FCP-1: Cell Biology

1st contact session: cell membranes, cytoplasmic organelles, the cytoskeleton, intercellular

connections, cell adhesion molecules, transport across cell membranes, ATP production

Part 1: intracellular structures and organelles

Simplified depiction of a cell

Cell membrane components

• Main component: phospholipids (hydrophilic outside, hydrophobic inside, spontaneous bi-layer)• Selectively permeable• Inner membranes have similar structure• Proteins: integral vs peripheral• Modifications• Anchors

Cell adhesion molecules, pumps, channels, receptors, enzymes

Mitochondria (1)

Main function: energy production throughoxidative phosphorylation

Mitochondria (2)

• Used to be free-living bacteria• Contains the components of the electron transport chain

(energy production) in the inner membrane• Contains own genome (smaller than nucleus) and

ribosomes (protein synthesis machinery)• Zygote mitochondria come from the ovum: maternal

inheritance of mtDNA• Very ineffective DNA repair leads to mistakes: results in

a large number of rare diseases associated with defects in energy metabolism

Mitochondria (3)Electron transport chain (oxidative phosphorylation, generation of ATP/energy):

Later…

Lysosomes: rubbish bins

• Large, irregular structures in the cytoplasm• Acidic interior, digest endocytosed bacteria and

discarded cell components• Filled with acid hydrolases, cannot function at normal

cellular pH, will not destroy other cell components• Lysosomal storage diseases result from absence of

enzyme, accumulation/engorgement of lysosomes

Peroxisomes: detox and more • Catalyse various anabolic and catabolic reactions, e.g.

breakdown of very long chain fatty acids, production of plasmalogen (myelin), production of bile acids

• Enzymes oxidize substrates, generating toxic H2O2, used to oxidize other substrates, neutralizing H2O2

• NB for the detox of ethanol• PXR gene product is outer pxome receptor, PEX gene

products import proteins into pxome, and enzymes are targeted into pxome by PTS signal

• Errors in pxome assembly result in Zellweger syndrome, neonatal adrenoleukodystrophy and infantile Refsum disease (lethal in infants)

Nucleus: command HQ

• Contains all of the DNA (nuclear genome) required for gene expression, in the form of chromatin• Site of gene expression (DNA → mRNA)

Nucleus

• DNA (chromosomes) normally unravelled, disorganized: chromatin

• Individual chromosomes condense before cell division• Nucleolus contains RNA, proteins:

ribosome assembly• Nuclear envelope a double-layer membrane• Contains pore complexes for shuttling of

proteins, ribosomes and RNA: ribosomes and RNA produced in nucleus, must shuttle to cytoplasm for protein synthesis, some proteins (i.e. transcription factors) must shuttle back to nucleus

Ribosomes: protein assembly lines

Endoplasmic reticulum: processing

• Complex series of tubules in the cytoplasm• Contiguous to the nuclear membrane• Smooth ER: steroid synthesis • Rough ER: covered with ribosomes,

protein synthesis, folding and

modification

Golgi apparatus: add some sugar

• Stacked membrane-enclosed sacs• Proper glycosylation (sticking on carbohydrate/sugar

chains) of lipids and proteins• Directional (cis→trans)• Vesicles shuttle from the ER, through the Golgi, out for

secretion

Cytoskeleton: intracellular highways

• Maintains structure, helps to move and change shape• Also moves proteins and organelles around

Molecular motors to move cargo

Kinesin, dynein, myosin: all use ATP (energy)

Part 2: Intercellular connections

Holding cells together: Tight junctions

• Surround the outer layer of epithelial cells (intestinal mucosa, renal tubules, choroid plexus in brain)• Also contribute to endothelial barrier function• Totally obliterates the gap between cells, prevents protein leakage between cells

Holding cells together: zonula adherens

Holding cells together: desmosomes

- Adhesion protein = cadherin, helps to withstand shear stress in epithelium, particularly in epidermis- Defining feature: dense plaques on cytoplasmic side, attached to cytoskeletal filaments- Blistering diseases (Pemphigus) are auto-immune, attack desmogleins(cadherins), cause layers of skin to pull apart

Attaching cells to the basal lamina: hemidesmosomes and

focal adhesions

Gap junctions: intercellular communication

• 1 subunit = connexin• Pore with 6 connexins = connexon• permit passage of ions and small metabolites between cells• highly selective (20 diff connexin genes, each for different flow-through)

Cell adhesion molecules

• All intercellular connections consist of cell adhesion molecules (CAMs)

• 4 broad families: integrins, cadherins, selectins and IgG adhesion molecules

• Not just for adhesion, but also for signalling:• cells that lose contact with other cells undergo dissociation-induced apoptosis (anoikis)• collagen-integrin interaction essential for osteoblast differentiation

Part 3: transport across cell membranes

Exo- and endocytosis

Note that the cytoplasmic side of the membrane always remains the cytoplasmic side

Endocytosis continued

• Phagocytosis: eating of bacteria, dead tissue by leukocytes

• Pinocytosis: drinking of solutes• Both processes involve invagination of the plasma

membrane before pinching off vesicle inside the cell• Clathrin-mediated endocytosis: three-legged clathrin

molecules cover endocytotic vesicle

(NB for receptor internalization and

synaptic function)

How do molecules move across the cell membrane?

• Small non-polar and neutral polar molecules diffuse directly across (O2, N2 CO2)

• Everything else needs help!• Transport proteins form channels for transport of various

molecules• Even water! (through aquaporins)• Some are non-selective ion

channels, some are very selective

How do molecules move across the cell membrane?

• Some channels are gated

(opened upon a particular

stimulus):

How do molecules move across the cell membrane?

• Carrier proteins transport molecules WITH a concentration or electrical gradient: facilitated diffusion, does not require energy (example: glucose)

• Other carriers transport molecules AGAINST a gradient: active transport, requires energy

• Many carrier proteins are therefore ATPases: hydrolyses ATP for energy for transport

• Secondary active transport: transport of one molecule coupled to the transport of another (often Na+)– Symport: two molecules moving in the same direction– Antiport: exchange of molecules in opposite directions

Ion channels

Possible configurations:

Part 4: Energy (ATP) production

ATP hydrolysis = energy

ATP → ADP + Pi + 30-50 kJ energy

Energetically unfavourable (unstable) Energetically more stable

Interesting factoid: 60% of energy goes towards maintenance of body temp

Main site of ATP production: the citric acid cycle

cytoplasm

mitochondria

But before we get to this point…..

Glycolysis (Embden-Meyerhof pathway)

1x 6-carbon

2x 3-carbon

Net gain (1 mol glucose): 4 ATP – 2 ATP = 2 ATP; 2 pyruvate; 2 NADH

Or….Glycogen breakdown

Net gain from 1 mol glucose-6-phosphate: 4 ATP – 1 ATP = 3 ATP; 2 pyruvate; 2 NADH

Or…Beta-oxidation of fatty acids

- Takes place in mitochondria: long-chain fatty acids transported in by carnitine- 18-C fatty acid generates 8 acetyl-CoA

Main site of ATP production: the citric acid cycle

cytoplasm

mitochondria

From NADH/FADH2 to ATP

ATP production: adding it up

• 1 pyruvate generates 4 NADH, 1 FADH2 and 1 GTP (ATP)

• 1 NADH = 3 ATP, 1 FADH2 = 2 ATP

• 1 pyruvate = (4x3) + (1x2) + 1 = 15 ATP• 1 glucose (2 ATP; 2 pyruvate; 2 NADH) = 2 +

(2x15) + (2x3) = 38 ATP• 1 glucose-6-P (from glycogen) = 39 ATP• 1 18-C fatty acid = 8 x 15 = 120 ATP• 1 triglyceride ≥ 360 ATP