Ch. 3 Lab Flash Cards Cell Biology, Mitosis, and Molecular Genetics

Components and Operation of the Compound Light Microscope

• Eyepiece Lens (Ocular Lens): Serves as the primary point of magnification, typically providing a 4x $10\times$ or $40\cdot$ or 100x magnification power.

• Barrel (Body Tube): Connects the eyepiece lens directly to the objective lenses.

• Arm: Acts as the structural backbone of the microscope, elevating the objective lenses above the stage. Serves as one of the two designated holding points when transporting the instrument.

• Stage: The flat platform where the specimen slide is placed for observation. The slide is secured using slide clips and can be adjusted across four directions using the stage control.

• Coarse Focus Knob: Moves the stage toward or away from the objective lenses to bring the specimen into general focus.

• Fine Focus Knob: Fine-tunes the focus to sharpen the image and reveal detailed cellular structures.

• Lamp Source and Dimmer: Emits light upward through the slide; light intensity can be modified using an adjustable dimmer knob.

• Iris Diaphragm: A rotating disc located beneath the stage that regulates the beam diameter and intensity of light projected upward into the slide.

• Base: The bottom structural support of the microscope. Proper carrying procedure requires using both hands—one supporting the base and the other gripping the arm.

Cell Size Constraints and Microscopy Techniques

• Physical Limits on Cell Size:

  • Upper Size Limit (Surface Area to Volume Ratio): Nutrients (such as oxygen) enter cells and wastes (such as carbon dioxide) exit via diffusion. As cell volume increases, the internal distance material must travel increases significantly. Small cell size keeps diffusion distances short and efficient.

  • Lower Size Limit: Cells cannot be infinitely small because they must contain sufficient volume to house essential molecular machinery, proteins, organelles, and genetic material ($DNA$).

• Cytoskeletal Infrastructure:

  • The cytoplasm is not a loose fluid bag; it contains an organized internal structural framework (lattice).

  • Organelles move along cytoskeletal tracks like a monorail system, powered by walking motor proteins.

• Microscopy Modalities:

  • Optical (Light) Microscopes: Utilize visible light and optical glass lenses to magnify specimens (e.g., standard classroom microscopes).

  • Electron Microscopes: Utilize electromagnetic fields to focus electron beams for significantly higher resolution.

    • Transmission Electron Microscope (TEM): Transmits electrons through ultrathin specimen sections.

    • Scanning Electron Microscope (SEM): Bounces electrons off specimen surfaces coated in heavy metals to generate 3D topography.

    • Limitations: Sample preparation for conventional electron microscopy destroys and kills the biological specimen.

  • Fluorescent Optical Microscopes: Employ fluorescent dyes and markers to label specific live or dead cellular structures, allowing high-resolution visualization without destroying live cells.

Cellular Taxonomy and Comparative Structural Features

• Universal Cellular Characteristics: All living cells share three fundamental features:

  1. A selectively permeable plasma membrane regulating internal vs. external environments.

  2. An aqueous jelly-like cytoplasm (specifically cytosol).

  3. Genetic material in the form of $DNA$.

• Prokaryotic vs. Eukaryotic Cells:

  • Prokaryotic Cells: Simple, single-celled organisms lacking a membrane-enclosed nucleus or internal membrane-bound organelles. Genetic material resides freely in the cytoplasm. Examples include Bacteria and Archaea.

  • Eukaryotic Cells: Larger, structurally complex cells containing a distinct membrane-bound nucleus and specialized membrane-enclosed organelles. Examples include plants, animals, fungi, and protists.

• Plant vs. Animal Cell Differences:

  • Plant Cells: Contain a rigid external cell wall composed of cellulose, chloroplasts for photoautotrophic photosynthesis, and a large central vacuole for water storage and turgor pressure. They lack centrioles.

  • Animal Cells: Lack cell walls and chloroplasts; possess centrioles/centrosomes and smaller, dynamic vacuoles for endocytosis and exocytosis.

• Specialized Cellular Extensions:

  • Cilia: Microscopic, hair-like membrane projections that move in coordinated wave patterns. Found on epithelial cells lining the human respiratory tract to sweep inhaled debris and mucus away from lungs.

  • Flagella: Long, whip-like tails used for cellular propulsion. Found on specific bacteria and human sperm cells (the only flagellated cell in the human body).

  • Microvilli: Tiny, finger-like folds of the plasma membrane that increase surface area for cellular absorption (e.g., intestinal epithelial cells).

Detailed Tour of Eukaryotic Cell Organelles

• Nucleus and Nucleolus:

  • Nucleus: The central genetic control center of the cell, enclosed by a double-membraned nuclear envelope perforated with nuclear pores that regulate substance traffic.

  • Nucleolus: A dense region within the nucleus where chromosomes transcribe ribosomal RNA ($rRNA$). Ribosomal proteins synthesized in the cytoplasm enter the nucleolus, assemble with $rRNA$ into ribosomal subunits, and export back to the cytoplasm through nuclear pores.

  • Chromatin: Uncoiled, thread-like structures of $DNA$ wrapped around histone proteins distributed throughout the nucleus during interphase.

• Ribosomes:

  • Non-membrane-bound complexes of $rRNA$ and proteins consisting of a small subunit and a large subunit.

  • Function as sites of protein synthesis (translation), binding messenger RNA ($mRNA$) and transfer RNA ($tRNA$).

  • Occur as free ribosomes floating in the cytosol or bound ribosomes attached to the rough endoplasmic reticulum.

• Rough Endoplasmic Reticulum (Rough ER):

  • A continuous membrane network of flattened sacs connected directly to the nuclear envelope, studded with bound ribosomes on its outer surface.

  • Functions as a protein manufacturing factory where nascent proteins enter the internal lumen, fold into functional secondary, tertiary, and quaternary structures, receive carbohydrate attachments (glycoproteins), and package into transport vesicles.

• Smooth Endoplasmic Reticulum (Smooth ER):

  • Membrane network lacking attached ribosomes.

  • Responsible for lipid, phospholipid, and steroid hormone synthesis, carbohydrate metabolism, and enzymatic detoxification of drugs, toxins, and alcohol.

  • Adaptation: Frequent toxic exposure (such as chronic alcohol consumption) induces cellular proliferation of smooth ER in liver cells, increasing chemical tolerance.

• Golgi Apparatus (Golgi Body):

  • A stack of flattened, curved membranous sacs (resembling stacked pita bread).

  • Functions as the cell's processing and shipping center (analogy: corporate shipping terminal). Receives transport vesicles containing proteins from the rough ER at its receiving face, modifies them (e.g., trimming or adding sugar chains), and packages them into vesicles at its maturing face for secretion, membrane repair, or lysosomal target delivery.

• Lysosomes:

  • Membranous vesicles ("suicide sacs") containing powerful hydrolytic digestive enzymes formed by the Golgi apparatus.

  • Fuse with phagocytosed food vesicles or damaged organelles (autophagy) to break down macromolecular contents, bacteria, and cellular debris.

  • Rupture of lysosomes across a cell triggers autolysis or programmed cell death ($apoptosis$).

• Peroxisomes:

  • Membranous sacs containing oxidase and catalase enzymes.

  • Detoxify harmful substances (including alcohol and formaldehyde) and neutralize free radicals by converting them to toxic hydrogen peroxide ($H_2O_2$), which catalase subsequently degrades into harmless water ($H_2O$) and oxygen.

  • Replicate independently via binary fission or by budding off from the ER.

• Mitochondria:

  • Double-membraned powerhouse organelles responsible for cellular respiration and ATP generation ($ATP$).

  • Features a smooth outer membrane and an intensely folded inner membrane forming shelf-like projections called cristae, maximizing surface area for metabolic enzymes.

  • Endosymbiotic Theory Evidence: Mitochondria contain their own circular $DNA$, possess distinct ribosomes, and self-replicate through binary fission independently of nuclear division.

• Vacuoles:

  • Membrane-bound storage sacs. Plant cells feature a large central vacuole storing water, ions, and nutrients to generate turgor pressure against the cell wall.

  • Freshwater protists utilize contractile vacuoles to actively pump out excess water entering via osmosis.

• Cytoskeleton:

  • An internal structural network maintaining cell shape, securing organelles, and facilitating motility.

  • Microtubules: Thick, hollow tubes composed of tubulin protein subunits. Resist cellular compression (analogy: structural support columns) and act as monorails for vesicle transit.

  • Microfilaments: Thin, solid protein threads composed of actin. Provide tensional resistance (analogy: suspension bridge cables) and drive amoeboid movement, cytoplasmic streaming, and division furrow formation.

• Centrosome and Centrioles:

  • Centrosome: The primary microtubule-organizing center located near the nucleus.

  • Centrioles: A pair of barrel-shaped structures composed of microtubule triplets situated within the centrosome of animal cells. Initiate and anchor the mitotic spindle fibers during cell division. Higher plants lack centrioles.

• Cytosol and Inclusions:

  • Cytosol: The aqueous fluid portion of the cytoplasm containing water, dissolved electrolytes, metabolic solutes, and distinct concentration gradients.

  • Inclusions: Non-functioning chemical substances stored in the cytoplasm, such as glycogen granules, lipid droplets, or pigment granules.

Plasma Membrane Structure and Transport Mechanisms

• Fluid Mosaic Model:

  • The plasma membrane comprises a dynamic double layer of phospholipids (phospholipid bilayer) with scattered embedded proteins, cholesterol, and surface sugar chains.

  • Phospholipid Orientation: Hydrophilic (water-loving) polar phosphate heads face outward toward the aqueous intracellular cytosol and extracellular interstitial fluid. Hydrophobic (water-fearing) nonpolar fatty acid tails face inward toward each other, creating an impermeable nonpolar core.

  • Cholesterol: Embedded between fatty acid tails to stabilize membrane fluidity across temperature fluctuations.

  • Membrane Proteins: Function as selective transport channels, carrier proteins, signal transduction receptors, biological enzymes, or structural anchors.

  • Glycoproteins and Glycolipids: Branching sugar chains attached to proteins or lipids on the extracellular face. Serve as biological ID tags for cell-to-cell recognition and act as cellular glue.

• Passive Transport Mechanisms (No cellular $ATP$ energy required):

  • Substances move spontaneously down their concentration gradient from an area of higher concentration to lower concentration.

  • Diffusion: Movement of solutes down concentration gradients. Particles diffuse directly if they are small enough to pass through membrane channels or if they are lipid-soluble (e.g., oxygen, carbon dioxide).

  • Facilitated Diffusion: Passive movement of larger or charged molecules (e.g., glucose, ions) across the membrane aided by specific transmembrane protein channels or carriers.

  • Osmosis: The simple diffusion of water molecules across a selectively permeable membrane toward higher solute concentrations.

• Active Transport Mechanisms (Requires cellular $ATP$ energy):

  • Substances are moved against their concentration gradients (from low concentration to high concentration) or transported via energy-consuming bulk membrane manipulation.

  • Solute Pumping: Transmembrane protein pumps utilize $ATP$ hydrolysis to force ions or small molecules across the membrane against concentration gradients.

  • Vesicular Transport (Bulk Transport):

    • Exocytosis: Secretory vesicles containing intracellular products (e.g., hormones, digestive enzymes, mucus) move to the plasma membrane, fuse with the bilayer, and discharge their contents into the extracellular space.

    • Endocytosis: Cell engulfs extracellular material by invaginating a segment of the plasma membrane to form a membrane-bound vesicle.

      • Phagocytosis ("Cell Eating"): Large cytoplasmic extensions called pseudopods fold around large particles, foreign bacteria, or dead cellular debris, enclosing them in a phagosome that fuses with a lysosome for destruction.

      • Pinocytosis ("Cell Drinking"): Non-specific engulfment of droplets of extracellular fluid containing dissolved solutes via small invaginating vesicles (prevalent in intestinal mucosa).

      • Receptor-Mediated Endocytosis: Highly selective bulk transport mechanism wherein specific target molecules (e.g., hormones, iron, cholesterol) bind to specialized membrane receptor proteins prior to vesicle internalization.

The Cell Cycle and Mitosis

• The Cell Cycle: The sequence of growth and division events experienced by a cell, divided into two major phases: Interphase and M-Phase.

  • Interphase: Non-dividing metabolic phase accounting for roughly $90\%$ of a cell's life cycle. Divided into three sub-phases:

    • $G_1$ Phase (Gap 1): Active cell growth, metabolic activity, and organelle duplication.

    • $S$ Phase (Synthesis): Replication of nuclear genetic material ($DNA$).

    • $G_2$ Phase (Gap 2): Final growth phase, protein synthesis, and centrosome duplication in preparation for division.

    • $G_0$ Phase: A non-dividing state entered by mature cells that permanently cease division (e.g., nerve cells, skeletal muscle cells).

  • M-Phase: Division of the cell nucleus (Mitosis) and cytoplasm (Cytokinesis).

• Purpose of Mitosis: Ensures exact duplication and equal distribution of nuclear genetic material ($DNA$) into two genetically identical daughter cells. Allows a single fertilized egg ($zygote$) to develop into a complex organism and provides tissue renewal and repair.

• Structural Elements of Mitosis:

  • Replicated Chromosomes: Composed of two identical strands of condensed $DNA$ termed sister chromatids, joined together at a central constricted region called the centromere.

  • Kinetochore: Specialized protein complexes located on the centromere of each sister chromatid that attach directly to mitotic spindle microtubules.

  • Mitotic Spindle: A lattice structure composed of microtubules originating from centrosomes that physically manipulates and separates chromatids.

• Phases of Mitosis (Mnemonic: IPMAT c):

  • Prophase: Chromatin fibers condense into tightly coiled visible chromosomes. Centrosomes begin moving to opposite cellular poles, assembling the mitotic spindle between them. The nucleolus disappears and the nuclear envelope breaks down.

  • Prometaphase: The nuclear envelope completely fragments. Microtubules invade the nuclear region, attaching to kinetochore protein targets on the centromeres.

  • Metaphase: Chromosomes align precisely along the equatorial midpoint of the cell, forming the metaphase plate (one of the straightest structural alignments in nature).

  • Anaphase: Centromeres split simultaneously, converting sister chromatids into individual daughter chromosomes. Kinetochore microtubules shorten, pulling chromatids apart toward opposite poles. Non-kinetochore microtubules elongate to push the cell poles further apart.

  • Telophase: Reverse of prophase. Chromosomes reach opposite poles and uncoil back into loose chromatin. Mitotic spindle disassembles. Nuclear envelopes reform around each set of daughter chromosomes, and nucleoli reappear within the new nuclei.

  • Cytokinesis: The division of the cytoplasm.

    • Animal Cells: A contractile ring of microfilaments pinches the plasma membrane inward, forming a cleavage furrow along the metaphase plate that divides the cell into two separate daughter cells.

    • Plant Cells: Vesicles carrying cell wall materials accumulate along the equator, fusing to form a cell plate that expands outward to synthesize a new rigid dividing cell wall.

    • Exceptions: If mitosis occurs without cytokinesis, multinucleate cells result (common in human liver cells and skeletal muscle fibers).

Molecular Genetics: Transcription and Translation

• The Central Dogma: Formulated by Francis Crick, defining the directional flow of genetic information:   $\text{DNA} \rightarrow \text{RNA} \rightarrow \text{Protein}$

  • Culinary Analogy: Master $DNA$ = Reference Cookbook in the nuclear library; Messenger $RNA$ ($mRNA$) = Transcribed recipe written on an index card; Ribosome = Kitchen chef; Amino Acids = Raw ingredients; Final Protein = Prepared dish.

• Transcription (Nucleus):

  • The enzymatic process of copying a specific $DNA$ gene segment into a complementary single strand of messenger $RNA$ ($mRNA$).

  • RNA Polymerase: The primary enzyme that binds to a promoter region on $DNA$, unzips the double helix, and constructs an $mRNA$ strand utilizing complementary base pairing rules:

    • $DNA$ Adenine ($A$) pairs with $RNA$ Uracil ($U$).

    • $DNA$ Thymine ($T$) pairs with $RNA$ Adenine ($A$).

    • $DNA$ Cytosine ($C$) pairs with $RNA$ Guanine ($G$).

    • $DNA$ Guanine ($G$) pairs with $RNA$ Cytosine ($C$).

  • RNA Processing: Non-coding intervening sequences (introns) are spliced out of the pre-$mRNA$ transcript, and coding sequences (exons) are joined together. The finished $mRNA$ strand exits the nucleus through nuclear pores into the cytoplasm.

• Translation (Cytoplasm):

  • The process wherein the genetic code carried by $mRNA$ is decoded by a ribosome to assemble a linear chain of amino acids (polypeptide).

  • Codons: Nucleotide triplets on $mRNA$. Each codon specifies a single amino acid.

  • Transfer RNA (tRNA): Adapter molecules that transport specific amino acids from the cytoplasm to the ribosome. The base of each $tRNA$ features an anticodon triplet complementary to an $mRNA$ codon.

  • Ribosomal Dynamics: The ribosome holds the $mRNA$ strand; matching $tRNA$ molecules enter the ribosome, bind their complementary codons, deposit their amino acid onto the growing peptide chain via peptide bonds, and exit to retrieve another amino acid.

  • Genetic Code Rules:

    • Start Codon: Every gene transcript begins translation at codon $AUG$, which codes for the amino acid Methionine ($Met$).

    • Stop Codons: Translation terminates when the ribosome reaches a stop codon ($UAA$, $UAG$, or $UGA$), which binds a release factor rather than an amino acid, releasing the finished polypeptide chain.

• Step-by-Step Translation Decoding Demonstration:

  • Given a transcribed messenger $RNA$ strand segment:       $\text{5'-AUG-ACG-GAG-CAA-UAA-3'}$

  • Codon 1: $AUG \rightarrow \text{Methionine (Met) [Initiation/Start]}$

  • Codon 2: $ACG \rightarrow \text{Threonine (Thr)}$

  • Codon 3: $GAG \rightarrow \text{Glutamic Acid (Glu)}$

  • Codon 4: $CAA \rightarrow \text{Glutamine (Gln)}$

  • Codon 5: $UAA \rightarrow \text{Stop Sequence [Termination]}$

  • Resulting Polypeptide Sequence:       $\text{Methionine} - \text{Threonine} - \text{Glutamic Acid} - \text{Glutamine}$

  • Following translation, the polypeptide chain folds into its distinct tertiary protein shape, ready to perform structural or enzymatic functions within the organism.