Signal transmission with cell surface and intracellular receptors Prof.Dr.Gönül Kanıgür

Hormones Can exert their specific effects in three ways By influencing the rate of synthesis of enzymes and other proteins By effecting the rate of enzymatic catalysis By altering the permeability of cell membranes

The steroid hormones. Steroid hormones, small and fat soluble –made from cholesterol –travel through bloodstream bound to carrier proteins –enter cells by diffusion –include glucocorticoids (including cortisol), estradiol, testosterone, progesterone. –cross membranes by diffusion and enter the cell. –high-affinity receptors present in cytoplasm –hormone receptor complex binds to DNA at promoter regions upstream of certain genes –Transcriptional enhancement. gene expression increases. relatively slow, longer term response they regulate gene expression

Steroid hormones ;testosterone,estrogene and progesterone are the sex steroidsWhich are produced by the gonads -Corticosteroids are produced by the adrenal gland.glucocorticoids stimulate production of glucose, mineralocorticoids regulate salt and water balance Thyroid hormone,vitamin D3,retinoic acid are structuraly and functionaly distinct from steroids but they share a common mechanism of action in their target cells

steroid hormones,thyroid hormones,retinoids,and vitamin D are small hydrophobic molecules. They diffuse across the plasma membrane of the target cell And activate intracellular receptor proteins They directly regulate the transcription of specific genes

Protein hormones Some terminology The ligand is the signaling molecule [first messenger] Receptor protein that binds the first messenger [hormone] Effector,a hormone responsive enzyme in the cell. often they make seccond messengers [enzymes] or import them into the cell [ion channels]

There are three main classes of cell surface receptorsof hydrophilic proteins.peptides.other water soluble molecules. -ion channel linked receptors G-protein linked receptors Enzyme linked receptors İon channel linked receptor opens in responce to binding of its ligand fore exm.when the ligand binds to the, receptor,it alters its conformation so as open or close a channel for the flow of specific types of ions such as Na,K,Ca or Cl across the membrane

(1) Ligand-gate ion channels type (cyclic receptor) ligand→receptor→ion channel open or close 1. membrane receptors

2-Enzyme linked receptors They are directly linked to intracellular enzymes The largest family of this group is the receptor protein tyrosine kinases. Which phosphorylate their substrate proteins on thyrosine residues. İncludes the receptors polypeptide growth factors

More than 50 receptor protein tyrosin kinases have been identified,EGF,NGF,PDGF, İNSULİNand many growth factor They are involved in the control of animal cell growth and differentiation

Receptor tyrosine kinases Each receptor consists of an N-terminal extracellular ligand binding domain, transmembrane domain and a cytosolic C-terminal domain with tyrosine kinase activity. The EGF receptor consists of one polypeptide,whereas insulin receptor is a dimer form of two pairs of polypeptide chains(alfa and beta)

Growth factor binding induces Reseptör dimerization. It leads to autophosphorylation as the two polypeptide chains phosphrylate one another. SH2 domains of downstream signaling molecules bind to phosphotyrosine part of the activated receptors. It leads to their association with other proteins,promotes their phosphorylation and stimulates their enzymatic activities

3-G-protein linked receptors. The largest family of cell surface receptors Transmit signals to intracellular targets via the action ofG proteins Gprotein consist of alfa,beta,gama subunits Ligand binds tothe receptor.this interaction activates the G-protein,and than it dissociates from the receptor and carries the signal to an intracellular target which may be either an enzyme or an ion channel.

1) 7-helices transmembrane receptor G Protein-Coupled Receptors (serpentine R)

G protein refers to any protein which binds to GDP or GTP and act as signal transduction. G proteins consist of three different subunits ( , ,  -subunit).  -subunit carries GTPase activity, binding and hydrolysis of GTP. 2) G protein (Guanylate binding protein)

G s →  s →AC→cAMP↑ G i →  i →AC→cAMP↓ G q →  q →PI-PLC→IP 3 +DAG G o →  o →ion channel G t →  t →cGMP PDE→cGMP→ Rhodopsin 3) Classes of G protein

The molecules of signal transductions Ligands Receptors G-proteins Protein kinase and phosphatase Second mesengers [cAMP,cGMP,ca- calmodulin,IP3,DAG] Substrats of second mesengers

Pathway of G protein linked receptor HRG protein Es secondary messeger Protein kinase Phophorylation of Es or functional protein Biological effect

cAMP The cAMP is a second messenger in hormonal signaling cAMP is formed from ATP by the action of adenylate siklase Epinephrine receptor stimulates enzymatic activity via G protein.thereby increasing the intracellular concentration of cAMP cAMP binds to the regulatory subunits of protein kinase A.

Protein kinase A The inactive form of protein kinase A consists of two catalytic and two regulatory subunits.Binding of cAMP to the regulatory subunits induces a conformational change that leads to dissociation of the catalytic subunits. Catalytic subunit active and able to phosphorylate serine residues on target proteins. For example,it stimulates glycogen brekdown cAMPdependent protein kinase(protein kinase A) also activates the transcription of specific target genes that contain cAMP response element [regulatory sequence]

1. cAMP dependent-protein kinase A pathway HR G protein Phosphorylation of Es or functional proteins Biological effects PKA cAMPAC

Phosphorylate specifically Ser/Thr residues in several proteins （ 1 ） Regulation of metabolism （ 2 ） Regulation of gene expression

Ribosylation of Arg of G  G  s -ATPG  s -ADP AC cAMPCl - + H 2 O Cavity of intestine diarrhea HCO 3 - Cholera toxin ATPase

Pertussis toxin  i -ADP-ribosylation G i AC↑ allergy of histamine cAMP ↑

Phospholipids and calsium Many hormones and growth factors stimulate the hydrolysis of PIP2(phosphatidylinositol4,5 biphosphate) by phospholipase C Produces two distinct second messengers,diacylglycerol,and inositoltriphospate DAG activates protein kinase C and mobilize Ca from intracellular stores. Protein kinase C play important role in the control of cell growth an differentiation

Ca 2+ dependent PK pathway (1) Ca 2+ -DAG -dependent PKC pathway H R PIP 2 G protein PLC IP 3 DG ER PKC Ca 2+ PS Biological effects Phosphorylation of Es or functional proteins

Calmodulin The effects of Ca are mediated by the Ca- binding protein Calmodulin İncreased levels of cytosolic Ca/calmodulin complex activates a variety of target proteins,including Ca/calmodulin –dependent protein kinases.

Ca 2 ＋ -CaM dependent protein kinase pathway HR G protein PLC IP 3 Ca 2+ CaM CaMK Biological effects Phosphorylation of Es or functional proteins

IP3 and Protein kinase C have different effects in different cells a. IP3 i. Vascular smooth musclecontraction ii. smooth muscle contraction iii. skeletal muscle contraction iv. blood platelet aggregation of platelets b. Protein Kinase C. cell growth, differentiation (development into different tissue types), metabolism. i. blood plateletsserotonin release ii. mast cellshistamine release iii. smooth muscle contractility iv. nerve cells neurotransmitter release v. adipose tissuefat synthesis vi. liver cellsglycogen hydrolysis

Termination of the signal adenylyl cyclase remains active while hormones are present. the cell has a way to stop adenylyl cyclase activity when it the hormones are removed. – (see below) cAMP is continually broken down by phosphodiesterase – thus limiting the cell response Phosphatase continually reverses the phosphorylation of the three proteins, I.e. phosphorylase kinase, glycogen synthase and phosphorylase this also limits the cell response

4. end the response: a. a subunit is a GTPase, which hydrolyzes GTP to GDP and inactivates itself inactive  subunit reassociates with  and  this process requires an additional factor: GTPase activating protein (GAP). enhances GTPase and speeds up the timing step. b. drop in ligand concentration, resulting in dissociation and inactivation of the receptor c. active inactivation of the receptor, a process of desensitization. This means that the cell stops responding, even when ligand is still present around the cell – G-protein receptor kinase (GPRK), phosphorylates the receptor, – the phosphorylated receptor binds another protein called arrestin, thus inactivating it (even though hormone is still present and bound to the receptor) – bound arrestin molecules act as adaptors for Clathrin, allowing receptors to be internalized, thus further desensitising the cell