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Cell Biology

Here are the notes for four of the nineteen topics of an upcoming examination.

Cell signalling

The technicalities

Paracrine, endocrine, synaptic.

How does basically anything meaningful happen in a cell?

Reception

Signalling molecules bind to receptors.

G-protein coupled receptors: cell-surface transmembrane receptor which works with G-protein (GTP-binding)

  • G-protein: attached but mobile along interior of cell membrane, molecular switch (state: GDP or GTP)
  • signalling molecule binds to extracellular binding site
  • receptor activated and changes shape, binding to an inactive g-protein; GTP displaces GDP
  • active g-protein dissociates from receptor, moves along membrane and binds to enzyme
  • g-protein serves as GTPase and hydrolyses GTP to GDP; available for reuse

Receptor tyrosine kinases: monomers with intracellular tails containing multiple tyrosines

  • signalling molecule binds to extracellular binding site
  • monomers dimerise and undergo auto-/cross-phosphorylation: phosphate moved from ATP to tyrosine
  • recognised by specific relay proteins which bind to tyrosines

Ligand-gated ion channels: membrane channel receptor containing a “gate”

  • ligand binds to receptor, opening channel
  • specific ions flow through channel and rapidly change concentration of ion in cell
  • ligand dissociates; channel closes

Intracellular receptors: for hydrophobic or small signalling molecules which pass through cellular membrane

  • steroid / thyroid hormones; nitric oxide
  • hormone passes through plasma membrane and binds to receptor protein, activating it
  • hormone-receptor complex enters nucleus and binds to specific genes
  • bound protein serves as transcription factor, stimulating transcription into mRNA and translation to protein

Transduction

Receptors undergo a conformational change and the signal is converted to a form eliciting cellular response, often via signal transduction pathways

Phosphorylation: phosphate groups added by protein kinases (phosphorylation cascade) and removed by protein phosphatases

Second messengers: small water-soluble non-protein molecules or ions which diffuse throughout cell (less expensive)

  • Commonly cAMP and Ca2+
  • Example: G-protein activates phospholipase C which cleaves PIP2 into DAG and IP3
  • IP3 serves as second messenger and opens IP3-gated calcium channel in ER
  • Calcium ions (second messenger) activate proteins

Response

just do your thing I guess

  • Protein synthesis regulation via gene regulation (activate transcription factors)
  • Protein activity regulation (opening/closing ion channels, etc)

Signal amplification: an effect of cascades (every step)

Specificity and coordination: branching pathways, cross-talk, etc

Signalling efficiency: scaffolding proteins attach several relay proteins

Signal termination: binding reverses via several means (conversion of second messengers; phosphatases; G-protein hydrolysis)

Cell cycle and Cancer

Necrosis: accidental cell death; swelling and lysis; results in inflammation

Apoptosis: programmed cell death; membrane blebbing, chromosome condensation, packaging into apoptotic bodies for phagocytosis

  • Discovery via studies of C. elegans (consistent decrease in number of cells, implying apoptosis)

Cell cycle

G1: pre-synthesis; S: DNA replication; G2: post-synthesis (prepare for mitosis)

Three important checkpoints: G1 (enter G0), G2 (abort mitosis), M (ensure kinetochore attachment before anaphase)

Cyclin-dependent kinases: a certain class of kinases which are activated by cyclin partner, always present

Cyclin: protein named for its cyclic expression changes, synthesised or degraded at various stages of cell cycle

Cyclin and CDK partnerships not exclusive

Six hallmarks

Cancer occurs when:

1. cells want to live (self-sufficiency in growth signals)

Refer to section Cell Signalling.

Self-sufficiency could develop due to any one of:

  • Reception
    • Faulty receptor: always activated
  • Transduction
    • Proteins involved in phosphorylation cascade always activated
  • Response
    • Transcription factor always activated
    • Production of more growth factor

2. cells don’t want to die (insensitivity to anti-growth signals)

Ignoring cell checkpoints (continuing mitosis, etc.)

Ignoring typical inhibitions

  • Contact inhibition
  • Anchorage dependence
  • Density-dependent inhibition

resulting in chaotic growth.

3. killing them doesn’t kill them (apoptosis evasion)

Ineffectiveness of tumour-suppressor genes (e.g. p53)

4. time doesn’t kill them (replicative potential; telomerase)

Telomerase results in telomeres extending during mitosis; infinite replicative potential

5. food doesn’t kill them (angiogenesis)

Tumours secrete angiogenic factors, enabling angiogenesis (blood vessel formation)

This enables rapid tumour growth and later metastasis.

6. space doesn’t kill them (metastasis)

Cancer cells eventually circulate through bloodstream and form tumours elsewhere.

Multistep progression model

1. Mutations

Tumour-suppressor genes (preventing abnormal cell growth) inactivated / absent.

Example: p53 “guardian of genome”

Inactive form: p53-mdm2 complex

Upon DNA damage or cell cycle abnormalities: p53 is activated and results in:

  • Cell cycle arrest, DNA repair, cell cycle restart
  • Apoptosis, elimination of damaged cell

Recessive loss-of-function mutations: with two mutated genes no normal protein is expressed.

Proto-oncogenes (encouraging cell proliferation etc.) mutate into oncogenes.

This results in overproduction of growth factors; mutant receptors; mutant relay proteins; mutant transcription factors.

Example: Ras (G-protein)

A hyperactive Ras protein issues signals regardless of receptor conformation, resulting in protein overexpression and increased cell division.

2. Activation of telomerase

Telomerase: present in germ cells (elongate telomeres during replication)

Immortal cells: after 10-20 cell doublings, telomerase is reactivated to maintain telomere length

3. Angiogenesis

4. Metastasis

respiration

Okay.

Respiration: synthesis of ATP from glucose (and oxygen).

ATP: energy currency

NADH, FADH2: high-energy electron shuttles

1. Glycolysis: oxidising glucose into pyruvate

Ten-step process; energy investment and payoff phases

Beginning with glucose:

  1. Add 6-phosphate (-ATP)
  2. Convert glucose to fructose
  3. Add 1-phosphate (-ATP)
  4. Cleave into G3P and DHAP
  5. G3P ⇌ DHAP (never reaches equilibrium, G3P used)

From here on everything happens twice per glucose (once per G3P).

  1. G3P oxidised and 1-phosphate added (+NADH)
  2. Phosphorylation (+ATP)
  3. Relocate 3-P to 2-P
  4. H2O removed to form PEP
  5. Phosphorylation (+ATP) to form pyruvate

Census per glucose:

  • ATP investment: -2
  • ATP payoff: 2(+2)
  • Net ATP: +2
  • Net NADH: +2

Occurs as pyruvate enters mitochondrion

  • Decarboxylation to form acetyl (+CO2)
  • Redox reaction (+NADH)
  • Binding with CoA forming acetyl-CoA

Census per glucose:

  • Net NADH: 2(+1)
  • Net CO2: 2(+1)

2. Krebs cycle

Or citric acid cycle or whatever you call it, really.

  • Oxaloacetate (4C) + acetyl-CoA (2C) to form citrate (6C)
  • which forms isocitrate (6C)
  • which is oxidised to form ketoglutarate (5C) (+NADH, +CO2)
  • which is oxidised to form succinyl CoA (4C) (+NADH, +CO2)
  • from which CoA is removed, forming succinate (4C) (+GTP, which is just ATP in a trench coat)
  • which is oxidised to form fumarate (4C) (+FADH2)
  • which is hydrated to form malate (4C)
  • which is oxidised to form oxaloacetate (4C) (+NADH)

Census per glucose:

  • Net ATP: 2(+1)
  • Net NADH: 2(+3)
  • Net CO2: 2(+2)
  • Net FADH2: 2(+1)

Overall census so far:

  • Net ATP: 4
  • Net NADH: 10
  • Net FADH2: 2

3. Oxidative phosphorylation

Electron transport chain

Electrons are passed along membrane proteins with increasing electronegativity.

NADH: I → Q → III → Cytochrome C → IV → O2 (forming H2O)

FADH2: II → Q → III → Cytochrome C → IV → O2 (forming H2O)

I, III, IV are transmembrane proton pumps which pump protons out of the mitochondrion.

II is not; FADH2 is slightly less efficient.

Chemiosmosis

H+ gradient across mitochondrial membrane: protons diffuse back in across ATP synthase due to proton-motive force, forming ATP.

ATP: 4 ATPNADH: 2.5(10)=25 ATPFADH2: 1.5(2)=3 Possible -2

Total: 4 + 25 + 3 − (0 or 2) = 30 or 32 ATP.

Anaerobic respiration

Glycolysis still occurs as usual.

For anaerobic respiration: the electron transport chain still occurs (but not ending in O2).

Alternatively fermentation occurs outside the mitochondrion.

  • Products: ethanol, lactate. etc.

Obligate anaerobes/aerobes; facultative anaerobes

Versatility

Various molecules can undergo catabolism at different points

  • Fructose enters directly at step 3
  • Amino acids converted to intermediates of glycolysis / citric acid cycle
  • Glycerol converted to glyceraldehyde
  • Fatty acids converted to acetyl CoA

Regulation

Phosphofructokinase (step 3) used to control respiration

  • Excess ATP inhibits
  • Excess citrate inhibits
  • AMP stimulates

Photosynthesis

…I don’t need to give an introduction, do I?

Chloroplasts

These comprise stroma (fluid), and grana (singular: granum) with folds known as thylakoids.

Chlorophyll comprises a light-absorbing porphyrin ring and hydrocarbon tail.

  • Electron excited when struck by photon
  • Energy emitted as heat and photon (fluorescence)
    • Increased wavelength

Chlorophyll α: key light-capturing pigment, participates directly in reactions

Chlorophyll β: accessory pigment (transfers energy to other pigments)

Carotenoids (carotene, xanthophylls): accessory pigments (prevent overexposure; broaden absorption spectra)

1. Light reaction

Photosystem: collection of light-harvesting complexes around a reaction-centre complex in a thylakoid membrane

Photosystem II: reaction-centre chlorophyll α: P680 (absorbs 680nm best)

Photosystem I: reaction-centre chlorophyll α: P700

Process:

  • Photon strikes chlorophyll of PS II, exciting electron
  • Electrons excite each other; energy relayed between chlorophyll molecules until striking P680s
  • Excited electron transferred to primary electron acceptor
  • Photolysis (bad name) of water into oxygen, hydrogen ions and electrons (replenishing P680)
  • Electron passed to P700s at PS I via electron transport chain: plastoquinone (Pq), cytochrome complex, plastocyanin (Pc)
    • H+ pumps create a proton gradient across thylakoid membrane; ATP synthase
  • (repeat but with PS I)
  • Electron passed to ferredoxin (Fd) and transferred (via enzyme) to NADP+ → NADPH

Cyclic electron flow

The electron passed to Fd can be passed back to the cytochrome complex, forming a cycle.

  • Inefficient; ultimately counterproductive
  • “Evolutionary leftover”

2. Dark reaction (Calvin cycle)

Analogous to Krebs cycle

For 3 CO2:

  1. Carbon fixation
    • 3 RuBP + 3 CO2 → 6 3-phosphoglycerate
    • Add 1-phosphate (-6 ATP)
  2. Reduction
    • Reduced (by NADPH) to form 6 G3P (1 G3P output)
  3. Regeneration
    • 5 G3P to 3 RuBP (-3ATP)