Class notes on organelles and the enomembrane system
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Tour of the Cell 1 (Ch. 6) Dead White Men Who Discovered (and were made of) Cells: Anton Van Leeuwenhoek Robert Hooke Where the Magic Happened Microscopy is certainly a Perspective Changer (sorry in advance) Light Microscopy TECHNIQUE (a) RESULTS Brightfield (unstained specimen). Passes light directly through specimen. Unless cell is naturally pigmented or artificially stained, image has little contrast. [Parts (a)–(d) show a human cheek epithelial cell.] 50 µm (b) Brightfield (stained specimen). Staining with various dyes enhances contrast, but most staining procedures require that cells be fixed (preserved). (c) Phase-contrast. Enhances contrast in unstained cells by amplifying variations in density within specimen; especially useful for examining living, unpigmented cells. (d) Differential-interference-contrast (Nomarski). Like phase-contrast microscopy, it uses optical modifications to exaggerate differences in density, making the image appear almost 3D. (e) Fluorescence. Shows the locations of specific molecules in the cell by tagging the molecules with fluorescent dyes or antibodies. These fluorescent substances absorb ultraviolet radiation and emit visible light, as shown here in a cell from an artery. 50 µm (f) Confocal. Uses lasers and special optics for “optical sectioning” of fluorescently-stained specimens. Only a single plane of focus is illuminated; out-of-focus fluorescence above and below the plane is subtracted by a computer. A sharp image results, as seen in stained nervous tissue (top), where nerve cells are green, support cells are red, and regions of overlap are yellow. A standard fluorescence micrograph (bottom) of this relatively thick tissue is blurry. 50 µm Electron Microscopy TECHNIQUE (a) Scanning electron microscopy (SEM). Micrographs taken with a scanning electron microscope show a 3D image of the surface of a specimen. This SEM shows the surface of a cell from a rabbit trachea (windpipe) covered with motile organelles called cilia. Beating of the cilia helps move inhaled debris upward toward the throat. RESULTS Cilia Longitudinal section of cilium (b) Transmission electron microscopy (TEM). A transmission electron microscope profiles a thin section of a specimen. Here we see a section through a tracheal cell, revealing its ultrastructure. In preparing the TEM, some cilia were cut along their lengths, creating longitudinal sections, while other cilia were cut straight across, creating cross sections. 1 µm Cross section of cilium 1 µm Cell Fractionation APPLICATION Cell fractionation is used to isolate (fractionate) cell components, based on size and density. TECHNIQUE First, cells are homogenized in a blender to break them up. The resulting mixture (cell homogenate) is then centrifuged at various speeds and durations to fractionate the cell components, forming a series of pellets. RESULTS In the original experiments, the researchers used microscopy to identify the organelles in each pellet, establishing a baseline for further experiments. In the next series of experiments, researchers used biochemical methods to determine the metabolic functions associated with each type of organelle. Researchers currently use cell fractionation to isolate particular organelles in order to study further details of their function. Homogenization Tissue cells 1000 g Homogenate (1000 times the force of gravity) Differential centrifugation 10 min Supernatant poured into next tube 20,000 g 20 min Pellet rich in nuclei and cellular debris 80,000 g 60 min 150,000 g 3 hr Pellet rich in mitochondria (and chloroplasts if cells are from a Pellet rich in plant) “microsomes” (pieces of plasma membranes and Pellet rich in cells’ internal ribosomes membranes) The size range of cells 10 m Human height Length of some nerve and muscle cells Unaided eye 1m 0.1 m Chicken egg 1 cm Frog egg Most plant and animal cells 10 µm 1 µm 100 nm nucleus Nucleus Most bacteria Most bacteria Mitochondrion Smallest bacteria Viruses Ribosomes 10 nm Electron microscope 100 µm Light microscope 1 mm Proteins Lipids 1 nm 0.1 nm Small molecules Atoms Measurements 1 centimeter (cm) = 102 meter (m) = 0.4 inch 1 millimeter (mm) = 10–3 m 1 micrometer (µm) = 10–3 mm = 106 m 1 nanometer (nm) = 10–3 µm = 10 9 m Comparing the size of a virus, a bacterium, and an animal cell Virus Bacterium Animal cell While we’re on the topic of size... Animal cell nucleus 0.25 m Why Cells Are So Small: The SA:V Ratio Surface area increases while total volume remains constant 5 1 1 Total surface area (height width number of sides number of boxes) 6 150 750 Total volume (height width length number of boxes) 1 125 125 Surface-to-volume ratio (surface area volume) 6 12 6 Prokaryote bacteria cells Types of cells - no organelles - organelles Eukaryote animal cells Eukaryote plant cells Why organelles? • Specialized structures – specialized functions • Containers – Compartments = different local environments • pH, concentration differences – distinct & incompatible functions • lysosome & its digestive enzymes • Membranes as sites for chemical reactions – Unique lipids & proteins – embedded enzymes & reaction centers • chloroplasts & mitochondria Cells gotta work to live! – make proteins • proteins control every cell function – make energy • for daily life • for growth – make more cells • growth • repair • renewal Proteins do all the work! proteins cells DNA organism Repeat after me… Proteins do all the work! Cell functions • Building proteins – copy DNA – DNA -> RNA – build proteins – process proteins • Folding, modifying –Remove amino acids –Add molecules (e.g. glycoproteins) – address & transport proteins Protein Synthesis • Organelles involved – nucleus – ribosomes – endoplasmic reticulum (ER) – Golgi apparatus – vesicles The Protein Assembly Line nucleus ribosome ER Golgi apparatus The Endomembrane System vesicles Nucleus • Function DNA chromosome – protects DNA • Structure histone protein – nuclear envelope • double membrane • membrane fused in spots to create pores nuclear pores What kind of molecules need to pass through? nuclear pore nucleolus nuclear envelope 1 nuclear membrane production of mRNA from DNA in nucleus DNA Nucleus mRNA 2 nuclear pore mRNA travels from nucleus to ribosome in cytoplasm through nuclear pore small ribosomal subunit mRNA large ribosomal subunit cytoplasm Nucleolus • Function – ribosome production • build ribosome subunits from rRNA & proteins • Ribosome assembly is completed in cytoplasm large subunit small subunit rRNA & proteins ribosome nucleolus Ribosomes large subunit • Function – protein production • Structure – rRNA & protein – 2 subunits combine small subunit 0.08m Ribosomes Rough ER Smooth ER Types of Ribosomes • Free ribosomes – suspended in cytosol – synthesize proteins that stay in cytosol • Bound ribosomes – attached to endoplasmic reticulum – synthesize proteins for export or membranes membrane proteins Endoplasmic Reticulum • Function – processes proteins – manufactures membrane – synthesis & hydrolysis of many compounds • Structure – membrane connected to nuclear envelope & extends throughout cell Types of ER rough smooth Smooth ER function • Membrane production • Metabolic processes – Lipid Synthesis – Glycogen hydrolysis (in liver) – Drug detoxification (in liver) Membrane Factory • Build new membrane – synthesize phospholipids – ER membrane expands • buds off & transfers to other parts of cell. Rough ER function • Produces proteins for export out of cell – protein secreting cells – packaged into transport vesicles for export Which cells have lot of rough ER? Synthesizing proteins cisternal space polypeptide signal sequence ribosome ribosome mRNA membrane of endoplasmic reticulum cytoplasm Golgi Apparatus • Function – finishes, sorts, tags & ships products • like “UPS shipping department” – ships products in vesicles • membrane sacs • “UPS trucks” secretory vesicles Which cells have lots of Golgi? transport vesicles Golgi Apparatus Vesicle transport protein vesicle budding from rough ER ribosome migrating transport vesicle fusion of vesicle with Golgi apparatus Putting it together… The Endomembrane System nucleus nuclear pore cell membrane protein secreted rough ER ribosome vesicle proteins smooth ER transport vesicle cytoplasm Golgi apparatus Any Questions!! Can I offer you something in A Computer Animation? Or perhaps something more in a silly rap song?!? Review Questions 1.. In which cell would you expect to find the most smooth endoplasmic reticulum? A. Muscle cell in the thigh muscle of a long-distance runner B. Pancreatic cell that manufactures digestive enzymes C. Macrophage (white blood cell) that engulfs bacteria D. Epithelial cells lining the digestive tract E. Ovarian cell that produces estrogen (a steroid hormone) 2. In which cell would you expect to find the most bound ribosomes? A. Muscle cell in the thigh muscle of a long-distance runner B. Pancreatic cell that manufactures digestive enzymes C. Macrophage (white blood cell) that engulfs bacteria D. Epithelial cells lining the digestive tract E. Ovarian cell that produces estrogen (a steroid hormone) 3. Of the following, which is probably the most common route for membrane flow in the endomembrane system? A. Golgi → lysosome → ER → plasma membrane B. tonoplast → plasma membrane → nuclear envelope → smooth ER C. nuclear envelope → lysosome → Golgi → plasma membrane D. rough ER → vesicles → Golgi → plasma membrane E. ER → chloroplasts → mitochondrion → cell membrane
