Week 3

Cytoskeleton: a network of fibers that organizes the structure and activities of the cell

• Supports the cell and maintains its shape

• Anchors organelles in specific locations

• Provides a ‘monorail system’ for vesicles to move on and reach their destination (platform and directionality)

• Used for cell motility and contractility

Three cytoskeleton structures: 

• Actin microfilament: subunit is actin

  • Thinner, dynamic (Can serve as tracks for vesicle transport)

• Intermediate filaments: ropes of protein strands many different types

  • Thinner, more permanent

• Microtubules: polymers of globular protein subunits; subunit = tubulin dimer

  • Large & hollow, dynamic (Can serve as tracks for vesicle transport)

Actin Filament/ Microtubules: dynamic and polarized

• Constant assembly (“polymerization”) and disassembly (“depolymerization”) of globular subunits

• They have structural polarity via +/- ends (Subunits can add to either end, but more rapid at “+” end)

Intermediate Filament:

• Assembly: Wrapping of dimers into tetramer and tetramers into octamers and so on, generates a rope-like filament with tremendous tensile strength and no polarity

• Keratin:assembles into filaments in epithelial cells (skin)

• Desmosomes: connects keratin filaments between cells to enhance tissue rigidity and flexibility

  • Stretching sheet of cells with intermediate filaments keep cells intact, without causes cells to rupture

• Nuclear lamins form a dense mesh inside the nuclear envelope that anchors chromosomes, defines the shape of the nucleus, and stabilizes the envelope. By controlling the interactions between these lamins, many eukaryotic cells will break down and reform the nuclear envelope during cell division.

• Various types of intermediate filaments are found throughout the cytoplasm of the cell.

  • Projects from the nucleus to hold it in place or runs parallel to the cell surface and interact with proteins embedded in the plasma membrane

• To summarize, intermediate filaments function like a flexible internal scaffolding to help secure the shape and stability of the cell.

Actin Filament: Flexible filament composed of two protofilaments

• Two strands coiled around one another; Pointed end is -, barbed end is +

• Myosin attaches to + end and hydrolyzes ATP to ADP, and then contracts to pull itself along the actin filament, releasing ADP + P. These changes cause the actin and myosin to slide past each other. After repeated rounds of this attachment and contraction cycle, the myosin gradually moves toward the plus end of the actin filament

Microtubules: 

• Made up of 2 subunits: a-tubulin (-) and b-tubulin (+) to form dimers

• Polymerizes in head to tail fashion via noncovalent interactions to form thin filaments called protofilaments which interact with one another to form hollow tubes

• Microtubule organizing center (MTOC): organization of microtubules, plus ends grow out, plants have many but animals/fungus have one near nucleus (centrosome/centrioles)

• Involved in mitosis/meiosis anaphase to pull away

• Squid experiment: observed large nerve cell in squid axon, found vesicle transport still occurred when cytoplasm was removed from axon, noticed vesicles move along microtubule tracks

  • Additional research found that vesicle movement requires ATP and kinesin (protein) converts the energy into movement towards plus end (dynein to minus end)

  • Kinesin composed of: head section, tail of small polypeptides, stalk connecting the two (all together makes one large subunit, two large subunits make most of kinesin)

• Motor proteins: carries cargo vesicles/organelles bound to tail of motor transported across cell

• Nuclear lamins, which make up the nuclear lamina layer introduced in Section 7.4, also are intermediate filaments. Nuclear lamins form a dense mesh inside the nuclear envelope that anchors chromosomes, defines the shape of the nucleus, and stabilizes the envelope. By controlling the interactions between these lamins, many eukaryotic cells will break down and reform the nuclear envelope during cell division.

Flagella/Cilia:

• A prokaryotic flagellum consists of a single helical rod made of flagellin (in bacteria) or other types of proteins (in archaea); a eukaryotic flagellum consists of several microtubules constructed from tubulin dimers.

• Prokaryotic flagella move the cell by rotating like a ship’s propeller; eukaryotic flagella move the cell by undulating—they whip back and forth.

• Eukaryotic flagella are surrounded by the plasma membrane; prokaryotic flagella are not.

Prokaryotes:

• No Nucleus 

• One circular chromosomes supercoiled in nucleoid

• Plasmids: circular supercoiled DNA (not always expressed)

• Cytoskeleton

• Organelles

• Internal membrane complexes (can increase SA for photosynthesis)

• Moves via flagellum (assembled from many diff protein at cell surface) vs fimbriae (needle like projection extending from plasma membrane and attached to other cell surface)

Cytoplasm is hypertonic: 

• Causes water to enter cell via osmosis increasing cell volume

• Stiff cell wall prevents this

Eukaryotes:organelles compartmentalized which allows incompatible chemical reactions to be separated, making them more efficient

• Nucleus: center for information storage + processing

  • Enclosed by nuclear envelop[e (double membrane) 

  • Nuclear lamin stiffens it to keep shape

  • Nucleolus: manufactures/processes RNA molecules into RNA

• Ribosomes: scattered free in cytosol

• Rough/smooth ER: rough has ribosomes attached to surface to translate mRNA to proteins while smooth synthesizes lipids

• Golgi apparatus: cis (receives vesicles or cargo from ER) to trans (delivers to other organelles)

• Lysosome: animal cells, hydrolytic enzymes that breaks down macromolecules in acidic environment

• Vacuoles: storage deposit for water/nutrient

• Peroxisomes: site of oxidation reaction originates when empty vesicles from ER filled with peroxisome specific enzymes from cytosol

• Mitochondria: enzymes for ATP synthesis, fusion/fission, has mDNA

• Chloroplast: third membrane with thylakoids in interior arranged in interconnected stacks called grana with stroma (fluid filled space around grana) containing enzymes to convert chemical energy to sugar

Endomembrane system: 

• Nuclear envelope: separates nucleus from cell but has openings called nuclear pore complex which extends through inner/outer nuclear membrane + connects inside of nucleus with cytosol

  • DNA does not leave through pores but RNA molecules produced in nucleus can

• Nucleoplasmin experiment: when nucleoplasmin is injected into cytosol, they quickly move into the nucleus. Used proteases to cut nucleoplasmin into core/fail, results showed core remains in cytosol while tail moves into nucleus. 

  • Hypothesis: nuclear proteins contain zip code (nuclear localization signal) to pass into nucleus)

• Secretory pathway hypothesis:

  • Protein enters ER while being synthesized by ribosomes + further processed

  • Proteins exit er via vesicle

  • Protein enters golgi apparatus (cis) then exists (trans) via vesicle

  • Protein vesicle travels to plasma membrane to be secreted 

• Pulse-chase experiment: 

  • Cells exposed to high concentration of radioactive amino acids to proteins synthesized will be radio-labelled

  • Stops exposure by washing away radioactive amino acids to observe movement of the labelled

  • Observed proteins inside rough ER then golgi then secreted

• Signal Hypothesis: 

  • ER signal sequence synthesized by ribosome

  • ER signal sequence binds to signal recognition particle (SRP) and halts synthesis

  • SRP binds to receptor in ER membrane

  • SRP released and protein enters ER via translocon

  • ER signal sequence removed + protein synthesis continues

• In golgi apparatus:

  • Proteins have diff tags for diff destinations

  • Proteins sorted in trans-golgi binding to diff receptors

  • Transport vesicles bud off trans-golgi to respective destinations

  • Transport vesicles attach/fuse at destinations

• Recycling via lysosome: 

  • Receptor-mediated endocytosis forms vesicle to deliver cargo (macromolecules0 to early endosome (acidifies then matures into late endosome then lysosome)

  • Phagocytosis: brings in food/smaller molecules via phagosome which fuses with lysosome to digest its contents

  • Autophagy: encloses damaged organelle forming autophagosome which delivers to lysosome for digestion

  • Lysosome releases small molecules from digestion into cytosol

Ribosomes: 

• If ribosome is free, the protein will end up in cytosol

• If ribosome is on the ER, the protein will be in the lumen of the endomembrane system of cell 

Snares: proteins that help transport vesicles fuse with plasma membrane 

Endocytosis: can be non-specific (phago/pinocytosis) or specific to cargo (has receptors)

• Clathrin mediated endocytosis: clathrin assemble around plasma membrane creating clathrin-coated pit which pinches off and forms clathrin-coated vesicle, clathrin coat falls off to be reused so vesicle can fuse at destination

Proteins made by Free Ribosomes either…

• stay in cytoplasm 

• get selectively imported into organelles that are not part of the dynamic ER / Golgi system connected by vesicles

Proteins made by ER-docked Ribosomes get sorted to different destinations: 

• reside in ER 

• reside in Golgi 

• reside in lysosomes 

• get exocytosed out of the cell