Chapter 3 Cells: The Living Unit Part B Shilla Chakrabarty, Ph.D.

Membrane Transport: Active Processes Active processes use energy (ATP) to move solutes across a living plasma membrane Two types of active processes: Active transport Vesicular transport

Active Transport Requires carrier proteins (solute pumps) Moves solutes against a concentration gradient Types of active transport: Primary active transport Secondary active transport

Primary Active Transport Figure 3.10 Extracellular fluid K+ is released from the pump protein and Na+ sites are ready to bind Na+ again. The cycle repeats. Binding of Na+ promotes phosphorylation of the protein by ATP. Cytoplasmic Na+ binds to pump protein. Na+ Na+-K+ pump K+ released ATP-binding site Na+ bound Cytoplasm ATP ADP P K+ K+ binding triggers release of the phosphate. Pump protein returns to its original conformation. Phosphorylation causes the protein to change shape, expelling Na+ to the outside. Extracellular K+ binds to pump protein. Na+ released K+ bound Pi 1 2 3 4 5 6 Energy from hydrolysis of ATP causes shape change in transport protein so that bound solutes (ions) are “pumped” across the membrane Sodium-potassium pump (Na+-K+ ATPase) Located in all plasma membranes Involved in primary and secondary active transport of nutrients and ions Maintains electrochemical gradients essential for functions of muscle and nerve tissues

1 Extracellular fluid Na+ Na+-K+ pump K+ ATP-binding site Cytoplasm Cytoplasmic Na+ binds to pump protein. Figure 3.10 step 1

Na+ bound P ATP ADP 2 Binding of Na+ promotes phosphorylation of the protein by ATP. Figure 3.10 step 2

Na+ released P Phosphorylation causes the protein to change shape, expelling Na+ to the outside. 3 Figure 3.10 step 3

K+ P 4 Extracellular K+ binds to pump protein. Figure 3.10 step 4

K+ bound Pi K+ binding triggers release of the phosphate. Pump protein returns to its original conformation. 5 Figure 3.10 step 5

K+ released 6 K+ is released from the pump protein and Na+ sites are ready to bind Na+ again. The cycle repeats. Figure 3.10 step 6

Stages of Primary Active Transport Extracellular fluid Na+ Na+-K+ pump ATP-binding site K+ Na+ bound Cytoplasm 1 Cytoplasmic Na+ binds to pump protein. P ATP K+ released ADP 6 K+ is released from the pump protein and Na+ sites are ready to bind Na+ again. The cycle repeats. 2 Binding of Na+ promotes phosphorylation of the protein by ATP. Na+ released K+ bound P Pi K+ 5 K+ binding triggers release of the phosphate. Pump protein returns to its original conformation. Phosphorylation causes the protein to change shape, expelling Na+ to the outside. 3 P 4 Extracellular K+ binds to pump protein. Figure 3.10

Secondary Active Transport Depends on an ion gradient created by primary active transport Energy stored in ionic gradients is used indirectly to drive transport of other solutes Cotransport — always transports more than one substance at a time Symport system: Two substances transported in same direction Antiport system: Two substances transported in opposite directions

Secondary Active Transport Cotransport—always transports more than one substance at a time Symport system: Two substances transported in same direction Antiport system: Two substances transported in opposite directions

Secondary Active Transport Symport System Transporting Sodium Ions And Glucose Into Cells Figure 3.11 The ATP-driven Na+-K+ pump stores energy by creating a steep concentration gradient for Na+ entry into the cell. As Na+ diffuses back across the membrane through a membrane cotransporter protein, it drives glucose against its concentration gradient into the cell. (ECF = extracellular fluid) Na+-glucose symport transporter loading glucose from ECF symport transporter releasing glucose into the cytoplasm Glucose Na+-K+ pump Cytoplasm Extracellular fluid 1 2

Vesicular Transport Transport of large particles, macromolecules, and fluids across plasma membranes Requires cellular energy (e.g., ATP) Functions: Exocytosis — transport out of cell Endocytosis — transport into cell Transcytosis — transport into, across, and then out of cell Substance (vesicular) trafficking — transport from one area or organelle in cell to another

Exocytosis Examples: Hormone secretion Neurotransmitter release Figure 3.14a 1 The membrane- bound vesicle migrates to the plasma membrane. 2 There, proteins at the vesicle surface (v-SNAREs) bind with t-SNAREs (plasma membrane proteins). Extracellular fluid Plasma membrane SNARE (t-SNARE) Secretory vesicle Vesicle SNARE (v-SNARE) Molecule to be secreted Cytoplasm Fused v- and t-SNAREs 3 The vesicle and plasma membrane fuse and a pore opens up. 4 contents are released to the cell exterior. Fusion pore formed Examples: Hormone secretion Neurotransmitter release Mucus secretion Ejection of wastes

Endocytosis Phagocytosis — pseudopods engulf solids and bring them into cell’s interior Example: Macrophages and some white blood cells Pinocytosis — Fluid-phase endocytosis. Plasma membrane infolds, bringing extracellular fluid and solutes into interior of the cell Example: Nutrient absorption in the small intestine

Endocytosis: Phagocytosis The cell engulfs a large particle by forming pro- jecting pseudopods (“false feet”) around it and en- closing it within a membrane sac called a phagosome. The phagosome is combined with a lysosome. Undigested contents remain in the vesicle (now called a residual body) or are ejected by exocytosis. Vesicle may or may not be protein- coated but has receptors capable of binding to microorganisms or solid particles. Phagosome Figure 3.13a

Endocytosis: Pinocytosis (b) Pinocytosis The cell “gulps” drops of extracellular fluid containing solutes into tiny vesicles. No receptors are used, so the process is nonspecific. Most vesicles are protein-coated. Vesicle Figure 3.13b

Endocytosis: Receptor-mediated Endocytosis Clathrin-coated pits provide main route for endocytosis and transcytosis Example: Uptake of enzymes low-density lipoproteins, iron, and insulin

Endocytosis: Receptor-mediated Endocytosis Vesicle Receptor recycled to plasma membrane (c) Receptor-mediated endocytosis Extracellular substances bind to specific receptor proteins in regions of coated pits, enabling the cell to ingest and concentrate specific substances (ligands) in protein-coated vesicles. Ligands may simply be released inside the cell, or combined with a lysosome to digest contents. Receptors are recycled to the plasma membrane in vesicles. Figure 3.13c

Endocytosis and Transcytosis Coated pit ingests substance. Protein- coated vesicle detaches. Coat proteins detach and are recycled to plasma membrane. Uncoated vesicle fuses with a sorting vesicle called an endosome. Transport vesicle containing membrane components moves to the plasma membrane for recycling. Fused vesicle may (a) fuse with lysosome for digestion of its contents, or (b) deliver its contents to the plasma membrane on the opposite side of the cell (transcytosis). Protein coat (typically clathrin) Extracellular fluid Plasma membrane Endosome Lysosome Transport vesicle (b) (a) Uncoated endocytic vesicle Cytoplasm 1 2 3 4 5 6 Involve formation of protein-coated vesicles Often receptor mediated, therefore very selective Figure 3.12

Coated pit ingests substance. Extracellular fluid Plasma membrane 1 Extracellular fluid Plasma membrane Protein coat (typically clathrin) Cytoplasm Figure 3.12 step 1

Coated pit ingests substance. Extracellular fluid Plasma membrane 1 Extracellular fluid Plasma membrane Protein coat (typically clathrin) Cytoplasm 2 Protein- coated vesicle detaches. Figure 3.12 step 2

Coated pit ingests substance. Extracellular fluid Plasma membrane 1 Extracellular fluid Plasma membrane Protein coat (typically clathrin) Cytoplasm 2 Protein- coated vesicle detaches. Coat proteins detach and are recycled to plasma membrane. 3 Figure 3.12 step 3

Coated pit ingests substance. Extracellular fluid Plasma membrane 1 Coated pit ingests substance. Extracellular fluid Plasma membrane Protein coat (typically clathrin) Cytoplasm 2 Protein- coated vesicle detaches. 3 Coat proteins detach and are recycled to plasma membrane. Endosome Uncoated endocytic vesicle Uncoated vesicle fuses with a sorting vesicle called an endosome. 4 Figure 3.12 step 4

Coated pit ingests substance. Extracellular fluid Plasma membrane 1 Coated pit ingests substance. Extracellular fluid Plasma membrane Protein coat (typically clathrin) Cytoplasm 2 Protein- coated vesicle detaches. Coat proteins detach and are recycled to plasma membrane. 3 Transport vesicle Endosome Uncoated endocytic vesicle 4 Uncoated vesicle fuses with a sorting vesicle called an endosome. Transport vesicle containing membrane components moves to the plasma membrane for recycling. 5 Figure 3.12 step 5

Coated pit ingests substance. Extracellular fluid Plasma membrane 1 Extracellular fluid Plasma membrane Protein coat (typically clathrin) Cytoplasm 2 Protein- coated vesicle detaches. 3 Coat proteins detach and are recycled to plasma membrane. Transport vesicle Endosome Uncoated endocytic vesicle Uncoated vesicle fuses with a sorting vesicle called an endosome. 4 Transport vesicle containing membrane components moves to the plasma membrane for recycling. 5 Lysosome 6 Fused vesicle may (a) fuse with lysosome for digestion of its contents, or (b) deliver its contents to the plasma membrane on the opposite side of the cell (transcytosis). (b) (a) Figure 3.12 step 6

Summary of Active Processes Energy Source Example Primary active transport ATP Pumping of ions across membranes Secondary active transport Ion gradient Movement of polar or charged solutes across membranes Exocytosis Secretion of hormones and neurotransmitters Phagocytosis White blood cell phagocytosis Pinocytosis Absorption by intestinal cells Receptor-mediated endocytosis Hormone and cholesterol uptake

Membrane Potential A membrane potential is caused by the separation of oppositely charged particles (ions) across a membrane. (The potential energy is measured as voltage) Resting membrane potential (RMP): Voltage measured in resting state in all cells Ranges from –50 to –100 mV in different cells Results from diffusion and active transport of ions (mainly K+)

Generation and Maintenance of RMP The Na+- K+ pump continuously ejects Na+ from cell and carries K+ back in Some K+ continually diffuses down its concentration gradient out of cell through K+ leakage channels Membrane interior becomes negative (relative to exterior) because of large anions trapped inside cell Electrochemical gradient begins to attract K+ back into cell RMP is established at the point where the electrical gradient balances the K+ concentration gradient A steady state is maintained because the rate of active transport is equal to and depends on the rate of Na+ diffusion into cell

Generation and Maintenance of RMP K+ diffuse down their steep concentration gradient (out of the cell) via leakage channels. Loss of K+ results in a negative charge on the inner plasma membrane face. 1 Extracellular fluid K+ also move into the cell because they are attracted to the negative charge established on the inner plasma membrane face. 2 A negative membrane potential (–90 mV) is established when the movement of K+ out of the cell equals K+ movement into the cell. At this point, the concentration gradient promoting K+ exit exactly opposes the electrical gradient for K+ entry. 3 Potassium leakage channels Protein anion (unable to follow K+ through the membrane) Cytoplasm Figure 3.15

Cell-Environment Interactions Involves glycoproteins and proteins of glycocalyx Cell adhesion molecules (CAMs) Membrane receptors

Roles of Cell Adhesion Molecules (CAMs) Anchor cells to extracellular matrix or to each other Assist in movement of cells past one another CAMs of blood vessel lining attract white blood cells to injured or infected areas Stimulate synthesis or degradation of adhesive membrane junctions Transmit intracellular signals to direct cell migration, proliferation, and specialization

Roles of Membrane Receptors Contact signaling — touching and recognition of cells; e.g., in normal development and immunity Chemical signaling — interaction between receptors and ligands (neurotransmitters, hormones and paracrines) to alter activity of cell proteins (e.g., enzymes or chemically gated ion channels) G protein–linked receptors — ligand binding activates a G protein, affecting an ion channel or enzyme or causing the release of an internal second messenger, such as cyclic AMP

Mechanism Of Action Of G Protein-Coupled Receptor 2 1 3 4 Plasma membrane G protein (inactive) CYTOPLASM Enzyme Activated receptor Signaling molecule Inactive enzyme Activated enzyme Cellular response GDP GTP P i

Cascade of cellular responses (metabolic and structural changes) Mechanism Of Action Of G Protein-Coupled Receptor 1 Ligand (1st messenger) binds to the receptor. 2 The activated receptor binds to a G protein and activates it. 3 Activated G protein activates (or inactivates) effector protein (e.g., an enzyme) by causing its shape to change. Extracellular fluid Effector protein (e.g., an enzyme) Ligand Receptor 4 Activated effector enzymes catalyze reactions that produce 2nd messengers in the cell Inactive 2nd messenger G protein 5 GDP Active 2nd messenger Second messengers activate other enzymes or ion channels 6 Activated kinase enzymes Kinase enzymes transfer phosphate groups from ATP to specific proteins and activate a series of other enzymes that trigger various cell responses. Cascade of cellular responses (metabolic and structural changes) Intracellular fluid Figure 3.16