Signal networks and pathways

Chitta Baral

Arizona State University

Goal: Decoding the cell and …

• Ultimate goal– Decoding the cell (understand what is happening

inside the cell)– Control cell behavior

• It involves– Accurate description of cellular biochemistry that

allows• Explanation of particular cell behaviors and phenotypes

(such as proliferation, cell migration, cell death, etc.)• Prediction of future behavior• Planning interactions with the cell to control the cell behavior

Biochemistry, network of interactions

• Relation between various molecules is often expressed using network of interactions (called Biochemical networks)

• Three kinds of biochemical networks– Metabolic networks

• Represent the chemical transformation between metabolites• Example: Glycolysis (breakdown of glucose)

– Signal networks (also called protein networks)• Representing protein-protein interactions

– Gene networks • Representing relationship between genes that encode the impact of

the expression level of one gene on another

• Pathways: Specific series of interaction in a network

G proteins

• 1994 Nobel prize. Discovery of G-protein coupled receptors and their role in signal transduction. Gilman, Alfred G. and Rodbell, Martin. http://www.nobel.se/medicine/laureates/1994/index.html

• 43,409 articles in Medline that mention G proteins.• 1. Basic facts about G proteins

– Each G-protein consists of 3 sub-units: G, G, and G. – These sub-units can be together or separated. – Each G-protein has a binding site that can be occupied by either

GDP or GTP. – A G-protein is said to be inactive when it is bound to GDP.– A G-protein is said to be active when it is bound to GTP.

cyclicAMP (cAMP) pathway

• 2. Up regulation: When a ligand binds to a receptor in a cell membrane the effect is that– A. the receptor becomes activated– B. the nucleotide binding site on the G-protein is

altered,– C. GTP replaces GDP,– D. GDP is released,– E. and G-GTP gets disassociated from GG.– F. G being active triggers the binding of G-GTP to a

membrane bound adenylate cyclase molecule,• activating it for production of cyclicAMP (cAMP).

G protein activation/inactivation cycle

cAMP

cyclicAMP (cAMP) pathway

• 3. Down regulation – when the ligand dissociates from the receptor the effects are– A. GTP is hydrolyzed by a GTPase activity on G

– B. G-GTP becomes G-GDP,

– C. and disassociates from adenylate cyclase molecule, making the later inactive.

– D. Gthen reassociates with GG

– E. CyclicAMP (in the cytoplasm) is then inactivated by the enzyme phosphodiesterase, which hydrolyzes it to AMP.

Interference with cAMP

• A. Cholera toxin inhibits the GTPase activity of the G-proteins of Gs subfamily, thus impacting 3A.

• B. The ras gene produces a G-protein that lacks GTPase activity, thus impacting 3A.

• C. Pertussis toxin inactivates the process that downregulates adenylate cyclase activity with respect to G-proteins of the Gi subfamily, with a few exceptions such as Gz, thus impacting 3C.

• D. G-proteins of the Gq subfamily are not modifiable by pertussis toxin or cholera toxin, thus nullifying A and C above.

A specific example of CAMP mediated regulation – Glycogen

degradation• In liver or muscle cells in presence of the ligand

epinephrine hormone (also called adernalin) increase in the cAMP concentration (in the cytoplasm)– activates protein kinase,– active protein kinases then converts inactive phosphorylase

kinase to active form by ATP dependent phosphorylation,– active phosphorylase kinase then phosphorylates (i.e.,

converts) less active phosphorylase-b to more active phosphorylase-a, and

– phosphorylase-a then catalyzes the phosphorolytic cleavage of glycogen into molecules of glucose-1-phosphate. (i.e., glycogen breakdown happens.)

References (for stuff so far)

• The world of cell 2nd ed. Becker, Deamer. Chapter 21. (Latest one is 5th ed, by Becker, Kleinsmith and Hardin; has a nice Cd-Rom with the book and explains signal transduction very well)

• Biochemistry of signal transduction and regulation. G. Krauss. Sec 5.5.1

• http://www.mun.ca/biology/desmid/brian/BIOL2060_W2003/CellBiol10/CB10.html

• Look for signal transduction, pathways, signal pathways, G protein, biochemical networks etc.

Reasoning about cAMP and Glycogen degradation

• Observation: cAMP concentration way above normal– Possible explanation: presence of Cholera toxin or

Pertussis toxin– Planning: How to overcome it using drugs.

• Predicting the impact of not having enough adernalin.

• Observation: Lack of glycogen breakdown.– Explained by low cAMP concentration

• Explained by G proteins not getting activated in adequate concentration

Computer Science (AI) challenges

• Represent signal networks– Such that we can reason with it– We can elaborate on it (add more details)

without making wholesale changes

• Reason with them– Explain observations.– Predict effect of particular actions.– Plan to make the cell behave a particular way.

Analogy with current AI research

• Goal: To represent effect of actions on the world, executability condition of actions, relation between objects in the world, etc.

• An example: – Description D.

• S1 Load causes loaded.• S2 Shoot causes ~ alive if loaded.• S3 Intially alive.• S4 Initially ~loaded.

– Planning: D |= ~alive after X. X=Load;Shoot.– Explanation: {S1, S2, S3, ~alive after shoot} |= Initially loaded– Prediction:

• D |= ~alive after Load? D |= loaded after Load?

Home work 3 (Due Feb 17th) – 100 pts

• Describe a particular signal pathway. – Draw the figure.– Write in English similar to the slides – Write in English-like syntax. (such as in the

previous slide and make up specific terms like `causes’ that you may need.)

– Source (journal: Cellular signaling; signal transduction chapter in books on cell biology; www.afcs.org. etc.) 50% bonus if you use a journal instead of a book.

Glossary• Activation

– A process of (i) initiating a chemical or biochemical reaction (ii) converting an inactive component to a functionally active form.

• Adenylate Cylase – The enzyme that catalyzes the synthesis of cyclic AMP (cAMP) from ATP,– ATP <--> cAMP + PP

• Allosteric – Pertaining to the topologically distinct sites on a protein or an enzyme molecule.

• AMP – An Adenosine 3'-monophosphate or Adenosine 5'-monophosphate nucleotide

with the phosphate group linked to the carbon 3 (or 5 resp.) of the ribose.• Arrestin

– Family of inhibitory proteins that bind to tyrosine-phosphorylated receptors, thereby blocking their interaction with G-proteins and effectively terminating the signaling.

Glossary (cont)• Channel Protein

– Proteins that form water-filled pores or channels across the membrane and are responsible for transporting solutes across the membrane.

• conformational change – change in the form differing in secondary or tertiary structure.

• cyclic AMP (cAMP) – An abbreviation for adenosine 3',5'-monophosphate or adenosine 2',3' -monophosphate. – An important intracellular regulator or second messenger for a number of cellular

processes in animals, bacteria, fungi and plants.• Effector molecule

– Small, biologically active molecule that acts as a regulator to control the activity of a protein or an enzyme by binding to a specific region on the protein or enzyme.

• Enzyme – Bioactive protein that catalyzes the biochemical reactions in the living cell.

• GAPs (growth-associate proteins) – Promote the hydrolysis of bound GTP, thereby switching the G-protein to the inactive form.

• GDP (Guanosine 5'-Diphosphate)• GEFs (guanine nucleotide exchange factors)

– Family of proteins that facilitate the exchange of bound GDP or GTP on small G-proteins such as ras and rho and thus activate them. (act in the opposite way to GAPs.)

Glossary (cont.)• G-Protein

– A GTP binding membrane protein that is capable of hydrolyzing GTP, activating membrane bound CAMP, and mediating a variety of signal transducing systems.

• GRK: G-Protein receptor kinase• GTP: Guanosine 5'-Triphosphate• GTPase (Guanosine triphosphatase)

– Enzyme that catalyzes the reaction GTP +H20 <---> Guanosine + Triphosphate• Guanine: A constituent base in nucleic acids.• Guanosine: A nucleoside and constitutent of nucleotides• Hydrolysis: Splitting of 1 molecule to 2 by incorporation of 1 water molecule.• Induction

– An increase in the rate of enzyme synthesis due to the presence of substrate or inducer.• Inhibition

– (i) reduction or prevention in the rate of enzymatic activity (ii) repression of physical or chemical activity.

• Kinase – The enzyme that catlyzes the transfer of a phosphate group from one compound to another.

• Nucleoside – A component of a nucleotide that consists of a nitrogenous base (purine or pyrimidine)

linked to a pentose sugar (ribose or deoxyribose)

Glossary (cont.)• Nucleotide: The basic building blocks of nucleic acids it consists of a nucleoside and a

phosphate. • phosphatase (two kinds: acidic and alkaline)

– acidic phosphatase is an enzyme that catalyzes the hydrolysis of a number of phosphomonoesters at acid pH but not phosphodiesters.

– Phosphoric monoester +H2O <--> Alcohol + Phosphoric acid.– alkaline phosphatase is an enzyme that catalyzes the hydrolysis of phosphomonoester at alkaline pH.

• phosphodiesterase – The enzyme that catlyzes the hydrolysis of phosphodiester bond in the polynucleotides or cyclic

nucleotides.• PKA: Protein kinase A• Protein

– A polymer of L-amino acids that folds into a conformation specified by the linear sequence of amino acids nd functions as an enzyme, a hormone, an antibody or a structural component of the cell.

• Regulatory enzyme – The enzyme that possesses a regulatory site for binding effector molecules in addition to the catalytic

binding site. • RGS (Regulators of G-Protein signalling)

– RGS is a protein that can increase the GTPase activity by more than one order of magnitude. • Transport protein: A protein that mediates the entry of specific substances into a cell.