3-Ch.4_Cell Biology of Prokaryotes and Eukaryotes_Micro50_LMC_F24.pdf

Ch.4 – Cell Structure o cell theory and cell size o eukaryotic vs. prokaryotic o prokaryotic cell structure o prokaryotic cell walls o bacterial shapes o preparing samples for light microscopy smears and stains o eukaryotic cell structure  EVERY form of life is……. Human...

Ch.4 – Cell Structure o cell theory and cell size o eukaryotic vs. prokaryotic o prokaryotic cell structure o prokaryotic cell walls o bacterial shapes o preparing samples for light microscopy smears and stains o eukaryotic cell structure  EVERY form of life is……. Human embryonic stem cell a single cell or composed of cells life starts with the cell any exceptions?..... a virus? virus REQUIRES a cells machinery to reproduce virus does not metabolize or use energy  The Cell Theory all organisms are composed of one or more cells. cells are the basic living unit of structure and function in organisms. all cells come only from other cells. Prokaryotic and eukaryotic cell images single cell organisms 2 cell types in life: 1. Prokaryotic o Bacteria o Archea (“extreme” bacteria) 2. Eukaryotic o Protist Protozoa (amoeba) Algae o Plants o Fungus Prokaryotic vs. eukaryotic cell structure comparison o Animals multi-cell organisms  Cell Size cells are small! (10-100um) being small an advantage for a cell…especially multicellular organisms easier to obtain nutrients such as glucose and expel waste such as CO2 small cell creates a high membrane surface area / cell volume ratio Why are cells so small? Size scale of living things – from atoms to whales  Surface Area / Volume Ratio being small creates a high surface area / volume ratio lots of surface area and little volume Surface Area to Volume Ratio example Increases with decreasing size as a cell increases its volume, the proportionate surface area DECREASES! all 3 have the same volume, but group on the right has 4 times the surface area  Ch.4 – Cell Structure o cell theory and cell size o eukaryotic vs. prokaryotic o prokaryotic cell structure o prokaryotic cell walls o bacterial shapes o preparing samples for light microscopy smears and stains o eukaryotic cell structure  Organizing Life: life’s family tree Three Domains of Life: 1. Bacteria Prokaryotes 2. Archaea 3. Eukarya - Eukaryotes Protist, plant, animal and fungi Cast of Microbiology: Bacteria Prokaryotes Archaea Protists Algae and Protozoa Eukaryotes Fungi Helminths Tree of life diagram Viruses “Non-Living” Evolution of eukaryotic kingdoms from bacteria  Prokaryotic vs. eukaryotic cell structure comparison Prokaryotes vs. Eukaryotes 2 cell types: Prokaryotic and Eukaryotic o Prokaryote Bacteria and archaea From Greek word for pre-nucleus o Eukaryote Prokaryotic: Eukaryotic: smaller larger Protist, fungi, plants, and animals simpler complex From Greek words for true nucleus unicellular uni or multicellular no nucleus or organelles nucleus and organelles binary fission mitosis and meiosis  Comparing Prokaryotic and Eukaryotic Cells: an overview Prokaryotic vs. eukaryotic cell structure comparison Prokaryote 1.One circular chromosome, no histones, not membrane enclosed 2.No organelles 3.Cell Wall Bacteria: peptidoglycan Archaea: pseudomurein 4.Divides by binary fission Eukaryote Histone: Protein that DNA wraps around to condense 1.Paired chromosomes, histones, within nucleus 2.Organelles 3.Cell Wall - Polysaccharide, when present Plants: cellulose Fungi: chitin 4.Divides by mitosis Histone proteins of eukaryotic chromosomes  Comparing Prokaryotic and Eukaryotic Cells: An Overview Characteristic: Prokaryotic Eukaryotic One circular chromosome Paired chromosomes Chromosome Nucleus NO YES Histones NO YES Organelles NO YES Cell Division Binary Fission Mitosis Bacteria – Peptidoglycan Polysacchardie if present Cell Wall Archaea - Pseudomuerin (Cellulose or Chitin)  Ch.4 – Cell Structure o cell theory and cell size o eukaryotic vs. prokaryotic o prokaryotic cell structure o prokaryotic cell walls o bacterial shapes o preparing samples for light microscopy smears and stains o eukaryotic cell structure  Prokaryotes: Bacteria and Archaea Escherichia coli Methanococcus bacteria jannaschii archaea o Unicellular and microscopic - size of cell organelle! o No nucleus; single circular chromosome o Have cell wall Bacteria cell wall made of peptidoglycan Archaea cell wall made of pseudomeurin  Bacterial Cell Structure Nucleoid: nuclear “area” where chromosome located Chromosome: bacteria have a single, circular chromosome Plasma membrane: phospholipid bilayer like eukaryotic cells Cell wall: made of “peptidoglycan”. Protection, filter, shape and prevent lysing from osmotic pressure Several different bacterial cell wall “forms” Capsule: additional protection made of polysaccharide Similar to cellulose plant cell wall Not all bacteria have Bacterial cell structure Fimbriae: hairlike appendages that allow for attachment  Bacterial Cell Structure Glycocalyx: Flagella: External to cell wall – viscous / gelatinous Filamentous appendages external to cell Made of polysaccharide and/or polypeptide Protein “tail” used for locomotion Two types o capsule: neatly organized and firmly attached Propel bacteria o slime layer: unorganized and loose Made of protein flagellin Contributes to bacteria's virulence - ability to cause Can have more than one disease Capsule Slime Layer Capsulated bacteria – capsule stain bacteria cell showing flagella  Bacterial flagella structure Bacterial Flagella Structure: Three parts 1. Filament: outermost region 2. Hook: attaches to the filament 3. Basal body: consists of rod and pairs of rings Anchors flagellum to the cell wall and membrane Bacteria can have more than one flagella: o monotrichous - one o amphitrichous - one flagellum at each end o lophotrichous - more than 2 flagella at one or both ends Different arrangements of bacterial flagella o peritrichous - uniform flagella around bacteria  Axial filaments Pili: o Aid in motility - gliding and twitching motility o Function to attach to substrate or other cells Axial filament of Spirochetes o Conjugation pili – Transfer DNA cell to cell Bacterial structure indicating sex pili an endoflagella – internal flagella In spirochetes electron micrograph of bacterial Anchored at one end of a cell conjugation pili Rotation causes cell to move  Motile Cells Rotate flagella to move One direction to run (straight trajectory) Reverse direction to tumble (re-orientation) Move toward or away from stimuli (taxis) Bacterial motility – tumble and roll – reversing flagella direction o Flagella proteins are H antigens Letters indicate antigen type: examples, H is flagella; O antigen is membrane protein e.g., E. coli O157 : H7 – H7 refers to flagella H antigen Can differ within bacterial species Help to distinguish between serovars o Serovar – antigen variations (serotypes) within one species e.g. Salmonella enterica is divided into 2400 serovars VIDEO: microbial motility - Tumble and Roll VIDEO: Bacteria motility: Tumble and Roll  Bacterial Endospores: “tough” protein coat to “protect” DNA Chromosome and essential proteins wrapped in very strong protein coat Form in “bad” environments o Low nutrients, unfavorable temp etc.. Resistant to desiccation, heat, chemicals Only some bacterial species have o Bacillus, Clostridium species Sporulation: Endospore formation Germination: Return to vegetative state Plasma Membrane: Life’s Border A “floating” phospholipid bilayer Two layers of phospholipids, oriented… Hydrophobic “tails” IN Hydrophilic “heads” OUT o many proteins interspersed in membrane Cell membrane indicating phospholipid bilayer Functions: control entry/exit of material protect from outside environment sending/receiving signals 4 major components of plasma membrane “Fluid Mosaic” model of membrane 1. Phospholipid bilayer Components “float” and “move” along 2. Proteins membrane 3. Cholesterol Membrane held together by hydrogen bonds 4. Glycocalyx “movement” across plasma membrane 1. Simple diffusion 2. Facilitated diffusion 3. Active transport 4. Osmosis Cell plasma membrane  Passive Transport 2. Facilitated Diffusion o Passage of hydrophilic (polar) molecules 1. Simple Diffusion o With help of transport proteins o Small hydrophobic (non-polar) molecules o Down concentration gradient (H to L) o Freely pass by diffusion through phospholipid o Some open and close bilayer example: glucose o Down concentration gradient (H to L) o Some open all the time example: gasses: O2 and CO2 pass freely example: Aquaporin (H O channel) 2 Simple Diffusion: Examples of facilitated O2 into cells membrane diffusion CO2 out of cells Passive diffusion of gas through plasma membrane  Passive processes Types of passive membrane diffusion  Active Transport: o Transport against concentration gradient Active transport through membrane ATP structure from low to high concentration requires energy expenditure Requires Energy......ATP! always through a membrane transport protein cell uses energy (ATP) to PUMP molecule in or out  Osmosis: o Water moving from high H2O concentration to low H2O concentration through a semi-permeable membrane o H2O moves from area of lower [solute] to area of higher [solute] o From Low solute / high H2O to high solute / low H2O o Osmosis Rule: H2O FOLLOWS high solute Osmosis example Left: Water and solute Right: Water only can pass can’t pass through through semi-permeable membrane membrane  bacteria in isotonic solution Three Osmosis Conditions: o Water moving from high H2O concentration to low H2O concentration through a semi-permeable membrane o H2O moves from area of lower [solute] to area of higher [solute] o From Low solute / high H2O to high solute / low H2O Isotonic Solution Osmosis Rule: H2O FOLLOWS high solute Equal solutes inside/outside cell Equal H2O movement in/out of cell bacteria in bacteria in hypertonic hypotonic solution solution Hypertonic Solution Hypotonic Solution Higher solute outside cell Higher solute inside cell Left: Water and solute Right: Water only can pass H2O moves out of cell H2O moves into cell can’t pass through through semi-permeable Called Plasmolysis Can “lyse” cells membrane membrane VIDEO: Cellular Structure  Ch.4 – Cell Structure o cell theory and cell size o eukaryotic vs. prokaryotic o prokaryotic cell structure o prokaryotic cell walls o bacterial shapes o preparing samples for light microscopy smears and stains o eukaryotic cell structure  Prokaryotic Cell Wall: Two Peptidoglycan Cell Wall Types: o Bacteria - usually contains peptidoglycan o Gram positive and gram negative o Archaea cell walls contains pseudomurein o Special cell wall type: mycolic acid cell wall Contains “waxy” glycolipid mycolic acid o Rigid cell walls provide: Protection against osmotic pressure Layer of protection against the environment Many highly pathogenic Protection against osmotic pressure Genus “Mycolata” o Cell wall contributes to pathogenicity Mycobacterium tuberculosis Bacterial cell wall helps prevent cell lysis A few bacteria lack cell walls Mycoplasma pneumoniae  Bacterial cell structure Bacterial Cell Wall: composed of peptidoglycan Polymer of a repeating disaccharide in rows: o N-acetylglucosamine (NAG) o N-acetylmuramic acid (NAM) Rows are linked by polypeptides Peptidoglycan similar to cellulose NAG and NAM structure Peptidoglycan fibers linked by polypeptides Two main bacterial cell wall types: Gram positive: thick, outer cell wall Gram negative: thin, “inner” cell wall; thin “outer” lipopolysaccharide (LPS) layer  Gram positive cell wall gram positive cell wall A thick multi-layer of cross-linked peptidoglycan and teichoic acid molecules Thick, rigid structure Porous, will allow many types of small molecules to pass through  gram negative cell wall Gram negative cell wall A single thin layer of peptidoglycan Surrounded by an outer membrane of phospholipids and lipopolysaccharides. Thin, flexible structure. Less penetrable by small molecules thanks to alternating hydrophobic and hydrophilic barriers Periplasm – concentrated, gel-like substance in space between plasma and LPS membranes Porins – membrane “barrel” proteins act as “pores”; molecule specific  Gram Positive Cell Walls Gram Negative Cell Walls Thick peptidoglycan Thin peptidoglycan Teichoic acids No teichoic acids In acid-fast cells, contains mycolic acid Outer membrane (LPS layer) Gram staining  Gram Stain: 1. Crystal Violet (purple) stains peptidoglycan purple 2. Ethanol wash removes CV from gram negative cells; dissolves LPS 3. Stain with Safranin (pink) – stains both types pink Gram + : Crystal Violet overwhelms Safranin – so PURPLE Gram - : Crystal Violet washed away - so PINK Gram negative / Gram positive cell comparison Gram Positive Bacteria Gram Negative Bacteria  Rod (gram-negative) Who’s who? Who’s gm-……..who’s gm+? Coccus (gram-positive) Gram stained bacteria  Ch.4 – Cell Structure o cell theory and cell size o eukaryotic vs. prokaryotic o prokaryotic cell structure o prokaryotic cell walls o bacterial shapes o preparing samples for light microscopy smears and stains o eukaryotic cell structure Bacterial shapes: the size, shape and arrangement of bacterial cells Most bacteria monomorphic - single shape A few pleomorphic - many shapes Five “main” shapes overall: 1. Coccus (spherical) 2. Bacillus (rod-shaped) 3. Spiral Vibrio Spirillum Spirochete 4. Star-shaped 5. Rectangular 3 categories of bacterial shapes  different cocci arrangements Prokaryotic Cell Shapes: coccus arrangements coccus, cocci (plural) = spherical cells Cocci Arrangements: how cells are “grouped” Pairs: diplococci Chains: streptococci Clusters: staphylococci Groups of four: tetrads Cube-like groups of eight: sarcinae  Prokaryotic cell shapes – rod arrangements Also called bacillus bacillus, bacilli (plural) = rod-shaped cells different rod (bacillus) arrangements Single bacillus Coccobacillus - small rod Streptobacilli -chains Dipplobacilli - pairs  prokaryotic cell shapes – spiral types spiral = curved cells vibrio spirillum spirochete  Prokaryotic Cell Shapes: unusual shapes Unusual bacterial shapes  unusual prokaryotic cell shapes: Streptomyces Dr. Paul Hoskisson, Univ. of Strathclyde, Glasgow, UK Walsby’s Square Bacterium  Gram-stained Bacillus antracis Pathogen: causes anthrax; infecting skin and lungs Scientific name: Bacillus anthracis o what’s the shape? o arrangement? o gm+ or gm-? shape: bacillus or rod arrangement: streptobacillus gram positive Gram stain - Bacillus anthracis  Ch.4 – Cell Structure o cell theory and cell size o eukaryotic vs. prokaryotic o prokaryotic cell structure o prokaryotic cell walls o bacterial shapes o preparing samples for light microscopy smears and stains o eukaryotic cell structure  Staining: Preparing a Bacterial Smear o 1st step in bacterial staining…..always! Smear: a thin film of a material containing microorganisms spread over a slide Microorganisms are fixed (attached) to the slide Kills the microorganisms Prevents cell “auto-lyses” – maintains shape and morphology Step 1: From solid media: Spread inoculation loop of sterile water in circular motion on slide Sterilize loop, add loop of bacteria to water on slide and mix From liquid broth media: Add inoculation loop of bacteria directly to slide Step 2: Bacterial smear preparation Let “air” dry - can heat gently on top of bacinerator or hot plate  Staining: Coloring microorganisms with a dye that emphasizes certain structures Stains consist of a colored positive or negative ion The colored ion is called the chromophore Cell wall has slight negative charge Two Dye Types: 1. Basic dye - the chromophore is a cation 2. Acidic dye - the chromophore is an anion Negative staining - staining the background instead of the cell Simple stain - use of a single basic dye o Highlights the entire microorganism to visualize cell shapes and structures Mordant - may be used to hold the stain or coat the specimen to enlarge it  acid-fast stain: mycolic acid cell wall Binds only bacteria that have a mycolic acid – a waxy material in their cell walls Special cell wall type of certain gram positive bacteria Mycolic acid is not decolorized by acid-alcohol decolorization step Used for identification of: Mycobacterium and Nocardia Color of Color of Acid-Fast Non–Acid-Fast Primary Stain: Red Red Carbolfuchsin Decolorizing Agent: Red Colorless Acid-alcohol Counterstain: Red Blue Methylene Blue acid-fast stain: positive (red) and negative (blue) cells acid fast staining steps  “special” stains: capsule, endospore and flagella Used to distinguish parts of an organism Capsule stain: o Capsule – a “gelatinous” and protective covering o Stain cell 1st – then Negative stain to stain background o Enables visualization of “capsuled” bacteria o Does not except most dyes Capsulated bacteria viewed with capsule stain o India ink or Nigrosin to stain background o Capsule appears as a “halo” around cells  “special” stains: capsule, endospore and flagella Endospore Stain Used to distinguish parts of an organism endospores – green (blue) Endospore stain: o Endospore Dormant, resistant protein structures House DNA and essential proteins Form when exposed to unfavorable conditions Very, very, very durable (100’s of years!) o Steps for endospore stain: Primary stain – malachite green – apply with heat Decolorize cells: water Counterstain: safranin Spores appear green around/in pink or red cells  Flagella Stain Flagella are difficult to stain Very specialized and technique dependent procedure Uses a mordant and special dye called carbolfuchsin  Methods of Identification: Differential Staining Gram stain: Acid-fast staining: simple, fast, useful for some rare cases important stain other stains for special structures: Bacterial Staining: endospore, flagella, capsule Pros: Fairly fast, cheap and easy, broad Cons: Imprecise, need enough cells, endospore stain capsule stain flagella stain technique dependent  Ch.4 – Cell Structure o cell theory and cell size o eukaryotic vs. prokaryotic o prokaryotic cell structure o prokaryotic cell walls o bacterial shapes o preparing samples for light microscopy smears and stains o eukaryotic cell structure  Evolution of the Eukaryotic Cell “Endosymbiotic Theory” o Prokaryotic cells evolved into 1st eukaryote Evolution of the Eukaryote Prokaryote into a Eukaryote o “In-folding” of plasma membrane Creating nucleus and organelles o “Proliferation” of plasma membrane Creation of endomembrane system Evolution of the eukaryotic cell  Evolution of the Mitochondria and Chloroplast o Both were bacteria phagocytized by early eukaryote 1st - Mitochondria Plant and animal cell took in bacteria Bacteria broke down sugar for ATP Bacteria evolved into mitochondria organelle 2nd - Chloroplast (Plastid) Plant cell took in photosynthetic bacteria Created glucose from CO2, H2O and light Bacteria evolved into chloroplast organelle Mitochondria and chloroplasts similarities with bacteria: Enveloped by a double membrane Contain free ribosomes and circular DNA molecules Evolution of eukaryotic mitochondria and chloroplast organelles Grow and reproduce independently in cells  Eukaryotic cell organelles and structures Eukaryotic cell: a “cell factory” tour Organelles – membrane enclosed “tiny organs” each with “special function” all outside nucleus - exception: nucleolus  Eukaryotic Protein Synthesis Protein destined for secretion Cell protein factory tour! o “path to a protein” DNA mRNA o the endomembrane system organelles involved: nucleus  Nucleus – DNA and mRNA nuclear pores  Endoplasmic Reticulum ribosomes  Golgi Complex  Lysosome endoplasmic reticulum o inner membrane system involved in protein golgi complex synthesis o proteins destined for organelle or secretion secreted or organelle specific proteins Final Protein! to organelle or secreted think of cell as a Protein Factory Assembly Line!!!!  NUCLEUS: “cell control center” Eukaryotic nucleus and nuclear pores o “houses DNA” site of DNA synthesis and replication o “houses” nucleolus site of ribosomal RNA (rRNA) synthesis o site of transcription Parts of Nucleus: Nuclear Envelope  double membrane that encloses the nucleus Nuclear Pore  nuclear channel to transport molecules in/out Nuclear Lamina Lines the nuclear side of the envelope Made of protein – maintains shape Nucleoplasm semi-fluid filled medium different pH than cytoplasm  Rough Endoplasmic Reticulum or “Rough ER” o folded-up “extension” of the nuclear envelope Functions: site of translation for proteins to be secreted checks that proteins are folded correctly applies 1st sugar “ID tag” NAME: rough – “studded” with ribosomes endo – “within” plasmic – “semi-fluid medium” reticulum – a “network” eukaryotic cell – rough endoplasmic reticulum  “Rough ER” – accepts proteins from ribosome 1. ribosome, bound mRNA, and growing protein migrate and attach to the rough ER. 2. protein chain “grows” INTO rough ER “lumen” (inside) and is released from ribosome 3. protein folds into final protein shape 4. protein transferred to golgi complex in transport vesicle rough ER and golgi complex eukaryotic cell – rough endoplasmic reticulum Golgi Complex – final protein processing o downstream of rough ER o organelle that is a series of “free-floating” membranes o CHECKS, PROCESSES, DISTRIBUTES proteins delivered from rough ER transport vesicles from rough ER FUSE with golgi complex Functions: 1. final check for mis-folded protein if so….to lysosome 2. adds sugars to the protein “labels” proteins for final destination serve as “chemical tags” or “identification markers” that “ID” proteins for routing 3. sorts proteins based on these “chemical tags” path of a protein nucleus to plasma membrane 4. transport vesicles “bud-off” golgi complex delivered to final destination – other organelle or secreted  Eukaryotic Cell Wall protective, but permeable, barrier exterior to plasma membrane gives structure resistance to hypertonic induced osmotic pressure all eukaryotic cells EXCEPT animal and protozoa (type of protist) Eukaryotic cell wall composition: o Algae – cellulose and silica (diatoms) o Plants – primary cell wall: cellulose o Fungi – chitin *NOTE* protozoa and animal cells DON’T have cell walls plant cell indicating cell wall  Mitochondria – The “Powerhouse” of the eukaryotic cell site of ATP synthesis Function: Energy Conversion site of aerobic (or anaerobic) respiration in eukaryotes generates ATP from breakdown of sugars and fats ATP (Adenosine Triphosphate): needed to supply energy for most cellular reactions almost ALL Food ultimately consumed in mitochondria Animal and plant cells and many eukaryotic microorganisms: o require O2 gas to maximize sugar breakdown and capture the energy as ATP process called aerobic respiration eukaryotic cell mitochondria Smooth Endoplasmic Reticulum (Smooth ER): no ribosomes ER – indicating smooth ER eukaryotic cell – indicating smooth ER Functions: o Site of LIPID SYNTHESIS (fats) o “lipid” transport vesicles: to other organelle(s) membrane or plasma membrane o detoxification of harmful substances (alcohol, toxins) o calcium ion (Ca2+) storage  Lysosome – a sealed off “acid vat” Lysosome digesting old mitochondria recycle / degradation center worn-out Lysosome – with organelle digestive enzymes Functions: o digest worn out cellular material 1. Lysosome “fuses” with o digest foreign material ingested by cell worn-out organelle o breaks down “large” molecules into “small” molecules recycles “usable” molecules to the cytoplasm 2. Organelle broken down expels “unusable” molecules out of cell 3. Small molecules returned to cytosol Lumen of lysosome: acidic pH: ~4.5pH 5. Waste contains strong digestive enzymes molecules expelled from enzymes are produced in ER and shipped to golgi cell BioFlix: Tour of an Animal Cell  Ch. 4 Learning Objectives After this lecture, you should be able to: 1. Describe the cell theory and relationship of surface area to volume ratio relative to cell size 2. Describe the basic differences between prokaryotic and eukaryotic cells; including cell wall differences 3. Describe the evolution of the 5 kingdoms of life 4. Describe the basic structure and function of the plasma membrane, cell wall, flagella, axial filament, fimbrae, pili, capsule, glycocalyx, endospore and chromosome of prokaryotes 5. Describe the four flagella arrangements 6. Describe the four components of the plasma membrane 7. Describe the four types of movement across plasma membrane 8. Describe the cell wall of bacteria and archaea; bacterial cell wall composition, structure and function; gram negative vs. gram positive cell wall 9. Describe the components, steps and outcome of the gram stain 10. Describe the basic cell shapes and arrangements of bacterial cells  Ch. 4 Learning Objectives After this lecture, you should be able to: 11. Understand the what a “bacterial smear” is; why and how to prepare 12. Differentiate an acidic dye from a basic dye; explain the purpose of simple staining. 13. Compare and contrast the gram stain, acid-fast stain, capsule and endospore stain 14. Why is the Gram stain so useful?; Which stain would be used to identify microbes in the genera Mycobacterium and Nocardia? 15. Why doesn't a negative stain color a cell?; Why’s fixing necessary for most staining procedures? 16. How do unstained endospores appear? Stained endospores? 17. Describe the evolution of the eukaryotic cell and the endosymbiotic theory 18. Describe the basic structure and function of the following eukaryotic organelles: plasma membrane, cell wall, endomembrane system - nucleus, rough /smooth ER, golgi complex; mitochondria and lysosome 19. Compare the overall cellular structure of prokaryotes and eukaryotes