signal transduction second lecture done by: ahmad … · 2020-01-22 · signal transduction second...
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SIGNAL TRANSDUCTION SECOND LECTURE
DONE BY: AHMAD ALSAHELE
*ما لون باألسود فهو جزء من الساليدات
*ما لون باألزرق فهو كالم الدكتور
*ما لون باألحمر فهو شرح اضافي
Cell-to-Cell Interactions
-Cells can identify each other by cell surface markers.
-Glycolipids are commonly used as tissue-specific markers
-Major histocompatibility complex (MHC) proteins are used by cells to distinguish "self"
from "non-self"
تم التفصيل عنهما مستقبالً ولهذين البروتينين دور مهم في المناعة وسي MHC-2و MHC-1يوجد منه نوعان MHC*ال
. immunologyفي ال
لخاليا الجسم الواحد ويستطيع الجسم من خاللها تمييز خاصة markersذه البروتينات تعتبر : ه 3*شرح العبارة
األجسام الطبيعية من الغير طبيعية حيث ترتبط هذه البروتينات باألجسام الغير طبيعية وتبرزها للخاليا المناعية ليتم
التخلص منها.
........................................................
Cells within a tissue are connected to each other by cell junctions
1. Tight Junctions – create sheets of cells (they make cells very close together )
2. Anchoring Junctions – connect the cytoskeletons of adjacent cells
3. Communicating Junctions – permit small molecules to pass between cells
a. gap junctions – in animal cells
b. plasmodesmata – in plant cells
........................................................
*the function of
tight junctions is to
make the cells close
together as possible
*anchoring
junctions are
attachment points
for the cytoskeleton
*permit small
molecules to pass
between cells
this projections are called microvilli
they exist in intestinal mucosa
Acetylcholine: common neurotransmitter
بواسطة االنزيم degradationعملية acetylcholine*لما يصير لل
acetylcholinesterase يعود الcholine الى الpresynaptic cell
. reuptake mechanismبعملية تسمى
the common function of
Ach:
opens ligand-gated Na+
channels on muscle cell
and some nerve cells
........................................................
*acetylcholine neurotransmitter act as ligand allows Na+ to pass inner structure of the cell
by change conformation of ligand-gated Na+ channels.
Signal transmitted to muscle cell across a synapse
• the whole mechanism of signal transmission as the
lecturer say in the lecture:
o acetylcholine permit transport of Na+ which
make action potential in postsynaptic
membrane
o action potential move across the axon as
electrical impulse until it reach presynaptic
membrane so that it depolarize
o a.Depolarization opens voltage-gated Ca+2
channels
o b.Ca+2 rushes in; Vesicles fuse with membrane
o c.Neurotransmitter(which is reserved in
presynaptic junction) released; opens ligand-
gated Na+ channels on muscle cell Depolarizes
muscle cell
action potentialوانتهت ب action potential*مما سبق نالحظ أن الرحلة بدأت ب
(action potential=>electrical signal)
Signal: electrical to chemical to electrical
acetate choline
a
c
b
GPCRs that Regulate Ion Channels: Muscarinic Acetylcholine Receptor
*Ach work in striated muscles (nicotinic receptor) & in cardiac muscles (muscarnic
receptors)
The neurotransmitter, acetylcholine (ACH) binds to two types of receptors known as the
nicotinic and muscarinic acetylcholine receptors.
The nicotinic receptor is itself a ligand-gated ion channel that opens on ACH binding. This
receptor is located in the neuromuscular junctions of striated muscle.
بتسمح أليونات Achبس يرتبط فيها ال بوابةعبارة عن nicotinic receptor*في هاي النقطة الزم تعرف انه ال
contractionوفي نهاية المطاف يحصل action potentialومن ثم polarizationالصوديوم انها تدخل وتعمل
للعضلة وتتم الحركة المطلوبة
The muscarinic ACH receptor, is a GPCR found in cardiac muscle cells that is coupled to an
inhibitory G protein
( أي أن سرعة نبض القلب تقل عندما يتم تحفيز inhabitoryمهمة تثبيطية)muscarinic ACH receptor *مهمة ال
. G proteinمستقبالت مرتبطة بهذه المستقبالت التي هي
........................................................
The binding of ACH to this
receptor(muscarinic) triggers dissociation
of Gai-GTP from Gßg, which in this case,
directly binds to and opens a K+ channel.
The movement of K+ down its
concentration gradient to the outside of
the cell, increases [K+] outside which
increases the positive charge outside the
membrane, hyperpolarizing(increase
electrical gradient) the cell. This results in
the slowing of heart rate.
*the main function of Gi protein is : increase K+ concentration out of the cell.
*there are different types of G proteins the most important is: Gi / Gs / G0 / Gq .
*[K+] is more important in heart ,[Na+] is more important in striated muscles.
*if hyperkalemia or hypokalemia happen many problems attack the heart especially in
ECG(a test that checks for problems with the electrical activity of your heart) & intensity of the heart
that can lead to heart attack because of little change in K+ concentration.
*any change(fluctuation) in [K+] may be severe because in the serum and blood the [K+] is
has a small limit(3.5 - 5.5) when we compare with [Na+] (135 - 145)
Structure of GPCRs
G protein-coupled receptors (GPCRs) are the most numerous class of receptors in most
eukaryotes. Receptor activation by ligand binding activates an associated trimeric G
protein, which in turn interacts with downstream signal transduction proteins. All GPCRs
are integral membrane proteins that have a common 7 transmembrane segment
structure (Fig.( The hormone/ligand binding domain is formed by amino acids located on
the external side of the membrane and/or membrane interior (Fig.). GPCRs interact with
G proteins via amino acids in the C3 and C4 cytoplasmic regions.
........................................................
*7 membrane segment protein is
represent 1 receptor only.
*transmembrane coupled receptor
segments are attached to each other by
exterior units(E1,E2,E3,E4) and
cytoplasmic units(C1,C2,C3,C4).
*the most important cytoplasmic units are
C3 C4 because they will interact with G
protein.
G Protein Activation of Effectors
The trimeric G protein cycle of activity in hormone-stimulated GPCR regulation of effector
proteins is summarized in (next slide). Initially, the G protein complex is a chain to the
inner leaflet of the cytoplasmic membrane via lipid anchors attached to the Ga and Gg
subunits. The trimeric GDP-bound form of the G protein is inactive in signaling. The
binding of a hormone to the GPCR triggers a conformational change in the receptor (Step
1) which promotes its binding to the trimeric G protein (Step 2). Binding to the activated
GPCR triggers the dissociation of GDP (Step 3). Subsequent binding of GTP to the Ga
subunit activates it, and causes its dissociation from the receptor and the Gßg complex
(Step 4). Ga-GTP then binds to the effector protein regulating its activity. The hormone
eventually dissociates from the receptor (Step 5). Over time (often less than 1 min), GTP
is hydrolyzed to GDP and Ga becomes inactive. It then dissociates from the effector and
outside
inside
recombines with Gßg (Step 6). A hormone-bound GPCR activates multiple G proteins, until
the hormone dissociates. Proteins known as regulators of G protein signaling (RGS)
accelerate GTP hydrolysis by Ga decreasing the time-period during which Ga is active (not
shown).
........................................................
*ملخص هذه الصورة:
في مما يؤثر receptorبال hormoneيتصل ال
الذي يعد G proteinعلى ال بنيته ويؤثر
(trimeric protein composed of a,b,g)
gو bوهي اكبر من GDPعلى aحيث تحتوي
يحصل receptorبال hormoneعندما يتحد ال
activation ه فيتصل باللG alpha subunit
حول conformational changeويحصل لها
فتتم عملية freeوتصبح GDPال
GTPلتصبح GDPلل phosphrylationال
وتنفصل alpha subunit =< activeفتصبح
أو GDPمرة اخرى GTPيصبح الوعندما activeجاعلة اياه effectorوتتصل مع ال beta & gammaعن
واذا عاد G protein =< trimericيعود كل شيء كما كان وبرجع ال receptorعن ال hormoneينفصل ال
جديدة. cycleتبدأ hormoneال
Trimeric G Proteins & Their Effectors
There are 21 different Ga proteins encoded in the human genome. The G proteins
containing these subunits are activated by different GPCRs and regulate a variety of
different effector proteins . The most common effectors synthesize second messengers
such as cAMP, IP3, DAG, and cGMP. In the case of cAMP, a stimulatory Gs subunit
activates adenylyl cyclase and cAMP production, whereas an inhibitory Gi subunit
inhibits adenylyl cyclase and cAMP production.
*there is another second messengers differ from the mentioned but the mentioned are the
most important.
GPCRs That Bind Epinephrine
Epinephrine is a hormone that signals the "fight-or-flight" response ( اي قاتل او اهرب وفي كال
,It elevates heart rate, cardiac output, blood supply .(الحالتين ستحصل نفس التغييرات للجسم
dilates the airway, and mobilizes carbohydrate and lipid stores of energy in liver and
adipose tissue. In the heart, liver, and adipose tissue, these effects are mediated via
binding to ß1- & ß2-adrenergic GPCRs. Both ß-adrenergic GPCRs signal via Gas, which
activates adenylyl cyclase and raises intracellular [cAMP]. The a2-adrenergic GPCR signals
via Gai, decreasing adenylyl cyclase activity and intracellular [cAMP]. The a1-adrenergic
GPCR is coupled to Gaq, which activates phospholipase C (PLC) and signaling via the
IP3/DAG pathway . a1-adrenergic GPCRs are present in the liver and blood vessels in
peripheral organs. Binding to a1-adrenergic GPCRs stimulates glycogen breakdown in the
liver, while blood flow to peripheral organs is decreased.
glycogenال عن طريق energy productionبسبب زيادة ال peripheral organs*يقل تدفق الدم لل
degredation
GPCRs that Regulate Adenylyl Cyclase
Adenylyl cyclase is an effector enzyme that synthesizes cAMP. Ga-GTP subunits bind to
the catalytic domains of the cyclase, regulating their activity. Gas-GTP activates the
catalytic domains, whereas Gai-GTP inhibits them. A given cell type can express multiple
types of GPCRs that all couple to adenylyl cyclase. The net activity of adenylyl cyclase thus
depends on the combined level of G protein signaling via the multiple GPCRs. In liver,
GPCRs for epinephrine and glucagon both activate the cyclase. In adipose tissue ,
epinephrine, glucagon, and ACTH activate the cyclase via Gas-GTP, while PGE1(or PGE2)
and adenosine inactivate the cyclase via Gai-GTP.
*activation mean increase cAMP while inhibition decrease cAMP
........................................................
حسب احتياجات الخلية. inhibitoryوقد يسيطر تأثير ال stimulatoryال*قد يسيطر تأثير
Activation of Gene Transcription by GPCR Signaling
GPCRs regulate gene transcription by cAMP and PKA signaling. As shown in the figure,
cAMP-released PKA catalytic domains enter the nucleus and phosphorylate the CREB
(CRE-binding) protein, which binds to CRE (cAMP-response element) sequences upstream
of cAMP-regulated genes. Only phosphorylated p-CREB has DNA binding activity. p-CREB
interacts with other TFs to help assemble the RNA Pol II transcription machinery at these
promoters. In liver, glucagon signaling via this pathway activates transcription of genes
needed for gluconeogenesis.
........................................................
*في الصورة التالية يحدث ما يلي:
(denylylلل activationيعمل GPCRفي البداية
(cyclase فيزداد تركيز الcAMP ويتحد الcAMP مع
ثم بصير PKAلل regulatory subunit 2ال
release للcatalytic domins الي بدخلوا
activation(phosphrylation)وبعملوا nucleusال
الي بعمل CREالي يتحد مع ال CREBلبروتين اسمه
activation ومن ثمtranscription .
alzheimerاكتشاف مهم فله عالقة بال CREB*يعد
وله long-term memoryوال Parkinsonوال
وتأثر الجسم drugsعالقة بتأثر الدماغ بال
المختلفة hormonesبال
Down-regulation of GPCR/cAMP/PKA Signaling
A number of events contribute to the termination of signaling by a GPCR. These include:
1. dissociation of the hormone from the receptor,
2. hydrolysis of GTP by Ga
3. hydrolysis of cAMP via cAMP phosphodiesterase,
4. phosphorylation and “desensitization” of receptors by kinases such as PKA and ß-
adrenergic receptor kinase (BARK).
5. In addition, GPCRs can be removed from the membrane by vesicular uptake.
Biological functions mediated by 7TM receptors
1. Smell
2. Taste
3. Vision
4. Neurotransmission
5. Hormone secretion
6. Exocytosis
7. Control of blood pressure
8. Embryogenesis
9. Cell growth and differentiation
10. Development
11. Viral infection
12. Carcinogenesis
G-protein activation
molecular switch
b) Ligand binds
G-protein associates
(c) GDP-GTP exchange
-Subunit dissociates
Active G-Protein-GTP
-> allosteric modulator
of target effector enzyme
*allosteric enzyme : a special type of
enzymes that :-
• have activation site & inhibition site
• have catalytic subunits and regulation subunits
• have 4o structure (it is multisubunit) not only one unit
• are the most important enzymes in metabolism because they had 2 control steps in
metabolism pathways which are responsible of irreversible steps
........................................................
All G-proteins – similar structure/activation
There are TWO broad subclasses of trimeric G-protein-activated signal transduction
pathways:
depends on their target effector enzymes
A. adenylyl cyclase
B. phospholipase C
........................................................
An activated Ga-protein-GTP
– Can trigger the formation of cAMP, which
then acts as a second messenger in
cellular pathways
........................................................
G-protein-GTP activation of Effector
Enzyme adenylyl cyclase produces the
2nd messenger cAMP
Adenylyl Cyclase & Protein Kinase A
Adenylyl cyclase is an integral
membrane protein that contains 12
transmembrane segments (Fig.). It also
has 2 cytoplasmic domains that
together form the catalytic site for
synthesis of cAMP from ATP. One of
the primary targets of cAMP is a
regulatory kinase called protein kinase
A (PKA), or cAMP-dependent protein
kinase.
*الدكتور اكد على هاي األرقام عأساس ما نخلط بينهم:
adenylyl cyclease : 12 transmembrane segments / GPCR : 7 transmembrane segments
trimerc G protein : 3 subunits / PKA : 4 subunits
.segments 6تقسم لمجموعتين كل مجموعة فيها transmembrane segments protein 12*ال
........................................................
PKA exists in two different states inside cells (Fig.). In the absence of cAMP, the enzyme
forms a inactive tetrameric complex in which 2 PKA catalytic subunits are non-covalently
associated with 2 regulatory subunits. When cAMP concentration rises, cAMP binds to
the regulatory subunits which undergo a conformational change, releasing the active
catalytic subunits.
عبارة PKA*الدكتور في هاي الفقرة أكد على انه ال
catalytic & 2 2فيه tetramericعن
regulatory
أما cAMPيتصلوا بالمهمتهم regulatory*ال
activation of next stepمهمتهم catalyticال
PKA =< activeيكون ال cAMP*في حالة وجود ال
inactiveوفي حالة عدم وجوده يكون
........................................................
Protein Kinase A
Phosphorylates downstream target enzymes
activationوظيفتها kinases*ال
وظيفتها phosphrylase*ال
inactivation
What are targets for Protein Kinase A??
cAMP regulated pathways
*the more specific place were water reabsorption take place is distal convolated tubules & collecting ducts.
How to shut it off (the G protein cycle)?
G-protein -subunit is *معنى جملة)
timeron a :)
ما بتكون طول G protien alpha subunitال
في phosphorylationالوقت من بعد عملية ال
بل يوجد هناك وقت محدد تبقى activeحالة ال
ثم تتحول من تلقاء نفسها activeفيه في حالة ال
-auto shutفي عملية تسمى ) inactiveالى
off وقد تصبح هذه العملية اسرع في حال وجود )
GAP =< GTPase activatingبروتين يسمى
protein ( وهذا يفسر عبارةinherent
GTPase activity ولهذا فإنه في حالة عدم )
فإن توقف ligandاستمرار تزويد الخلية بال
بالتالي مسألة وقت ال أكثر و G proteinنشاط ال
وتنتهي استجابة effectorsسيتوقف تحفيز ال
الخلية للهرمون
*auto shut-off => G protein lose its ability to activate target protein that because GTP turn
to GDP and the G protein become inactive
*this process take few seconds or minutes(it is mentioned before in this lecture usually less
than 1 minute)
........................................................
( H2O)بنضيف اله hydrolysisبصيرله cAMP*ال
phosphodiesterase-cAMPبواسطة انزيم
proteinلل activationاول ما يعمل cAMP*ال
kinase وينتهي دوره بصيرلهdegradation وبتحول
ATPثم ADPثم AMPل
: phosphodiesterase-cAMP*هناك مواد تثبط ال
1)caffeine
2)bronchial dilators(such as Theophylline)
وبالتالي بزيد phosphodiesterase-cAMPلما تشرب كافيين يتم تثبيط ال >= caffeine*آلية عمل ال
وهو المطلوب وبهذا فانك ستشعر بالنشاط عند cAMPتاع ال functionأو ال actionال( وبزيد level of cAMP)ال
شرب الكافيين
يستخدم في بعض االمراض التنفسية مثل الربو ويعتبر dilatorbronchialيعتبر Theophylline*ال
drugلهذا ال target proteinهو ال phosphodiesterase-cAMPال
........................................................
phosphorylate <=kinases – phosphatases=> dephosphrylate
Diametrically Opposed…
Remember: whether you active or inactivate by adding P
depends on the specific protein
What if you can’t turn off cascade?
ايش inactiveللبروتين ايش ممكن يصير؟ / لو ما رجع البروتين لحالة ال shut ofحصل ما*معنى السؤال: اذا
ممكن يصير؟ اذا حصل هذا الشيء رح ينصاب الجسم بالعديد من االمراض والمشاكل ومن ضمنها الكوليرا
Vibrio cholera - causes cholera
Normal gut: H20, NaCl, NaHCO3 secretion controlled by hormones via Gs/cAMP signal
pathways
بشكل منظم وكميات معقولة intestineفي الوضع الطبيعي األيونات تفرز الى ال*
V. cholera – secretes enterotoxin, chemically modifies Gs – no GTPase activity - stays ON
تساهم في جعل عملية االفراز مستمرة )ببطل في عملية toxic proteins*في الكوليرا تفرز البكتيريا المسببة للمرض
inhibition للcAMP وبضل البروتينactive وبصير في خلل بالmembrane )
Severe watery diarrhea – dehydration, death
,مما يؤدي الى الجفاف وبالتالي الموت diarrheaمرض نتيجة ال*يتم فقدان الكثير من السوائل بسبب هذا ال
*يموت سنوياً مئات اآلالف من دولة اليمن الشقيقة نتيجة الكوليرا الناتجة عن التلوث وبخاصة تلوث المياة
target effector enzyme is Phospholipase C
PLC cleaves a membrane phospholipid (Phoshatidyl
inositol) to
two 2nd Messengers:
Inositol-1,4,5-Trisphosphate(InsP3) &
Diacylglycerol (DAG)
........................................................
*you should know :
1. DAG
a. 2 fatty acids + glycerol
b. lipid soluble
2. InsP3
a. 3 phosphate groups +suger nucleus
b. water soluble
GPCRs That Activate Phospholipase C
Another common GPCR signaling pathway involves the activation of phospholipase C
(PLC). This enzyme cleaves the membrane lipid, phosphatidylinositol 4,5-bisphosphate
(PIP2) to the second messengers, inositol 1,4,5-trisphosphate (IP3) and diacylglycerol
(DAG) (Fig.). In this case, the Go and Gq G proteins conduct the signal from the GPCR to
PLC. This is the pathway used in a1-adrenergic GPCR signaling in the liver.
,phosphatidylethanolamineهما نوعين: phospholipidة لتشابتر سابق أهم انواع ال*مراجع
phosphatidylcholine
IP3/DAG Signaling Elevates Cytosolic Ca2+
*Ca++ is important for neurotransmitters
The steps downstream of PLC that make up the IP3/DAG signaling pathway are illustrated
in the figure. IP3 diffuses from the cytoplasmic membrane to the ER where it binds to and
triggers the opening of IP3-gated Ca2+ channels (Steps 3 & 4). Another kinase, protein
kinase C (PKC) binds to DAG in the cytoplasmic membrane and is activated (Step 6). In
liver, the rise in cytoplasmic [Ca2+] activates enzymes such as glycogen phosphorylase
kinase, which phosphorylates and activates glycogen phosphorylase. Glycogen
phosphorylase kinase is activated by Ca2+-calmodulin. In addition, PKC phosphorylates
and inactivates glycogen synthase.
........................................................
*IP3
increase [Ca++] in cytosol from ER
Ca++ ==> activate PKC
PKC ==> do its function
*DAG
activate PKC
PKC ==> phosphorylation of substrates
........................................................
DAG
Activates
Protein
Kinase C
(Starts
Cascade)
..........
InsP3
Ligand
for ER
ligand-
gated
Ca++
channels
when ligand-gated Ca++ channels open the cell response occur
*the form at which Ca++ present in the cell is calcium-calmodulin complex
........................................................
Response:
Protein Kinase C phosphorylates target proteins (ser & thr)
cell growth
regulation of ion channels
cytoskeleton
increases cell pH
Protein secretion
Ca++
Binds & activates calmodulin Calmodulin-binding proteins activated (kinases &
phosphatases)
Signal Trans. Components: GTPase Switches
GTPase switch protein also play important roles in intracellular signal transduction .
GTPases are active when bound to GTP and inactive when bound to GDP. The timeframe
of activation depends on the GTPase activity (the timer function) of these proteins.
Proteins known as guanine nucleotide-exchange factors (GEFs) promote exchange of GTP
for GDP and activate GTPases. Proteins known as GTPase-activating proteins (GAPs),
stimulate the rate of GTP hydrolysis to GDP and inactivate GTPases.
GTPase-عكس ال responseوظيفته يسرع ال GEFs( exchange factors-guanine nucleotide(*البروتين
)GAPs( activating proteins الي وظيفته يثبط
........................................................
We will cover two classes of GTPase switch proteins--trimeric (large) G proteins, and
monomeric (small) G proteins. Trimeric G proteins interact directly with receptors,
whereas small G proteins interact with receptors via adaptor proteins and GEFs.
activating proteins-GTPaseال *
accelerating-GTPase الهوه نفسه
proteins
Signal Trans. Components: 2nd Messengers
While there are a large number of extracellular receptor ligands ("first messengers"),
there are relatively few small molecules used in intracellular signal transduction ("second
messengers"). In fact, only 6 second messengers occur in animal cells. These are cAMP,
cGMP, 1,2-diacylglycerol (DAG), and inositol 1,4,5-trisphosphate (IP3) , and calcium and
phosphoinositides . Second messengers are small molecules that diffuse rapidly through
the cytoplasm to their protein targets. Another advantage of second messengers is that
they facilitate amplification of an extracellular signal.
........................................................
*the most common 2nd messengers
Nitric Oxide (NO)/cGMP Signaling
A related signaling pathway involving phospholipase C operates in vascular endothelial
cells and causes adjacent smooth muscle cells to relax in response to circulating
acetylcholine (Fig.) In the NO/cGMP signaling pathway, the downstream target of
Ca2+/calmodulin is nitric oxide synthase, which synthesizes the gas NO from arginine. NO
diffuses into smooth muscle cells and causes relaxation by activating guanylyl cyclase and
increasing [cGMP]. As a result arteries in tissues such as the heart dilate, increasing blood
supply to the tissue. NO also is produced from the drug nitroglycerin which is given to
heart attack patients and patients being treated for angina.
حيث انه يعمل على زيادة تركيز nitroglycerin دواء ال(angina*يتم اعطاء األشخاص المصابين بالذبحة الصدرية)
ويعالج المصاب بالذبحة الصدرية vasodilationالذي يحدث NOال
........................................................
"second messenger"
هو تحفيز 2nd messengers*الهدف من كل هذه ال
لتقوم target proteinsيحفز اللكي PKCال
. responseبال
Metabolism of Diacylglycerol
Diacylglycerol may be (1) phosphorylated to phosphatidate or (2) hydrolyzed to glycerol and fatty acids(degraded).
Summary
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