Question

Cell Biology Tutorial: Signal Transduction

Answer 1

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Answer 2

\({\bf{Signal~transduction}}\) is the process by which a cell releases a signal molecule and induces a response in another cell (sometimes the signal is induced in the same cell but we'll address that later). Some signaling molecules, such as steroids, can diffuse through the membrane, while others require cell-surface receptors.

A \({\bf{cell~surface~receptor}}\) contains three domains: an extracellular domain (exoplasmic), the plasma-membrane spanning domain, and the intracellular (cytosolic) domain. When a receptor binds to the receptor, it is transmitted through the membrane domain into the cytosolic domain, inducing a conformation change that will allow for the activation or inhibition of other proteins. These activated proteins catalyze the synthesis of small molecules or change the concentration of an ion.

Answer 3

\({\bf{types~of~signaling}}\)

endocrine: signaling molecules are transported through the ciruclatory system and act on distant target cells. ex: hormones

paracrine: signaling molecules act on nearby cells. ex: neurotransmitters, some growth factors

autocrine: signal molecules act on the cell that released it. ex. some growth factors, tumors

Answer 4

\({\bf{more~about~receptors}}\)

receptors:

have \({\bf{molecular~complementarity}}\)
bind noncovalently to ligands
have \({\bf{binding~specificity}}\)
may respond differently to the same ligand, ex. acetylcholine induces different responses in skeletal, muscular, heart, and pancreatic cells
lead to changes in protein phosphorylation

Answer 5

\({\bf{enzymes}}\)

kinases: add phosphate groups to target protein
phosphatases: remove phosphate groups from target proteins

in animal cells, there are two types of kinases - one adds phosphate to the hydroxyl on tyrosine and another adds phosphates to the hydroxyl groups on serine and threonine

Like most other enzymes, kinases have an N-terminal C-terminal, and a set of highly conserved amino acids around the catalytic sites. These kinases recognize the side chains and the amino acids on the phosphorylated residue.

Answer 6

\({\bf{GTP-ases}}\)

Two forms, the active form bound to GTP and the inactive form bound to GDP
\({\bf{GEFs}}\) mediate the switch from GTP and GDP forms

The time that a protein remains in the active conformation is dictated by the rate of GTP hydrolysis, which in turn is mediated by GTPase-activators (GAPs) and regulators of G protein signaling (RGSs) which accelerate GTP hydrolysis

\({\bf{Heterotrimeric G proteins}}\) bind directly to cell-surface receptors
\({\bf{Monomeric G proteins}}\) do not bind directly to cell-surface receptors. ex: Ras, Ran, Sar but can affect pathways like cell division/motility

Answer 7

\({\bf{second~messengers}}\) are molecules that bind to proteins and affect their activity

Calcium ions and cyclic adenosine monophosphate (cAMP) are the two most commonly discussed

The cystolic concentration of calcium is normally low, kept that way using ATP-pumps. Calcium channels can be opened to change the concentration very quickly.

In the human body an increase in cystolic concentration causes:

muscule contraction
exocytosis of hormones/neurotransmitters

the EF hand family proteins bind to calcium ions and in turn experience conformational changes that allow these proteins to bind to others, activating or deactivating them

Answer 8

\({\bf{amplification}}\) is the process by which the activation of an extracellular signal can increase thousands of second messengers, each of which in turn can activate other proteins. receptors are typically low-abundance so amplification is necessary to produce the needed response

Answer 9

\({\bf{the~dissociation~constant~Kd}}\) measures the affinity of the receptor for the ligand. high Kd = high dissociation of the ligand-receptor complex.

Kd = [R][L]/{RL]

Answer 10

\({\bf{binding~assays}}\) are used to study receptor affinity and specificity

Ligands are labelled, and thus, the number of ligand-receptor complexes can be studied as a function of ligand concentration.

It has been shown that only a very small percentage of receptors need to be bound to induce a cellular response.

Answer 11

\({\bf{affinity~chromotography}}\) is another analytical technique. First, the membrane is washed with a non-ionic detergent that separates the receptors from the other molecules without affecting the receptor. An antibody recognizes the receptor (or at least, a ligand for the receptor) and is linked in beads inside a column. The solubilized membrane is passed through the column, where the receptor protein binds to the receptor while all others are washed away.

Answer 12

\({\bf{immunoprecipitation~assays}}\) are used to measure the activity of kinases. The antibody is incubated with small beads of protein A. The beads are mixed with the cell/organelle, then washed to remove unbound proteins. The gamma-[P]ATP (only the gamma phosphate of the ATP is labelled) is used as a measure to quantify kinase activity.

Answer 13

\({\bf{western~blotting}}\) is used to determine the extend that a single amino acid side chain is phosphorylated. First, an amino acid is synthesized to have the same sequence around the phosphorylated amino acid. A monoclonal antibody is selected to only bind to the phosphorylated (not the non-phosphorylated) amino acid

Answer 14

\({\bf{GPR~receptors}}\) (G-protein coupled receptors) have:

1. a receptor with seven membrane-spanning alpha helices
2. heterotrimeric G protein
3. membrane-bound effecto protein
4. proteins involved in desensitization

Answer 15

The N-terminus of the G-receptor is on the exoplasmic side and the C-teminus is on the cytosolic side; it has four extracellular and four cystolic segments. A series of hydrophobic amino acids kanchor the receptor to the hydrophobic region of the plasma membrane. Segments of the alpha helices/loops form the ligand binding site on the exoplasmic side.

\({\bf{GTP~protein~subunits}}\) alpha, beta, and gamma. Only alpha binds to GTP/GDP, while the beta and gamma are bound to each other.

The steps of signalling/activation will be outlined below:

Answer 16

Upon binding, the receptor induces a conformation change that causes the release of GDP, so the system looks like:

Answer 17

the receptor is a GEF, causing GTP to bind to the G-alpha subunit, weakning the attachment to the receptor and allowing it to bind to and activate and effector protein

Answer 18

The GTP is then hydrolyzed by the G-alpha subunit, switching back to the inactive GDP state and begining the cycle again

the effector protein is a GAP (GTPase-activating protein) which enhances the binding of the GTP complex to the effector, another molecule that has this effect is RGS

Answer 19

\({\bf{This~is~the~end~of~my~tutorial;~I~hope~you~found~it~helpful.}}\)

\({\bf{If~you~have~any~*relevant*~comments~or~questions~I~will~attempt}}\)

\({\bf{to~address~them~to~the~best~of~my~ability.}}\)

\({\bf{Thank~you~for~reading!}}\)

Answer 20

The source of this information is \({\scriptstyle{Molecular~Cell~Biology~Eigth~Edition}}\)
\({\scriptstyle{Lodish,~et.al.}}\)