CHAPTER 1 INTRODUCTION TO THE S TUDY OF CELL & MOLECULAR BIOLOGY THE DISCOVERY OF CELLS
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Robert Hooke (1665), English microscopist (at age 27, became curator of the Royal Society) A. Described chambers in cork; called them cells (cellulae) since they reminded him of cells occupied by monks living in a monastery B. Found them while trying trying to explain why cork stoppers could hold air in a bottle so effectively C. Was looking at empty cell walls, walls, the remains of dead cells; no no internal structure
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Anton van Leeuwenhoek (1665-1675 (1665-1675), ), Dutch seller of clothes & buttons buttons in spare time, he was was first to describe living single cells; results were checked and confirmed by Hooke A. Saw “animalcules” in pond water using the scopes of remarkable quality that he made made
B. Described various forms of bacteria from tooth scrapings & water in which pepper was soaked C. Eventually, became celebrity visited visited by Russia's Peter the Great & the queen of England England III. 1830s - full & widespread importance of cells realized A. Matthias Schleiden , botanist (1838 (1838)) - all plant tissues composed of cells; plant embryos arise from single cell B. Theodor Schwann, zoologist (1839 (1839)) - same conclusion about animals; plants & animals similar C. Schwann then proposed first two tenets of Cell Theory are composed of one or more cells. 1. All organisms are 2. The cell is the structural unit of life for all organisms. D. However, the Schleiden-Schwann view view of the origin of cells was less insightful since both agreed that cells could arise from noncellular materials -> eventually disproved by others E. Rudolf Virchow, German pathologist (1855 (1855)) - added third tenet of Cell Theory derived from his cell division observations; it ran counter to Schleiden-Schwann view of cell origins 1. Cells can arise only by division from a preexisting cell. BASIC PROPERTIES OF CELLS
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Life – most most basic property of cells; they are the smallest units to exhibit this property; plant or animal cells c an be removed from organism & cultured in laboratory A. Can grow and reproduce for a long time in culture, culture, unlike their parts which soon deteriorate B. George Gey, Johns Hopkins Univ. (1951 (1951)) - first human cell culture (HeLa cells); donor was Henrietta Lacks (from her malignant tumor); still grown in laboratories today C. Cultured cells are simpler to study than cells in body; cells grown in vitro (in culture, outside body) are essential tool of cell & molecular biologists
II. Cells are highly complex and organized A. Each level of structure in cells is consistent from cell to cell – each each cell has consistent appearance in EM; organelles have particular shape & location in individuals of a species B. Organelles have consistent macromolecular macromolecular composition composition arranged in a predictable pattern C. Cell structure similar similar organism to organism despite differences in higher higher anatomical features III. Cells possess genetic program & the means to use it (a blueprint); encoded in collection of genes A. Blueprint for constructing cellular structures & ultimately organisms B. Directions for running running cell activities C. Program for making more cells IV. Cells are capable of producing more of themselves - mitosis mitosis and meiosis A. Contents of “mother” “mother” cell distributed to 2 “daughter” “daughter” cells
B. Before division, genetic material is faithfully copied; each daughter cell gets complete complete & equal share of genetic information C. Usually, daughter cells have roughly equal volume; during egg egg production, one cell gets most of cytoplasm & half of genetic material V. Cells acquire & utilize energy energy to develop & maintain complexity complexity - photosynthesis photosynthesis & respiration A. Virtually all energy needed by life arrives from sun sun B. This energy is trapped by light-absorbing pigments in photosynthetic photosynthetic cells C. Light energy turned to chemical chemical energy by photosynthesis; stored in energy-rich energy-rich carbohydrates D. Animals get energy energy prepackaged usually in form of glucose glucose E. Once in cell, glucose disassembled; most most energy is stored as ATP & used to run cell activities VI. Cells carry out many chemical chemical reactions - sum total of chemical reactions in cells (metabolism); (metabolism); to do this, cells require enzymes (molecules that greatly increase rate of chemical reactions) VII. Cells engage in numerous mechanical mechanical activities based on dynamic, mechanical changes changes in cell: A. Material moved moved from place to place B. Structures assembled assembled and disassembled C. Cells move move from place place to place
VIII. Cells able to respond to stimuli whether cells are uni- or multicellular - have receptors that sense environment & initiate responses (move away from object in path or toward nutrient source) A. Most cells covered with receptors that interact in specific ways with substances in environment 1. Receptors bind to hormones, growth factors, extracellular materials, surfaces of other cells 2. Allow ways for external agents to evoke specific responses in target cells B. Cells may respond to specific stimuli by: 1. Altering metabolic activities 2. Preparing for cell division 3. Moving from one place to another, or 4. Even committing suicide IX. Cells are capable of self-regulation A. Importance of regulatory mechanisms most evident when they break down 1. Failure of cell to correct error in DNA replication -> may lead to debilitating mutation 2. Breakdown in growth control -> may lead to cancer cell & maybe death of whole organism B. Example: Hans Driesch, German embryologist (1891) - separate first 2 or 4 cells in sea urchin embryo -> each produces normal embryo TWO FUNDAMENTALLY DIFFERENT CLASSES OF CELLS:
PROKARYOTES AND EUKARYOTES
I. With advent of EM, 2 cell types were distinguished by size & types of internal structures (organelles); exhibited a large fundamental evolutionary discontinuity (no known intermediates) A. Prokaryotes ( pro - before; karyon - nucleus) – all bacteria, cyanobacteria (blue-green algae); structurally simpler 1. Prokaryotes now living very similar to those fossilized in >3.5 billion year old rocks (Australia, S. Africa); sole life on planet for nearly 2 billion years before first eukaryote B. Eukaryotes (eu - true) - structurally more complex; protists, fungi, plants, animals II.
Similarities between prokaryotes and eukaryotes - reflect fact that eukaryotes almost certainly evolved from prokaryotic ancestors A. Both types of cells share an identical genetic language B. Both types of cells share a common set of metabolic pathways C. Both types of cells share common structural features - cell membrane, cell walls (same function, different structure) III. Characteristics that distinguish prokaryotic & eukaryotic cells - eukaryotic cells are internal much more complex (structurally and functionally) A. Eukaryotes have membrane-bound nucleus with complex nuclear envelope & other organelles 1. Prokaryotes have nucleoid (poorly demarcated cell region)[ no membrane-bound organelles B. Prokaryotes - relatively little DNA (0.25 - ~3 mm) coding for several hundred to several thousand proteins (1 mm of DNA = ~3 x 106 base pairs) 1. Simplest eukaryotes (4.6 mm in yeast encoding ~6200 proteins) have slightly more DNA than prokaryotes; most eukaryotes have an order of magnitude more DNA C. Eukaryotic chromosomes numerous; contain linear DNA tightly associated with protein; prokaryotes have single, circular chromosome with DNA that is nearly naked D. Cytoplasmic structures - eukaryotes have many; prokaryotes mostly devoid of such structures (except for infolded bacterial mesosomes & cyanobacteria photosynthetic membranes) 1. Intracytoplasmic communication smaller issue in prokaryotes due to size (diffusion works); in eukaryotes, interconnected channels/vesicles transport stuff around cell & out of cell 2. Eukaryotes have cytoskeletal elements generally lacking in prokaryotes – cell contractility, movement, support 3. Ribosomes of prokaryotes smaller than those of eukaryotes (essentially same function) 4. Both eukaryotes & prokaryotes may be surrounded by rigid, nonliving cell wall that protects, but their chemical composition is very different 5. Eukaryotes have more complex locomotor mechanisms – prokaryotes have rotating flagella; eukaryotes have more complex flagella with different mechanism (also cilia & pseudopodia) E. No mitosis or meiosis in prokaryotes (binary fission instead); prokaryotes proliferate faster (double in 20 - 40 minutes; exchange genetic information via conjugation) 1. In eukaryotes, chromosomes are compacted & separated by mitotic spindle which allows each daughter cell to get equal genetic material 2. In prokaryotes, no chromosome compaction & no spindle; DNA copies separated by growth of intervening cell membrane
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3. In conjugation, recipient almost never gets whole chromosome from donor; cell soon reverts to single chromosome F. Examples of some eukaryotic organelles and their functions – divide the cytoplasm into compartments within which specialized activities take place 1. 2. 3. 4. 5. 6.
Mitochondria (plants & animals) – make chemical energy available to fuel cell activities Endoplasmic reticulum (plants & animals) – where many cell lipids & proteins are made Golgi complexes (plants & animals) – sorts, modifies, transports stuff to specific locations Variety of simple membrane-bound vesicles of varying dimensions plants & animals) Chloroplasts (plants) – sites of photosynthesis Single large vacuole (plants) – occupies most of cell volume
IV. Prokaryotes not inferior - metabolically very sophisticated & highly evolved A. Have remained on Earth more than 3 billion years B. They live on and in eukaryotic organisms, including humans C. Make almost everything they need; need only simple carbon (only 1 or 2 low MW organic compounds), nitrogen source(s) & some inorganic ions; some live on only inorganic substances 1. One species found in wells >1000 m below Earth's surface; live on basalt rock & H2 made by inorganic reactions 2. Even most versatile cells in human require a variety of organic compounds (vitamins, etc.) D. Bacteria in our large intestine even make some essential dietary ingredients for us
Types of Prokaryotic Cells
I. Divided into two major groups or domains – Archaea & Bacteria II. ARCHAEA (archaeons or archaebacteria) - groups of primitive bacteria (related DNA sequences); closest relatives of first cells; live in extremely inhospitable environments (extremophiles) A. Methanogens - capable of converting CO2 & H2 gases into methane (CH4) gas B. Halophiles - live in extremely salty environments (Dead Sea & Great Salt Lake) C. Acidophiles – acid-loving prokaryotes that live at pHs as low as 0 D. Thermophiles - live at very high temperatures 1. Hyperthermophiles (ex.: Pyrolobus fumaril) - live in hydrothermal vents of ocean floor; reproduce at temperatures above 109°C & won't grow below 90°C III. BACTERIA (eubacteria) A. Bacteria are present in every conceivable habitat on earth – permanent Antarctic ice shelf to driest African deserts to internal confines of plants & animals, rock layers several km deep 1. Some of these bacteria cut off from life on surface for >100,000,000 years B. Example: Mycoplasma - smallest living cells (0.2 µm dia); only prokaryotes lacking cell wall C. Example: Cyanobacteria (formerly blue-green algae) - most complex; elaborate cytoplasmic membrane arrays which are sites of photosynthesis; similar to membranes in chloroplasts 1. Filled world with O2; need few resources to survive 2. Some do N2 fixation - convert N2 gas into reduced nitrogen forms (e. g. NH3) used to make amino acids & nucleotides D. Those species capable of both photosynthesis & nitrogen fixation survive on barest resources – light, N2, CO2, H2O 1. Not surprising that cyanobacteria are the first to colonize bare rocks left lifeless by volcano IV. Prokaryotic diversity A. To study prokaryotic diversity, cells can be concentrated, their DNA extracted & DNA sequences analyzed 1. All organisms share certain genes (genes for rRNAs or some metabolic pathway enzymes) 2. Sequences of these genes vary species to species 3. Carefully analyze variety of sequences for particular gene in habitat -> tells you the number of species living in the habitat B. By carefully analyzing sequences in extracted DNA & comparing sequences to those in known organisms, one can learn about phylogenetic relationships of these organisms 1. Prokaryotes living in single Yellowstone National Park pool – 30% of sequences were from bacteria that could not be grouped into any of the 14 known divisions in the domain Bacteria 2. Based on such differences, the previously unidentified bacteria were put in 12 new divisions C. Most habitats on earth teeming with previously unidentified prokaryotic life 1. Archaea once thought to be restricted to harshest environments 2. Now found to be common & abundant members of non-extreme habitats (oceans, lakes, soil) 3. >90% of these organisms now thought to live in subsurface sediments well beneath oceans & upper soil layers; not long ago, deeper sediments thought to be only sparsely populated 4. Carbon sequestered in world's prokaryotes is roughly comparable to total carbon in plants
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Types of Eukaryotic Cells
I. Unicellularity vs. multicellularity - most complex eukaryotic cells are among single-celled protists A. Protists - must do everything an organism must do to survive; one evolutionary pathway B. Multicellular organisms exhibit differentiation - different activities conducted by different types of specialized cells II. Example of multicellularity & differentiation - cellular slime mold Dictyostelium shows advantages provided by division of labor among cells A. During most of life, they are independent amoebas, each a complete, self-sufficient organism B. If food scarce, stream toward each other & form sluglike aggregate (pseudoplasmodium or slug) 1. Slug migrates slowly over substratum leaving slime trail 2. Previously single organisms now small part of larger, multicellular individual C. Cells are no longer a homogeneous population 1. Cells differentiate into prestalk cells of anterior third of slug and posterior prespore cells D. Soon, slug stops moving, rounds up on substratum & extends upward into air 1. Forms elongated fruiting body (sporangium) 2. Sporangium has slender stalk supporting rounded mass of dormant, encapsulated spores 3. Stalk (from prestalk cells) supports spore mass (from prespore cells) above substratum 4. Spores scatter and give rise to next generation of amoebas III. Differentiation – process by which a relatively unspecialized cell becomes highly specialized A. Fertilized egg develops into many cell types (hundreds) in mature organism 1. Cells specialized for varied functions, have distinctive appearance, carry unique materials 2. Cells have similar organelles but their number, appearance & location may differ & correlate with cell act. B. Differentiation of each eukaryotic cell depends primarily on signals received from environment 1. Signals, in turn, depend on position of cell within embryo 2. As a result, different cell types acquire distinctive appearance & contain unique materials C. Despite differences, various cells of multicellular plant or animal are made of similar organelles 1. Mitochondria are found in all cell types, but they may change shape (rounded or highly elongated & threadlike) 2. Brown adipose cell (main function is generation of heat from chemical energy stored in fat); has numerous fat droplets & lots of mitochondria where energy conversion occurs 3. Plasma cell specialized for antibody production – have relatively small number of mitochondria but extensive rough endoplasmic where protein synthesis occurs 4. Number, appearance & location of organelles can be correlated with activities of particular cell type IV. Cell & molecular biology research focuses on small number of representative or model organisms A. Saccharomycese cerevisiae , a budding yeast B. Arabadopsis thaliana – a mustard plant C. Caenorhabditis elegans – a nematode D. Drosophila melanogaster – a fruit fly E. Mus musculus – a mouse The Sizes of Cells and Their Components
I. Units of linear measure most often used to describe cell structures A. Micrometers (µm; 10-6 m), nanometers (nm; 10-9 m) B. Ångstroms (Å; 10-10 m) – often used by molecular biologists for atomic dimensions although no longer formally accepted in metric nomenclature); ~1 Å = diameter of H atom II. Examples of dimensions of cells and cell components A. Typical globular protein (myoglobin) - ~ 4.5 nm x 3.5 nm x 2.5 nm B. Highly elongated proteins (collagen, myosin) - over 100 nm in length C. DNA - ~2 nm in width D. Large molecular complexes (ribosomes, microtubules, microfilaments); 5 - 25 nm dia. E. Nuclei - about 10 µm diameter; mitochondria - about 2 µm in length F. Bacteria - 1 to 5 µm in length; eukaryotic cells - 10 to 30 µm in length III. Why are most cells so small? A. Most eukaryotic cells have single nucleus with only 2 copies of most genes 1. Thus, cells can only produce limited number of mRNAs in a given amount of time 2. The larger a cell's volume, the longer it takes to make the number of mRNAs the cell needs B. As a cell increases in size, the surface area/volume ratio decreases 1. If surface area/volume ratio gets too small, surface area not sufficient to take up substances needed to support metabolism (oxygen, nutrients, etc.) or get rid of wastes
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C. As cell gets larger, takes too long for diffusion to move substances in and out of active cell 1. Time required for diffusion is proportional to the square of the distance traversed 2. O2 required 100 µsec to diffuse 1 µm, but 106 times as long to diffuse 1 mm 3. As cell becomes larger, distance from surface to interior gets larger; diffusion time to move things in & out of metabolically active cell becomes prohibitively long IV. How do large cells get around the surface area/volume problems? - examples A. Ostrich egg & others - little living protoplasm spread over top of lots of inert yolk nutrient B. Giraffe (and other large animal) nerve cells - very long but very small diameter C. Plant cell interior filled with large fluid-filled vacuole; needs no support, unlike cytoplasm D. Intestinal epithelium specialized for absorption with microvilli to increase surface area Viruses
I. Pathogens smaller and, presumably, simpler than smallest bacteria; called viruses A. Late 1800s - thought infectious diseases caused by bacteria but other agent soon found 1. Sap from sick tobacco plant found to infect other plants while containing no bacteria 2. Sap still infective if forced through filter with pores smaller than smallest known bacteria 3. Infectious agent could not be grown in culture unless living plant cells also present B. Wendell Stanley, Rockefeller Institute (1935) - tobacco mosaic virus (TMV), a rod-shaped particle was crystallized & found to be infective; thought to be protein 1. Now know it is a single RNA molecule surrounded by helical shell of protein subunits C. Viruses responsible for many human diseases, some cancers - come in different shapes, sizes & constructions – AIDS, polio, influenza, cold sores, measles, a few types of cancers II. Common virus properties - not considered living since need host to reproduce, metabolize, etc. A. All are obligatory intracellular parasites (must reproduce in host cell [plant, animal, bacteria]) 1. Alone, they are unable to reproduce, metabolize or carry on other life-associated activities 2. Thus, they are not considered to be organisms & not considered to be alive 3. Once it has attached & passed through membrane, it can alter host cell activities B. Outside of living cell, it exists as particle or virion, essentially a macromolecular package C. Has genetic material (single/double stranded DNA or RNA); 3 or 4 genes up to several 100 1. The fewer the genes, the more it relies on enzymes & other proteins encoded by host genes D. Genetic material surrounded by protein capsule (capsid) usually made up of a specific number of subunits; efficient (need only a few genes to make capsid) 1. Capsid subunits often organized into polyhedron with planar faces (ex.: 20-sided icosahedron) like adenovirus which causes mammalian respiratory infections E. Many animal viruses have capsid surrounded by lipid-containing outer envelope derived from modified host cell membrane as virus buds from host cell surface (ex.: HIV) F. Bacterial viruses (bacteriophages) are among most complex – T bacteriophages polyhedral head (contains DNA), cylindrical stalk (injects DNA) & tail fibers (attach to bacteria) 1. Used in key experiments that revealed genetic material structure & properties G. Viruses have surface proteins that bind to particular host cell surface component (specificity) 1. HIV - glycoprotein of 120,000 dalton MW (gp120) interacts with specific protein (CD4) on surface of certain white blood cells facilitating virus entry into host cell 2. Viral & host protein interaction determines virus specificity, the hosts it can enter & infect H. Most viruses have relatively narrow host range (certain cells of certain host like human cold & influenza viruses, which are only able to infect human respiratory epithelium cells 1. But some can have wide host range, infecting cells from a variety of organs or species - rabies infects variety of mammalian host species (bats, dogs, humans) 2. Host cell specificity change can have dramatic effect – 1918 influenza epidemic killed >20 million people; flu strain may have been so virulent because it infected many cell types III. Two basic types of viral infection A. Lytic infection - virus usually arrests normal host activities, redirects cell to make new viral nucleic acids & proteins that self-assemble into new virions 1. Cell lyses to release new viral particles & infect neighboring cells B. Formation of provirus - integrates its DNA into host DNA, but no immediate host cell death IV. Effects of integrated provirus depend on type of virus & host cell - up to 1% of human DNA is DNA from proviruses that infected our ancestors (now just genetic garbage transmitted passively)
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A. Bacterial cells with provirus behave normally until exposed to some stimulus (e. g. UV radiation) that activates dormant viral DNA 1. Then cells make new virions & lyse releasing viral progeny - bacterial lambda () virus B. Animal cells with provirus may make new viruses by cell surface budding without lysis – HIV 1. Infected cell may stay alive for a period acting as a factory for production of new virions C. Animal cells with provirus may lose growth & division control -> malignant (tumor viruses) V. Viral origin A. Unlikely that viruses present before hosts since they need hosts for reproduction, etc. B. Since have same genetic language as hosts, they could not have arisen independently as primitive form after other cells had evolved C. Probably a degenerate form derived from more complex cellular organism - maybe evolved from small cell chromosome fragments able to maintain a type of autonomous existence in cell D. Over time, these autonomous genetic elements acquired protein coat, became infective agents E. Different viruses likely arose independently from various organisms (genes similar to host genes) 1. Corroboration – genes present in each group of viruses are different from those of other groups but similar to genes within host cells they infect 2. Difficult to find drugs not harmful to human host since viruses use host enzymes VI. Viruses have virtues - research tool to study host DNA replication/gene expression, insect-killing viruses (pest control), used to introduce foreign genes into human cells as treatment (gene therapy) Viroids
I. T. O. Diener, U. S. Dept. of Agriculture (1971) - discovered an agent causing potato spindle-tuber disease; potatoes get gnarled, cracked A. Infectious agent was small circular RNA lacking protein coat (viroids) B. Viroid traits 1. RNAs range from about 240 to 600 nucleotides (10% size of smaller viruses) 2. No evidence that RNA codes for proteins; viroids use host enzymes & proteins completely; ex.: duplication of viroid RNA in infected cell uses host RNA polymerase II C. May cause disease by interfering with cell's normal path of gene expression (e. g. monopolize RNA polymerase II to duplicate viroid RNA) II. Viroid diseases can have serious effects on crops A. Cadang-cadang - devastated coconut palm groves of Philippines B. Another has wreaked havoc on chrysanthemum industry in U. S.
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