Eukaryotic cell structure

What are the limits to prokaryotic cell size?

• surface area : volume - lower SA per volume as gets bigger

• diffusion rates of molecules

• high concentrations of compounds + enzymes

What does compartmentalisation allow?

allows enzymes + substrates to be localised + concentrated

• allows cells to differentiate + specialise into specific functions

What is the function of the plasma membrane?

Cell boundary. Selectively permeable with transport systems that allow cell-cell interactions, adhesion, secretion + signalling.

What is the function of the cytoplasm?

Composed of cytosol and organelles. Location of many metabolic processes

What is the function of the cytoskeleton?

Composed of actin filaments, intermediate filaments + microtubules. Provides cell structure + movement

What is the function of the endoplasmic reticulum?

transport of materials + lipid synthesis

What is the function of ribosomes?

protein synthesis

What is the function of golgi apparatus?

• packaging and secretion of materials

• lysosome formation

• protein modification

• protein sorting for movement around cell

What is the function of lysosomes?

intracellular digestion

What is the function of mitochondria?

Energy production

What is the function of chloroplasts?

photosynthesis

found in plants, algae + photosynthetic bacteria

light + CO₂ → organic molecule

What is the function of nucleus?

houses genetic information

What is the function of nucleolus?

ribosomal RNA synthesis + ribosome construction

What is the function of cell wall?

strength + shape of cells

What is the function of cilia + flagella?

cell movement

What is the role of vacuole?

temporary storage, digestion + water balance

• large - fluid filled vesicles

• contain hydrolytic enzymes

• 30-90% of cell volume

What is a compartment without membranes called?

biomolecular condensates

• based on selective aggregation of some macromolecules

• concentrated zones of enzymes

How are biomolecular condensates formed?

requires scaffolds e.g. nucleic acid or proteins

• scaffolds form multiple weak, fluctuating binding interactions with themselves (aggregate)

• recruit specific proteins + nucleic acids into condensate (client proteins)

What are the characteristics of biomolecular condensates?

• highly dynamic + reversible properties

• liquid-like behaviour

• condensates of different properties can coexist

• liquid-liquid phase separation

How are cells studied?

• microscopes - live imaging

• fluorescent proteins - bind GFP to protein of interest to track it

What are the similarities between plant + animal cells?

• membrane-bound organelles

• similar membrane, cytosol + cytoskeleton elements

• function of organelles

What are the differences between plant + animal cells?

• plant cells larger than animals

• additional structures in plants - chloroplast, cell wall, vacuole

What are the double membraned organelles?

Nucleus, Mitochondria + chloroplast (M + C = own DNA)

What is the endosymbiosis theory?

Archaea genome similar to eukaryotic genome. Mitochondria + Chloroplast originate from the engulfment of bacteria by ancient archaea

What are the characteristics of the nucleus?

• contains genetic information - DNA

• DNA organised into chromosomes

• Nuclear envelope = double membrane

What is the structure of the nucleus?

• nuclear envelope = two membranes

  • outer nuclear envelope

  • perinuclear space

  • inner nuclear envelope

What are nuclear pores?

• point of fusion between inner + outer nuclear membrane

• gated by Nuclear Pore Complex (NPC) - made of 30 different proteins

How does the nuclear pore control what can enter/exit nucleus?

• small molecules can diffuse by passive diffusion - large molecules cant enter passively

• proteins created in cytosol can be transported through NPC if they have a Nuclear Localisation Signal (specific amino acid sequence)

• Importin = recognises NLS

• Binding of Importin to NLS allows transport through NPC

How does nuclear import use Ran-GTPase?

In cytosol - release of Ran-GDP allows protein to bind to importin

In nucleus - Ran-GTP binds to receptor to release protein

GTP hydrolysed to GDP as returns to cytosol

What is the process of nuclear export?

• protein being exported has a Nuclear - Export Sequence (NES)

• NES binds to Exportin

• also requires Ran-GTPase + Ran-GTP/Ran-GDP gradient across nuclear membrane

• important for export of RNA

What are the characteristics of the the nuclear lamina?

• tough fibrous mesh under inner membrane

• made of intermediate filament

• acts as scaffold to maintain shape of nucleus

• disassembles + reforms during division

What is the structure of chromosomes?

Chromatins (DNA wrapped in proteins - histones) associates with nuclear lamina. Each chromatin has its own discrete location - not randomly distributed

What is the nucleolus?

It is a biomolecular condensate - liquid like behaviour. Centre for ribosomal RNA (rRNA) production. Responsible for synthesis + assembly of RNA and protein to for ribosome.

What is the structure of endoplasmic reticulum?

• Continuous structure with outer nuclear membrane

• interconnected tubes + flattened sacs = high SA

• share single internal space - ER lumen

• 10% of cell volume

What are the type of endoplasmic reticulum?

Rough ER - ribosomes on surface = protein synthesis

Smooth ER = biosynthesis + metabolism of lipids

Transitional ER = producing vesicles with protein or lipid for transport to Golgi

How does rough ER carry out its function?

proteins are translocated into ER lumen while being translated from mRNA. Carries out simple modification of proteins + folding quality check.

Where is smooth ER found?

found in cells involved in lipid metabolism. Synthesises lipids for production of lipoprotein also contains enzyme for drug detoxification.

Muscles - special type = sarcoplasmic reticulum

• stores calcium ions to regulate muscle contraction

What is the structure of the golgi apparatus?

collection of flattened, membrane enclosed sac = cisternae

Cis complex = adjacent to ER + recieves proteins from ER

Medial cisternae = in middle

Trans complex = towards plasma membrane

What are the two models that explain vesicle trafficking through cell?

Vesicular transport model:

• resident enzyme stays in same cistern (proteins that modify proteins)

• cargo proteins are moved between each cisternae

Cisternae maturation model:

• cargo proteins stay in same cisternae

• resident proteins move between cisternae - cisternae mature between through layer

Why are transport vesicles coated?

determines where they go

Clathrin coated - from Golgi, end-some, plasma membrane

COPI coated - from Golgi

COPII coated - from ER

Retromer coated - endosome retrieval to Golgi

How do proteins move across membrane barrier?

Gated transport - e.g. nuclear pore complex

Protein translocation - proteins are synthesised into compartments

Vesicular transport - membrane-bound vesicles ferry proteins from one compartment to another

How does co-translation translocation occur?

• newly made peptide with ER signal sequence bind to signal recognition particle (SRP)

• SRP guides ribosome + RNA to translocator by binding to SRP receptor on plasma membrane

• protein translation occurs through translocator into ER lumen

• Proteins unfold during translocation and refold after

How are transmembrane proteins produced?

Stop transfer sequence prevents complete entry into ER lumen - proteins become transmembrane

How are vesicles coated?

1. cargo selection

2. memory bending

3. protein coating

4. coat disassembly

How do vesicles know where to go?

Vesicle coat = address label

Phospholipid profile = post code (differently phosphorylated)

Rab GTPases = postman

• guides vesicle to destination

• different ones guide to different locations

• GTP-binding = active

• GDP-binding = inactive

How does vesicle-membrane fusion take place?

1. Rab GTPase interacts with tethering proteins on target membrane

2. v-snare + t-snare (transmembrane proteins) interacts - highly complementary + specific

3. v + t-snare ‘zipping’ is what forces membranes together by removing water between two phospholipid bilayers

4. v + t-snare determine if vesicle fuses in correct destination

What is exocytosis?

movement out of a compartment

What is endocytosis?

movement to a compartment

What is the process of exocytosis?

delivery of newly made proteins, lipids + carbohydrates to cell surface

ER → Golgi → Plasma membrane

What are the types of exocytosis?

Constitutive exocytosis - continuous supply of plasma membrane with new lipid + protein

Regulated exocytosis - secretary vesicles stored + released upon stimulus

How does regulated exocytosis occur in insulin secretion?

insulin produced + stored in vesicles in pancreatic beta-cells

• glucose uptake results in increased ATP

• ATP inhibits potassium channels causing membrane depolarisation

• opening up calcium channels that trigger exocytosis of insulin-containing vesicles.

What is the process of endocytosis?

internalisation of large/small molecules and liquid

phagocytosis = large

pinocytosis = small

• endocytic vesicles bud inwards and is delivered to either lysosome or endosome.

• lysosome = digestion, endosome = recycled to plasma membrane

What is the process of phagocytosis?

• Internalisation of large particles e.g. bacteria. - lysosomes

• Immune cells that carry out phagocytosis = macrophages + neutrophils

• cell surface receptors on phagocytic cell must be activated before phagocytosis

• important for scavenging dead/damaged cells

What is the process of pinocytosis?

• uptake of small volume of extracellular fluid + bits of own plasma membrane

• cell volume + total surface area dont change

How do viruses enter cells by endocytosis?

virus can hack receptor-mediated endocytosis trafficking to enter host cell. Virus use receptor-binding domain of spike protein on its surface to bind to receptor on plasma membrane and activate endocytosis.

What are the characteristics of endosomes?

• intracellular sorting organelles

• connected membrane tubes + large vesicles

• early endosome = under plasma membrane

• late endosome = closer to nucleus

• acidic interior maintained by ATP driven H⁺ pump

• role:

  • recycling membrane receptors

  • lysosomal degradation

What are the characteristics of lysosomes?

• degrade proteins, nucleic acids, lipids + oligosaccharides

• contain hydrolytic enzymes - optimally active in acidic environment

• H⁺ pump make lumen acidic

• have specialised transporters to export useful metabolites out to cytosol - recycling

What are the functions of the plant vacuole?

• storage - nutrients + waste products

• degredation compartment

• cell size increase

• control turgor pressure

• maintain pH homeostasis

What are the characteristics of peroxisomes?

• single membrane enclosed organelles - very small

• contain oxidative enzymes: catalase, urate oxidase

• produces hydrogen peroxide

• catalases use hydrogen peroxide to oxidise lipids + detoxify harmful metabolites

• myelin sheath = highly dependant on phospholipid metabolism

What are the characteristics of mitochondria?

• highly dynamic

• highly plastic

• vary among cells - rbc = none, liver cells = more than 1000

• often associated with cytoskeleton

What is the double membrane structure of mitochondria?

outer membrane

• contains large, channel proteins e.g. porin

• permeable to small molecules

Inner membrane

• folded into cristae

• contains proteins for electron-transport chain and ATP synthesis

• contain transport proteins into + out of mitochondria matrix

What is the structure of the mitochondial matrix?

highly concentrated mix of enzymes + dna

1. mitochondrial DNA genome

2. mitochondrial ribosomes + tRNA

3. enzymes for oxidation of pyruvate + fatty acids + for citric acid cycle

What is the structure of the intermembrane space?

• concentration of small molecules - same as cytosol

• higher concentration of proteins - cytochrome C

• release of cytochrome C - apoptosis

What makes mitochondria dynamic?

they can divide by fission + fusion, and move by cytoskeleton

How is ATP produced in mitochondria?

occurs in mitochonrial inner membrane - OXIDATIVE PHOSPHORYLATION

1. Acetyl CoA production

2. Activated electron carrier: NADH + FADH₂

3. stepwise electron movement along electron transport chain

4. electrochemical gradient

5. proton flow back into mitochondrial matrix

6. ‘turn’ ATP synthase + create ATP

What is the electron transport chain?

4 large protein complexes - 3 = proton pumps

• during electron transfer from NADH, a hydride ion is removed and converted to a proton + 2 electrons

  • electrons donated to electron transport chain

  • electron moves through complex - induces conformational changes in protein = pumps proton from matrix to intermembrane space

  • energy to pump H⁺ from energy of electron transfer

• build up of H⁺ in intermembrane space - creates electrochemical gradient

How does proton motive force make ATP?

membrane potential + pH gradient = proton motive force

• H⁺ carrier spins rapidly within stationary head of F1 ATPase

• F1 ATPase converts ADP to ATP

• ATP synthesised in matrix is pumped back into intermembrane space

• ADP back into matrix

• done by a ADP/ATP carrier protein on inner membrane

What are the similarities/differences between mitochondria + chloroplast?

chloroplast generally larger than mitochondria

similar structures:

• double membrane

• highly permeable outer membrane

• tight intermembrane space

chloroplast have extra structure = thylakoid

What is the internal structure of chloroplast?

stacks of thylakoid = grana - contain chlorophyll

What are the characteristics of chlorophyll?

• absorb red + green light

• green light absorbed poorly - reflected

• this is why plants look green

• different chlorphylls have different absorption preferences

What is stage 1 of photosynthesis?

photosystem II = water splitting enzyme produces O₂

excided electrons are passed down via electron carriers to photosystem I

electron proton gradient allows ATP synthesis

What is stage 2 of photosynthesis?

light independant reaction

thylakoid membrane is impermeable to ATP + NADPH - no diffusion back into thylakoid membrane

ATP and NADPH + CO₂

• used to produce simple 3-carbon sugar

• exported + used as precursor for synthesis of other metabolites

What is the key enzyme for the light independent reaction?

Rubisco

slow + abundant

How do mitochondria + chloroplast collaborate?

• sugars made in chloroplast can be stored as starch or exported

• exported can be broken down + imported into mitochondria for efficient ATP synthesis

• O₂ released from chloroplast - used in oxidative phosphorylation

• CO₂ released from mitochondria - used in carbon fixation

How are ribosomes structured?

ribozyme = RNA enzyme

• two subunits - large + small

• by mass: 2/3 rRNA + 1/3 ribosomal protein

• eukaryotic 80s ribosomes - 40s subunit + 60s subunit

• assembled in the nucleolus

How are ribosomes made?

rRNA is folded into highly compact + precise 3D structure

ribosomal protein on surface + fills gaps in folded RNA core

contains 4 binding sites:

• 1 for mRNA

• 3 for tRNA - only 2 occupied at once

How do ribosomes carry out translation?

1. tRNA ‘charged’ with amino acid form base pair with complementary codon on mRNA

2. A + P sites very close - no base between

3. peptidyl transferase activity

4. large subunit translocate relative to smal subunit

5. position tRNA into exit site

6. small subunit translocate exactly 3 bases back to original position

How does ribosome reset after translation?

• ejection of used tRNA

• empty A site ready for next ‘charged’ tRNA to bind

• repeated until stop codon reached

What determines translation speed?

single mRNA can have multiple translations simultaneously - aslong as previous ribosome moves out the way, new one can bind

polyribosomes = especially important for highly-tanslated proteins

What are the types of cytoskeleton?

intermediate filaments, microtubules + actin filaments

• dynamic

• flexible

What is the structure of microtubules?

• hollow tube

• 13 protofilaments per microtubule

• made from α + β tubulin heterodimers

• ‘plus’ + ‘minus’ ends - β tubulin at + end

• each dimer makes 2x lateral contacts + 2x longitudinal contacts

What are the characteristics of α + β tubulin?

• highly conserved - found in all organisms in a similar structure

• each bind to one GTP molecule

  • α tubulin bound GTP = not hydrolysed

  • β tubulin = hydrolysed to GDP

How do you describe microtubule dynamic?

Nucleation - initiation of MT = start from scratch

Polymerisation - growth of MT = elongation of existing

How does microtubule nucleation occur?

• spontaneous MT formation requires very high concentrations of α + β tubulin heterodimers

• MT nucleation is catalysed by third type of tubulin = γ tubulin

• γ tubulin + accessory proteins - γ tubulin ring complex = promotes MT nucleation

• γ tubulin ring complex - enriched in MT organising centre in specific intracellular location

What is the MT organising centre?

centrosome = mtoc in most cells

• γ tubulin ring complex are enriched at centromere

• + end microtubule grows outwards relative to centromere

What is microtubule dynamic instability?

microtubules can both grow + shrink. requires constant energy imput - GTP

How do microtubules grow?

Addition of GTP bound tubulin dimers to growing (+) end of MT (GTP cap)

addition is faster than GTP hydrolysis to unstable GDP bound β-tubulin

How do microtubules shrink?

GTP bound β tubulin is hydrolysed, GDP bound β tubulin makes MT unstable and leads to self-unpeeling - catastophe

What is the function of dynamic stability?

• + end of microtubule can be stabilised by attaching to another molecule or cellular structure - wont undergo dynamic instability

• dynamic instability allows MT to explore cellular space

How do drugs affect dynamic instability?

Taxol (anti cancer drug) stabilises tubulin into MT lattice

• prevents depolarisation

• no shrinkage

• results in mitotic arrest + death

Colchicine (treats gout) binds free tubulin dimers

• prevents incorportation into MT - no polymerisation

• results in mitotic arrest + death

What is the function of microtubules?

organisation is related to function

• organelle positioning

• vesicle movement

• mitotic spindle

How do motor proteins aid vesicle movement?

motor proteins bind to microtubule - each type only moves in one direction + occurs in pairs

Kinesin - plus-end directed (towards + )

Dynein - minus end directed (towards - )

energy for movement from hydrolysed ATP

How do microtubules act as mitotic spindles?

during mitosis:

• MT nucleates from centrosome

• MT attached to kinetochore on chromosome = stabilised

• selective stabilisation of interpolar MT through MT associated proteins

How do chromosomes move during mitosis?

pulling force = motor protein on kinetochore MT towards centrosome

pushing force = motor proteins on interpolar MT push to elongate mitotic spindle

Where do cilia + flagella originate from?

extensions from centrioles - centre of centrosome. Form base of cilia/flagella (basal bodies)

How are MT organised in cilia/flagella?

basal body = cylinder of 9 MT filaments

• 9” 2” arrangement - 9 outer MT doublets + 2 central MT pairs

• outer MT doublets connected by nexin proteins

• Dynein motor proteins bind to MT doublets - provides movement

This structure allows Active Transport System to occur = Intraflagellar Transport - for assembly + maintenance of cilia