Advanced Cell Biology

2014 1nd Semester

Department of Animal Science

Chungbuk National University

6th Lecture

1st week : Introduction

3rd week :Research Strategies For Cell Biology

5nd week : Nucleus, Transcription and Splicing

7nd week : Membrane and Channel

9nd week : Membrane Trafficking

11nd week : Cell Signaling

13nd week : Cytoskeleton

15nd week : Cell Cycle

• Eukaryotic cells need to adapt to changing environmental conditions

• Transducing chemical or physical stimuli -> activity

• Short term responses mediates changes in– Metabolic activity– Secretion– Cell motility

• Long Term responses mediates changes in- Gene Expression

Why cell need Cell Signaling?

• Cell uses about 20 different families of receptor proteins to detect and respond to the different incoming chemical and physical stimuli

Receptors

• GPCR : uses trimeric GTP-binding proteins to relay signals to effector protein insie cells

• Mammals has ~2,000 different seven-helix receptors to discriminate odorants, light, amino acids, peptide, hormones, lipids.

• S375 orphan receptors : Receptors with unknown ligands

Seven-Helix Receptors

Ligands: alkaloids (opioids), biogenic amines (epinephrine), lipids (prostaglandins), neurotransmitters (acetylcholine, glutamate, dopamine), organic molecules (odorants), peptide hormones (glucagon, ACTH), proteins (parathyroid hormone, thrombin, interleukin-8), rhodopsin (photoreceptor pigment activated by light). Many useful drugs.

Receptor Tyrosin Kinase

Ligands : All ligands are proteins

- Insulin- EGF- Platelet-derived growth factor (PDGF)- Fibroblast growth factor (FGF)

Activation

- Unbounded (Inactive) receptor diffuse in the membrane- Ligands bring together two receptor molecules- Activation of tyrosin kinase - transphosphorylation of kinase domains

Receptor Tyrosin Kinase

Epidermal Growth Factor Receptor Kinase

Effectors

Phospholipase C gamma : bind P-tyr, produced IP3 and diacylgycerol GEF : binds P-tyr and activate Ras or Map Kinase

Cytokine Receptor associated with Cytoplasmic Tyrosin Kinase

Receptor Serine/Threonine Kinase

Protein Phosphorylation

Protein Kinase

Protein Phosphatase

Most common posttranslational modifications

Effects of Phosphorylation on Protein Structure and Function

- Interference of substrate binding

Serine/Threonine Kinase and Tyrosine Kinase

Regulation of Protein Kinase

• Activation by phosphorylation

- Src-family tyrosine kinase- MAP kinase- cyclin-dependent kinase- calcium-depenent kinase

• Inhibition by phosphorylation

- Phosphorylation of Myosin-Light Chain Kinase by PKC : inhibit activity

• Regulation of substrate binding

- binding of regulatory subunit : inhibit substrate binding- autoinhibtion

Regulation of Protein Kinase

• Targeting

- PKA location was determined by RI and RII subunit = Kinase-anchoring proteins

• PH Domain and lipid tag : target some kinase to lipid bilayer

• Scaffolding Proteins : Cellular ‘Matchmaker’

Protein Phosphatases

Number of Protein Kinase Genes in Human/Mouse Genome

- Serine/Threonine Kinase : 289- Tyrosine Kinase : 123

~= more than 500

Number of Protein Phosphatases - Serine/Threonine phosphatase : 18- Tyrosine phosphatase : 40

Total Number of Protein Phosphtase : less than 60

Substrate Targeting Subunit of PP

Protein Phosphatase 1 + Myosin Light Chain Kinase Targeting Subunits (MYPT1)

Catalytic subunit of Protein Phosphatase relies on Targeting Subunit for substrate specifities

GTP binding protein (GTPase)

Small GTPase

Ras family

Rho family : Rho, Rac, cdc42..

Rab family

Ran

Trimeric G-Proteins

Linker between signaling molecules

Small-conserved domains can be utilized as protein-protein linker between signaling proteins

Second Messengers in signaling pathway

- Cyclic Nucleotide

Activated by Adenylate/Guanylyl cyclase

Deactivated by Phosphodiesterase

Adenylate cyclase

cAMP level up: 1. PKA activation2. CDC25b phospho3. Inactive CDC25b4. Maintain GV

MilironedbcAMP

Lipid Second Messangers

PKC family of Protein Kinases

Activated by binding of lipid second messangers

Phosphatidylinositol metabolism

Calcium (Ca2+)- ATP-driven calcium pump (SERCA) remove calcium from cytoplasm (0.1uM)- Activated Inositol Triphosphate Receptor release ER-calcium into cytoplasm

Calcium binding of calmodulin

- Calcium binding of calmodulin change protein structure of calmodulin

- Ca2+ bound calmodulin regulate various protein targets

CaM Kinase (Calmodulin-dependent Protein Kinase)

Detection of Odors by the Olfactory System

Vertebrate Visual Transduction

B-Adrenergic signaling

Signaling Modulating gene expression

MAP Kinase Pathways

Insulin signaling Pathways

TGF-beta signaling Pathways