The eukaryotic cell

Question 1

Cellular components and organelles

Answer 1

Functions = providing shape and support, locomotion, reproduction
Types = double membrane bound organelles, single membrane bound organelles, cellular components without membrane
Organelles – membrane-bound compartments or structures in a cell that performs a special function

Question 2

Nucleus

Answer 2

• Largest organelle in animal cells
• Consists of nuclear envelope, nucleoplasm, nuclear lamina, chromosomes and chromatin, nucleolus
• Nuclear envelope = two membranes both phospholipid bilayers with different types of proteins. Inner defines nucleus, outer is continuous with er. Perinuclear space between membranes is continuous w lumen of rer. Nuclear pores where two membranes fuse
• Nuclear pore = ring-like complexes composed of specific membrane proteins through which material moves between nucleus and cytosol. Made up mainly of multiple copies of different proteins calls nucleoporins. 34 distinct nucleoporins types and >456 individual proteins. Nucleoporins help cargo proteins traverse the nuclear pore. Ions, small metabolites and globular proteins diffuse through water-filled channel in nuclear pore complex. Larger proteins and ribonucleoprotein complexes need the assistance of soluble transporter proteins
• Nucleoplasm = suspends structures within nucleus that are not membrane bound e.g chromosomes, nuclear bodies, DNA, RNA, minerals, also called karyoplasm or nuclear sap, semi-solid, granular substance that contains many proteins, responsible for maintaining shape and structure of the nucleus, protein fibres form a crisscross matrix within nucleus
• Nuclear lamina = protein mesh associated w inner face of inner nuclear membrane, provides mechanical support for nucleus, fibrous proteins called lamins form a 2d network (nuclear lamina) along inner surface of inner membrane to give it shape, regulates important cellular events e.g DNA replication and cell division, when a cell divides the nuclear envelope disappears transiently due to reversible disassembly of the nuclear lamina
• Chromosomes and chromatin = genetic material packed into chromosomes, DNA wrapped around histones into nucleosomes, nucleosomes fold up to form a chromatin fibre which is packed into chromosomes, DNA organises in nucleosomes that condense and form chromatin, chromatin divides into euchromatin and heterochromatin representing two different degrees of DNA condensation, during interphase chromatin is in least condensed state appears loosely distributed throughout the nucleus, two forms of heterochromatin – constitutive (always remains heterochromatic) and facultative (regions of euchromatin converted to heterochromatic state and are silences by histone deacetylation/DNA methylation)
• Nucleolus = site of rRNA synthesis, ribosome biogenesis

Question 3

Mitochondria

Answer 3

• One of biggest organelles in the cell
• Outer membrane (50% lipid, 50% protein) contains porins making membrane permeable to molecules = similar to outer membrane gram-neg bacteria
• Mitochondria originated by permanent enslavement of purple non-sulphur bacteria
• PMF and ETC take place in inner membrane
• SA of inner membrane greatly increased by large number of infoldings (cristae)
• Cristae = expand SA of inner mitochondrial membrane, enhances ability to generate ATP, intermembrane space appears continuous w the lumen of each crista, F0F1 complexes which synthesise ATP are intramembrane particles that protrude from the cristae and inner membrane into the matrix
• Matrix = contains mitochondrial DNA, ribosomes, granules, glycolysis and beta oxidation occurs

Question 4

Chloroplast

Answer 4

• Contain extensive internal system of interconnected membrane-limited sacs called thylakoids
• Thylakoids = flattened to form discs and stacks (grana)
• Thylakoid membranes contain photosynthetic pigments and ATP synthases

Question 5

Ribosome biogenesis

Answer 5

mRNA, tRNA and rRNA made in nucleus
Ribosomal proteins made in cytoplasm, but ribosomes are assembled in nucleus/nucleolus

Question 6

Nuclear transport

Answer 6

All proteins in nucleus are synthesised in cytoplasm and imported into the nucleus through nuclear pore complexes (NPC), such proteins contain a nuclear-localisation signal (NLS) that directs their selective transport into the nucleus, NLS bound by importins and is one or more short sequences of positively charged lysines or arginines exposed on the protein surface
Small GTPase Rna = regulate interaction of transport receptors with cellular cargo proteins
Ran = a monomeric G protein that acts as a molecular switch it exists in two conformations = Ran + GDP, Ran + GTP
High affinity binding of the GTP-bound form to the import receptors promotes cargo release, whereas its binding to export receptors stabilises their interaction w the cargo

Question 7

Nuclear import

Answer 7

1. Importin binds the cargo protein – cargo complex
2. Cargo complex diffuses through the nuclear pore
3. Ran-GTP interacts w importin causing a conformational change that decreases affinity for the cargo protein, releasing the cargo protein
4. Importin Ran-GTP complex is exported back in the cytoplasm
5. Ran-GTP is hydrolysed to Ran-GDP w the help of GTPase-activating protein – GAP
6. Ran-GDP is returned to the nucleoplasm by nuclear transport factor 2 (NTF2)
7. Guanine nucleotide-exchange factor (GEF) causes release of GSP and rebinding of GTP

Question 8

Nuclear export

Answer 8

To export macromolecules from the nucleus to the cytoplasm requires proteins w a nuclear exporting signal (NES) = short target peptide containing 4 hydrophobic residues on the protein
Proteins going back and forward have a NLS and NES
1. Exportin 1 binds to the NES of the cargo protein to be transported with Ran-GTP – cargo complex
2. Cargo complex diffuses through and NPC
3. GAP converts Ran-GTP to Ran-GDP
4. Conformational change in Ran leads to dissociation of the cargo complex
5. NES-containing cargo protein is released into the cytosol. Exportin 1 and Ran-GDP are transported back into the nucleus. Ran-GDP is transported by NTF2
6. GEF causes release of GDP and rebinding of GTP
Once processing of an mRNA is completed in the nucleus it is exported out of the nucleus into the cytoplasm before it can be translated into the encoded protein
mRNA remains associated w specific heterogenic nuclear ribonucleoproteins (hnRPS) in a messenger ribonuclear protein complex (mRNP)
mRNA exporter binds to mRNPs
The mRNP-m-RNA exporter complex diffuses through the nuclear pore

Question 9

Lysosomes

Answer 9

Only present in animal cells
Contain digestive enzymes
Responsible for degrading obsolete or worn out components of the cell
Destroy invading viruses and bacteria
Involves in programmed cell death (apoptosis) if cell is damaged beyond repair
Objects transported to lysosomes via:
Endocytosis = invagination of the cell membrane forms a vesicle that buds off and transports objects through the cell to the lysosomes
Phagocytosis = a form of endocytosis where whole cells and other large insoluble particles move from the cell surface to the lysosomes
Autophagy = worn-out organelles and bulk cytoplasm are surrounded by a membrane and delivered to the lysosomes

Question 10

What happens in lysosome

Answer 10

The lumen of the lysosome is acidic and contains hydrolytic enzymes that degrade polymers into their monomeric subunits
Enzymes include = nucleases degrade RNA and DNA into mononucleotides, proteases degrade proteins and peptides, phosphatases remove phosphate groups from mononucleotides, phospholipids and other compounds, other hydrolases degrade complex polysaccharides and glycolipids into smaller units

Question 11

Types of lysosome

Answer 11

Primary = roughly spherical, do not contain obvious particulate or membrane debris, formed by the fusion of golgi vesicles with late endosomes
Secondary = larger and irregularly shaped, result from fusion of primary lysosomes with other membrane-bound organelles and vesicles
Residual bodies = where indigestible materials pass outwardly and fuse with the plasma membrane
Auto-phagic vacuoles = a fusion of a primary lysosome with an autophagosome

Question 12

Vacuoles

Answer 12

Mostly in plants and fungi, most plant cells contain at least on membrane-limited internal vacuole
Two main types:
Storage = PSV, vacuolar membrane contains a variety of transport proteins that allows plant cells to accumulate and store water, ions and nutrients
Lytic = LV, lumen of a vacuole has an acidic pH and contains degradative enzyme

Question 13

How vacuole gets big and keeps shape

Answer 13

Vacuolar membrane (tonoplast) is permeable to water like most cellular membranes but poorly permeable to the small molecules stored within it
Solute concentration therefore much higher in the vacuole lumen than in the cytosol or extracellular fluids
Water mostly moves by osmotic flow into vacuoles
Influx of water creates turgor inside the cell
Pressure is balanced by mechanical resistance of cellulose-containing cell walls surround the cell

Question 14

Peroxisomes

Answer 14

Peroxisomes
All animal cells (except rbc) and many plant cells
Play key role in lipid metabolism and control of reactive oxygen species
In liver and kidney cells, degrades various toxic molecules that enter the bloodstream producing harmless products
Oxidise organic substances like fatty and amino acids

Question 15

Oxidation of fatty acids in peroxisomes

Answer 15

Yields no atp
Acetyl coa generated during degradation of fatty acids cannot be oxidised further
It is transported into cytosol for use in the synthesis of cholesterol and other metabolites
In process of oxidation, hydrogen peroxide is formed
H2o2 removed by catalases in peroxisomes

Question 16

Endoplasmic reticulum

Answer 16

Enclosed by largest membrane in eukaryotic cell
Has extensive network of closed, flattened membrane-bound sacs called cisternae
Important for synthesis of lipids, membrane proteins and secreted proteins
SER = lacks ribosomes
- Synthesis of lipids, fatty acids and phospholipids
- Steroid hormones
- Detoxification of harmful metabolic products
- Storage and metabolism of ions within cell
- Abundant in hepatocytes
- Enzymes in ser of liver modify or detoxify hydrophobic chemicals by chemically concerting them into water-soluble products that can be extracted from the body, high doses of such chemicals can result in large proliferation of ser in liver cells
RER = cytosolic face is studded w ribosomes
- Ribosomes bound synthesise certain membrane and organelle proteins and secreted proteins
- Newly made membrane proteins remain associated w the rer membrane
- Proteins to be secreted accumulate in lumen of er
1. Membrane and secreted proteins have signal sequences
2. Signal sequence directs ribosomes to the er membrane
3. Ribosomes dock on the translocon – a protein complex
4. Polypeptide synthesis starts
5. Signal peptide is cleaved
6. – 8 completed polypeptide leave the ribosome and folds into final conformation
Both regions of ER differ structurally and functionally but remain a continuous system

Question 17

Golgi complex

Answer 17

Main sorting hub of the secretory pathway
Receives proteins and membrane lipids first synthesised in er, complete processing (chemical modifications), package and transport to appropriate destinations

Question 18

Transport through Golgi

Answer 18

Consist of series of flattened membrane vesicles/sacs (cisternae) surrounded by number of spherical vesicles
Stack of golgi cisternae has 3 distinct regions = cis, medial, trans
Transport vesicles from rer fuse w cis membrane of golgi
Proteins in vesicles move from cis to medial and finally to trans of golgi
Vesicles bud off the trans-golgi membranes
Vesicles containing proteins move to the cell surface and others move to lysosomes

Question 19

Major protein-sorting pathways in euk cells

Answer 19

Main sites of protein sorting = er, golgi, vesicles
Each site proteins sorted into separate vesicles based on sorting signals and cellular machineries that recognise those signals
Proteins synthesised in er have signals that directs the sorting system

Question 20

Secretory pathways

Answer 20

All nuclear-encoded mRNAs are translated on cytosolic ribosomes
Ribosomes synthesising secrete nascent proteins are directed to the rer by an er signal sequence (1 and 2)
Translation is completed on er. Proteins can move via transport vesicles to golgi (3)
Further sorting delivers proteins either to plasma membrane or to lysosomes (4a, 4b)

Question 21

Secretory cell

Answer 21

Secreted proteins after synthesis on the ribosomes of rer moved to lumen of rer
1. Transport vesicles bud off and carry these proteins to golgi
2. Protiens are conc and packaged into immature secretory vesicles
3. These vesicles coalesce to form larger mature secretory vesicles
4. Vesicles accumulated under apical surface of cell, fuse with plasma membrane releasing their contents (exocytosis)
Release is response to appropriate hormone or nerve stimulation

Question 22

Non-secretory pathways

Answer 22

Synthesis of proteins lacking an er signal sequence is completed on free ribosomes (1)
Proteins w no targeting sequence released into cytosol and remain there (2)
Proteins w an organelle-specific targeting sequence first released into cytosol but then imported into mitochondria, chloroplasts, peroxisomes, or nucleus (3-6)
Majority of proteins located in mitochondria and chloroplasts are encoded by genes in the nucleus and imported into the organelles after synthesis in cytosol
But proteins encoded by mitochondrial or chloroplast DNA are synthesised on ribosomes within these organelles then directed to the correct sub-compartment
e.g mitochondrial proteins for oxidative phosphorylation

Question 23

Targeting signals

Answer 23

Precursor proteins synthesised in cytosol destined for mitochondria chloroplasts contain specific uptake-targeting sequences that bind receptors on organelle surface
Short stretches of amino acids

Question 24

Cytoskeleton

Answer 24

Dynamic network of soluble proteins packing the cell interior
Provides number of functions but primary function is to provide internal structure e.g shape and mechanical resistance
Cytosol of eukaryotic cell contains 3 types of filaments
- Actin filaments = small diameter, microfilaments, twisted two-stranded structure. Each strand made of 2 monomeric actin subinuts
- Intermediate = intermediate diameter, assembled from different proteins depending on the cell
- Microtubules= large diameter, consist of hollow tube-like strucutres made of dimetric subunits of alpha and beta tubulin
= polymers comprising a chain of monomer protein subunits

Question 25

Cytoskeleton filament cellular localisation

Answer 25

Most eukaryotic cells contain all 3 filaments, located in different places in different cells
In absorptive epithelial cells in lumen of gut:
- Actin filaments = apical region
- Microtubules = oriented w the long axis of cell
- Intermediate = conc along cell periphery e.g specialised junctions w neighbouring cells or lining the nuclear membrane
Sensory hairs of inner ear:
- Actin = conc in apical region in particular stereocilia
- Microtubules = conc in apical region and kinocilia but also throughout cells
Cytoskeletal filaments are organised into bundles and networks
In bundles filament are closely packed in parallel arrays, in network filaments crisscross
Cytoskeletal components usually attached to plasma membrane proteins and form skeleton that helps support plasma membrane

Question 26

Functions of cytoskeleton

Answer 26

Confers structure
Involved in cellular movement
Intracellular movement – e.g cytoplasm streaming (flow of cytoplasm inside cell, driven by forces from cytoskeleton, function is to speed up transport of molecules and organelles around cell), transport of vesicles or separation chromosomes in cell division, transport of chloroplasts
Whole cell movement – e.g migration, hosts defences, changes in morphology like muscle contraction, nerve axon elongation and cell protrusion and constriction
Cell movement requires arrangement of microfilaments or microtubule and/or motor proteins
Intermediate fibres are not involved in cell movement, instead they play role in cell adhesion
Cell movements require ATP and proteins that convert the energy stored in ATP into motion
Cells evolved 2 basic mechanisms for generating movement
- One entails the assembly and disassembly of microfilaments and microtubules; responsible for many changes in cell shape
- Other requires a special class of enzymes called motor proteins (myosins, kinesisns and dyneins)
Few movements require both the action of motor proteins and cytoskeleton rearrangements

Question 27

Microfilaments

Answer 27

Made of actin = most abundant intracellular protein in most euk cells
Actin
- Encoded by large highly conserved gene family
- Single cell organisms 1-2 actin genes
- Multicellular organisms multiple genes
- Different genes in the family have different functions and are found in different cells
- Globular monomer called g-actin, separated into 2 lobes by deep cleft, lobes and cleft compose the ATPase fold = site where ATP and mg ion bound, cleft acts as hinge that allows lobes to flex relative to each other, g-actin monomer can assemble into chain to form filamentous polymer f-actin which can depolymerise to form g-actin
- In filament the subunits are organised as helix
- Filaments polarised w one end (-) end containing exposed ATP binding site
- Filaments organised into bundles and networks by variety of bivalent cross-linking proteins. When cross-linked by short protein actin filaments pack side by side to form bundle, long cross-linking proteins such as filamin are flexible and can cross-link actin filaments into network
Actin assembly
- Within cells actin cytoskeleton dynamic w filaments able to grow and shrink rapidly
- Filaments grow considerably faster at + end than – end
- Initial nucleation process acts as a focal point
1. In initial nucleation phase ATP G-actin monomers slowly form stable complexes of actin
2. Nuclei rapidly elongated in second phase by addition of subunits to both ends of filament
3. After their incorporation into filament subunits slowly hydrolyse ATP and becomes stable ATP-F actin
Actin treadmilling – actin subunits can flow through the filaments by attaching preferentially to the + end and dissociating preferentially from -end of filament, in this situation the length of the filament remains constant
Control of actin polymerisation – regulated by proteins that bind g-actin
- Thymosin β4 inhibits actin assembly.
- Profilin promotes actin assembly.
- Profilin binds to the G-actin coupled to ADP and catalyses the exchange of ADP with ATP
- The ATP-G-actin-profilin complex can be linked to the plus end of the filament and Prolifin dissociates
Polymerisation of actin causes changes in cell shape
Myosin powered cell moments – myosin is large superfamily of motor proteins that bind actin filaments or microfilaments. Responsible for
- Vesicular trafficking
- Cytoplasmic streaming
- Muscle contraction
- Cytokinesis
All myosins consist of one or two heavy chains and several light chains, head, neck and tail domain organisation
- Myosin head domains have ATPase activity. ATP hydrolysis allows myosin movement along an actin filament.
- Role of a myosin is related to its tail domain. Tail domains of myosins I, V, VI, and XI bind the plasma membrane or the membranes of intracellular organelles.
- Myosin II dimers associate to form bipolar thick filaments (muscle contraction): close packing of myosin II molecules into thick filaments allows many myosin head domains to interact simultaneously with actin filaments.
- In non-muscle cells the myosin II actin bundle are referred to as stress fibres and are responsible for cell and tissue migration and adhesion.
ATP hydrolysis allows myosin movement along actin filament
- ATP hydrolysed resulting in myosin attaching to actin subunit
- Phosphate released causing head to change position pulling myofilament
- ADP released and ATP attaches resulting in detachment of myosin from actin
- Repeats
Cell movement – initiated by formation of large, broad membrane protrusion at leading edge of a cell called lamellipodium in vertebrate cells
Microfilament based cell movement occurs when
1. Actin filaments at leading edge are rapidly cross-linked into bundles and networks
2. Lamellipodium adheres to substratum moves forward then the trailing edge de-adheres

Question 28

Microtubules

Answer 28

Tubulin dimer of monomers alpha and beta tubulin
Function:
- Maintenance of cell shape
- Cell motility
- Chromosome movement
- Organelle movement
Structure:
- Alpha tubulin monomer bound to GTP which is nonexchangeable
- Beta tubulin monomer bound to GDP which is exchangeable w GTP
- GTP or GDP bound to beta tubulin modulates the addition of tubulin subunits
Assembly:
1. Protofilaments assembled from alpha beta tubulin subunits
2. Protofilaments associate to form the wall of the microtubule
3. Addition of more subunits to the ends of the protofilaments elongates the microtubule
Like actin filaments microtubules have a distinct polarity. Tubulin polymerizes end to end, with the β-subunits of one tubulin dimer contacting the α-subunits of the next dimer. In a protofilament, one end will have the α-subunits (-) exposed while the other end will have the β-subunits (+) exposed. As the protofilaments bundle parallel to one another with the same polarity, in a microtubule, there is one end, the (+) end, with only β-subunits exposed, while the other end, the (−) end, has only α-subunits exposed. While microtubule elongation can occur at both the (+) and (−) ends, it is significantly more rapid at the (+) end.
Microtubule forms at a central site, the microtubule-organizing centre (MTOC), contains a Ɣ-tubulin which is the major site of microtubule nucleation.
Animal cells - MTOC is usually a centrosome, sometimes but not always contains a pair of centrioles.
Fungi - MTOC is called the spindle pole
Plants - nuclear envelope appears to act as MTOC
Kinesin and dynein powered cell movements:
- Microtubules function as tracks in the intracellular transport of various types of “cargo.”
- Two families of motor proteins, kinesins and dyneins, mediate transport.
- Examples of kinesins and dyneins transport include metaphase during mitosis and meiosis and beating of cilia and flagella.
Kinesins = composed of heavy chains and two light chains, most are + end directed motors (move to + end), divided into cytosolic and motitic kinesins based on cargo they transport
- Cytosolic = transport of organelles and vesicles
- Mitotic = participate in spindle assembly and chromosome segregation in cell division
Dyneins = composed of 2 or 3 heavy chains complexes and several intermediate and light chains Dynein cannot mediate cargo transport alone requires dynactin, that links vesicles and chromosomes to the dynein light chains. Cytosolic dyneins are (-) End–Directed motor proteins that bind cargo through dynactin.
- Cytosolic = role in movement of vesicles and chromosomes
- Anoxemal = responsible for beating of cilia and flagella

Question 29

Intermediate filaments

Answer 29

Found in nearly all animals but not plants and fungi.
Fibrous proteins supercoiled into thicker cables
protein subunits depend on cell type
Associated with the nuclear and plasma membrane principal function is structural.
Formation of nuclear lamina
Not involved in cell movement.

Question 30

Ribosomes

Answer 30

Site of protein synthesis in eukaryotes & prokaryotes.
Thousands to millions per cell depending on activity.
Found in cytosol, rER, mitochondria & chloroplasts.
2 Subunits made up of RNA molecules & proteins.
Ribosomes are NOT an organelles are not bound by a membrane and are much smaller than other organelles

Question 31

Cell wall

Answer 31

Structural layer that can be tough, flexible, and sometimes rigid.

Provides structural support and protection, and acts as a filtering mechanism.

Absent in animals, present in most other eukaryotes

Major function prevent over-expansion of the cell when water enters.

Composition depends on the cell type
e.g. plants have three layers - primary layer contains cellulose, hemicellulose and pectin.
Middle lamella lies between cells: Calcium and Magnesium pectates
Primary cell wall consists of:
Hemicellulose: polysaccharides of 5-carbon (xylose and arabinose) and 6-carbon (glucose, mannose, galactose and rhamnose)
Pectin: polysaccharides of mostly galacturonic acid
Cellulose microfibrils provide strength