC which, converts ATP–>cAMP, cAMP activates PKA

2) can activate K+ & Ca2+ channels

How cAMP made/deactivated?

1) made by conversion of ATP–>cAMP via Adenylate Cyclase

2) degraded by PDE (phosphodiesterase)

Typical G(alpha)I signaling?

1) inhibits AC, therefore inhibits cAMP production & PKA production
2) beta & gamma subunits can activate RAS

Typical G(alpha)q signaling?

1) couples to PLC(beta) 2) hydrolyzes PIP2 3) PIP2 makes DAG + IP3 which move calcium 4) DAG & Ca2+ activate PKC

What are Receptor Tyrosine Kinases?

- growth factor receptors - 6 families - some have intrinsic kinase activity - some have kinase activity only after dimerization (growth factor families) - cell surface receptors w/ into and extra cellular domains

intrinsic kinase activity?

insulin family | -is dimeric

How activate Receptor TK?

1) ligand binds extracellular domain, conformational change(dimerization) or autophosphoryaltion if is an intrinsic tyrosine kinase 2) RTK-P Phosphorylated sites recruit binding/adapter proteins 3) Proteins activated: Protein kinases, lipid kinases, Monomeric G protein

kinase activity only after dimerization?

-growth factor families: EGFR, PDGFR, FGFR

Extra cellular domain structure vs intracellular domain RTK?

extra: similar to IgG, fibronectin or cysteine rich repeats intra: conserved w/ single tyrosine kinase domain or 2 tyrosine kinase repeats

Monomeric GTPases?

- Ras - molecular switches for cell division, growth, gene expression - are downstream of membrane receptors; bind & activate other proteins in signaling cascade

How are GEF's activated?

1) y phosphorylation | 2) active Tyrosine Kinase association

What is RAS-GEF called?

SOS

How monomeric GTPases (Ras) activated/deactivated? Where located?

- activated by exchange of GTP for GDP by GEFs - deactived by GTP hydrolysis (Gap) which acts as intrinsic timer 2)RAS attach to membrane via farnesyl & myristoylation

ERK1/2 Cascade? (5 steps)

1) RAS activated by GTP, farnesylation RAF brings RAF to active RAS membrane. Phosphorylate it 2) Active RAF (MAPKKK) looses auto inhibition 3) RAF phosphorylates MEK1/2 (MAPKK) activating it 4) MAPKK (MEK1/2) phosphyrlates & activates ERK1/2 5) ERK1/2 phosphorylated can enter nucleus and effect early trxn factors that promote cell proliferation

What does the ERk1/2 cascade link?

Links external cues from the RTKs to internal processes (Ras) down to cell proliferation

PI3Kinase?

- is a lipid kinase that promotes cell survival, migration & glucose uptake - alternative to RAS/RAF pathway

PI3 Kinase mechanism?

1) activated by RTK & GPCR 2) converts membrane lipids PIP2--> PIP3 3) PIP3 attracts proteins containing PH-domains

Downstream effects of PI3-Kinase?

1) Akt (protein kinase B) ^IMPORTANT 2)RhoA family GTPases 3) Glut 4

Examples of cellular oncogenes? How do they work?

1) src 2) Raf 3) Ras 4) Myc * induce proliferation - act on the mitogen activated protein kinase (MAPK) pathway

Akt?

- downstream effector of PI3 Kinase - is a protein kinase B (ser-thre kinase) - involved in mitogenic proliferation & survival

Jak/STAT pathway?

- JAK Family (Janus Kinase) = 3 JAK isoforms & tyrosine kinase 2 (TYK2) - JAK inhibitors used to treat autoimmune diseases

Jak/stat pathway?

1) interferon ligand binds receptor, cytosolic cross phosphorylation of receptor 2) activates JAK, which phosphorylates tyrosine kinase 3) binding proteins activated by JAK phosphorylation become TF (Stats_ enter nucleus and effect immunity/inflammation gene

Non-receptor Tyrosine Kinases?

- some tyrosine kinases don't have extracellular/membrane domains - post translation modification allow them to attach to membrane - unlike receptor TKs, intracellular domains are highly variable - src=example

src?

1) first non-recepeptor tyrosine kinases 2) membrane localization =myristoylation 3) regulation= autoinhibition that suppresses kinase domain 4) phosphorylates: EGFR & cytosolic proteins 5) function: trxn cell cycle genes, proliferation, cell rounding OVEREXPRESSED IN CANCER