Cytology & Nucleus

nuclear envelope

Consists of two unit membranesseparated by a 10-30 nm space: a. outer nuclear membrane- faces the cytoplasm - continuous with (& shares biochemical and functional properties with) the rER - can have ribosomes attached to the cytoplasmic surface b. inner nuclear membrane- faces the nuclear matrix - supported by the nuclear lamina - associates with chromatin The perinuclear cisternae is the space between the two unit membranes The inner and outer nuclear membranes are continuous with one another at the nuclear pores

4. Where are carbohydrates located?

- carbohydrate residues associate with lipids and proteins and are confined to the membrane surface, the outer leaflet, facing away from the cytoplasm

diploid

- cells with the full complement of chromosomes (humans = 46)

C. Melanin

- found in keratinocytes, pigment cells of the retina, and some nerve cells of the brain. Melanin formation will be discussed in the lecture on the Integuments.

haploid

- having only one of each pair (i.e., 23) as in the germ cells (egg and sperm)

Clinical Aspects

I-cell disease - missing or defective Golgi enzyme Tay-Sachs - hexosaminidase A Fabry - alpha-galactosidase A Gaucher - B-glucocerebrosidase Niemann-Pick- sphingomyelinase

RNA

RNA is a linear molecule similar to DNA but contains ribose sugar instead of deoxyribose and the base uracil instead of thymine. RNA is synthesized by the transcription of DNA.

4. Compare the two types of chromatin present in the interphase nucleus in terms of their structure and function.

(1) Heterochromatin- electron-dense portions of the interphase nucleus - appears as basophilic clumps in the light microscope - represents genes that are not being transcribed - can be either constitutive (never transcribed) or facultative (transcription depends on cell type , etc.) (2) Euchromatin- also known as extended chromatin; uncoiled or loosely packed portions of chromosomes - represents genes that are being transcribed

5. Outline the packaging of genetic material in the nucleus. Include the terms chromatin, nucleosomes, histones, linker DNA, and chromosomes in your outline.

(1) Nucleosome-the simplest arrangement of chromatin packing in the nucleus -11 nm wide structure resembling beads-on-a-string. The beads are known as nucleosomes and the string is DNA - made up of eight proteins (an octomer) consisting of pairs of four types of histones. DNA (about 150 base pairs) makes 1.75 turns around this octomer of histone proteins. The "string" part is known as the linker DNA. (2) 30 nm chromatin fibril -the nucleosomes form 30nm threads (fibrils) by coiling of consecutive nucleosomes (six nucleosomes per turn of coil) bound by another type of histone (3) continued coiling of chromatin fibrils to form chromatin fibers in euchromatin and more tightly packed heterochromatin.The coiled 30 nm fibers are looped into 300 nm loops held together by specific protein-DNA complexes. The 300 nm loops are further coiled into 700 nm helical loops which form the maximally condensed chromosomes of metaphase. These can be seen with the light microscope in a cell undergoing mitosis or meiosis. 4) Chromosomes - extensively condensed chromatin seen during mitosis or meiosis

types of RNA:

(1) messenger RNA (mRNA)- contains sequences that code for amino acids thereby dictating the amino acid sequence of proteins (2) transfer RNA (tRNA) -transfers the correct amino acid to the ribosome for incorporation into the protein (3) ribosomal RNA (rRNA) -forms ribosomes in association with specific proteins. Ribosomes are the sites of protein synthesis.

Structure of the ribosome:

(1) one large subunit - composed of 5S, 28S, and 5.8S rRNA and ~ 49 proteins; catalyzes the peptide bond formation (2) one small subunit - composed of 18S rRNA and ~ 33 proteins; binds mRNA and tRNA

Nucleolus Areas

(1) pale-staining region (fibrillar center) (2) pars fibrosa or dense fibrillar region (3) pars granulosa or granular component

1. Draw the structure of cell membranes placing the three components - lipids, proteins and carbohydrates - in their proper orientation.

(A) lipids (50% of membrane mass) responsible for membrane form and permeability properties; (B) proteins responsible for most of the membrane's specific functions (amount and type of proteins varies); and (C) carbohydrates that are confined to membrane's surface. unit membrane- three distinct layers (trilaminar structure)

aneuploidy

- any deviation in the normal number of chromosomes

3. Discuss membrane proteins in terms of their arrangement in the membrane and their general functions.

- are either peripheral proteins (associate with membranes through ionic interactions) or integral proteins (buried in the lipid membrane such as transmembrane proteins) - membrane proteins perform most of the membrane's functions - the presence of membrane proteins is indicated by the freeze fracture technique

Nuclear pores Functions

- bidirectional "gates" for trafficking of molecules between the nucleus and the cytoplasm - small molecules (< 40 - 60 kd) pass through pores by diffusion - proteins of any size containing a nuclear localization amino acid sequence are transported by an energy-dependent mechanism (requiring either ATP or GTP)

c. autosomes

- chromosomes that are not sex chromosomes (humans = 22)

polyploid

- containing more than one complement of chromosomes

D. Crystals

- least common of the inclusions. They include the crystals of Reinke found in the interstitial cells (Leygig cells) of the human testis and the crystals of Charcot-Bottcher found in the Sertoli cells of the human testis.

Nuclear lamina location and structure

- located between the inner nuclear membrane and the peripheral heterochromatin - made up a class of intermediate filament proteins called lamins (lamins A, B, C) - lamin phosphorylation at mitosis causes dissolution of the nuclear lamina

Microtubules

- non-branching hollow cylinders of ~25 nm in diameter - formed from 13 parallel protofilaments, each protofilament is composed of dimers of α and β tubulin subunits - are polarized with a plus end and a minus end; the plus end is the rapidly growing end; minus end is the slow growing end and is usually embedded in the MTOC (microtubule-organizing center). The centrosome is a MTOC. - The centrosome is composed of: 1. fibrous centrosome matrix containing 50 or more copies of the γ-tubulin ring (γ-TuRC) 2. a pair of centrioles arranged at right angles to each other.Each centriole is composed of nine triplets of MT

Nuclear pores Structure

- nuclear pore proteins are known as nucleoporins - the nuclear pore contains a central cylindrical body between inner and outer octagonal rings; each ring contains 8 protein particles - protein filaments extend from each ring; they form a basket on the nuclear side

a. genome

- number of chromosomes in somatic cells; is species specific (humans = 46)

d. sex chromosomes

- one pair per cell; either xx (female) or xy (male)

Hemoglobin

- oxygen carrying pigment of red blood cells. Hemosiderin- granular pigment derived from hemoglobin - hemosiderin represents large aggregates of ferritin micelles (ferritin micelles are formed from the association of iron with the protein apoferritin and represents the storage form of iron in cells) - hemosiderin can be unambiguously identified by the Prussian blue histochemical reaction - small amounts of hemosiderin are normally located in macrophages of the spleen, bone marrow and liver where old red blood cells are normally degraded - excessive accumulations of hemosiderin is pathological and known as hemosiderosis

2. Identify the major lipid components of membranes and their functions

- phospholipids are the most abundant type of lipid in membranes - phospholipids are amphipathic: hydrophilic and a hydrophobic region - cholesterol is the second major lipid of membranes. does not freeze - cholesterol maintains the structural integrity of membranes - glycolipids are the third most abundant membrane lipids - glycolipids are always found on the outer leaflet

5. Describe MAPs and molecular motors.

- some MAPs function as molecular motors that are responsible for the movement of organelles, vesicles, etc.; they utilize ATP to move "cargo" in a single direction along the filament Examples: a. kinesin - movement toward the plus end b. dynein - movement toward the minus end (cytoplasmic and the axonemal dyneins

The Nucleolus location

- usually eccentrically placed in the interphase nucleus - not surrounded by a membrane -1-2 per cell depending on the species and synthetic activity of the cell

2. Name biological processes where apoptosis plays an essential role.

-Development - proper number, spacing and orientation of neurons -generating the middle ear space -generating the vaginal opening -generating the spaces between the fingers and toes -removal of unneeded structures such as male mammary tissuecontrol of cell number -Adult organism - to control cell number and changes in organ functions. To protect the organism from cells that may be harmful. Examples: -mammary gland and uterus during and after pregnancy -uterus during the menstrual cycle -changes in liver following drug (barbiturate) exposureviral infected or transformed cells

Chromatin

-complex of DNA, histone proteins found in the nucleus of eukaryotic cells. -one copy of a chromosome formed by DNA replication still joined at the centromere to the other copy.

Intermediate filament assembly

-dimer has central regions wound into coiled-coil -staggered side-by-side arrangement of two dimer forms tetramer -tetramer is basic subunit for assembly of filaments - IF are more stable than Mts or MFs

Microfilaments functions

-maintenance of cell shape; transport of organelles; separation of chromosomes during mitosis/meiosis; ciliary/flagellar movement. - many functions of microtubules are due to the property of dynamic instability; this refers to the rapid conversion between a growing and shrinkage state. - microtubule-associated proteins (MAPs) bind alongside MTs and organize MTs into bundles; stabilize MTs against disassembly; mediate interactions of MTs with other cellular components Cell shape cel-cell and cell matrix adhesion contraction stability and function of microvilli locomotion endo & exocytosis

The Nucleus

-membrane-limited organelle that contains the genetic information of the cell. -function: replication of DNA (as required by cell division); DNA repair; RNA transcription and processing

How a cell's shape affect its function?

-red blood cells (RBCs) are very small, flat discs, which allows them to easily fit through narrow capillaries and around sharp corners in the circulatory system to deliver oxygen throughout the body. -Neurons carry messages from the brain and spinal cord to the rest of the body, using electrical signals down their lengths and chemical signals between neurons. Since electrical signals travel much faster than chemical signals, neurons are long and thin to minimize the number of slower chemical signals that would be required between links in a chain of many shorter neurons. -The elongated shape of muscle cells allows the contraction proteins to line up in an overlapping pattern that makes muscle flexing possible..

aneuploidy examples:

-trisomy - presence of a third chromosome of one type (Down's syndrome, Klinefelter's syndrome) -monosomy - absence of one membrane of a chromosome pair -(Turner's syndrome = absence of one of the sex chromosomes (X0); all other types of monosomy are lethal)

Membranes

1. (1) they are not homogenous; (2) membrane components are in a constant dynamic flux; (3) membranes are asymmetric, and (4) the shape of the surface membrane is partly determined by the underlying cytoskeleton.

Microtubule-Associated Proteins

1. Binds along side of MT 2. Stabilize MTs against disassembly 3. Mediate the interaction of Mts 2ith other cellular components 4. Organize Mts into bundles 5. Organize the cytoplasm into functionally different areas -axons and dendrites of nerve cells 6. Increases the rate of nucleation (start of elongation )

5. Name the types of inclusions found in a cell.

1. Glycogen 2. Lipid 3. Pigments a. Lipofuscin b. Hemoglobin- Hemosiderin c. Melanin d. Crystals

1. Compare the two processes by which cells die: necrosis and apoptosis.

1. Necrosis- results from mechanical injury, exposure to toxins or anoxia. Characteristics include cell swelling, chromatin clumping, deterioration of organelles followed by cell lysis and phagocytosis by macrophages. This is usually associated with inflammation and tissue deterioration. 2. Apoptosis- programmed cell death important during development, organ reorganization, and the control of cell number. The characteristics include decrease in cell volume with the cell membrane and the organelles remaining intact. The cell ultimately breaks up into membrane-bound globules that are phagocytized by macrophages leaving no trace.

Mitochondria function

1. Production of ATP a. Reactions of the matrix space - conversion of pyruvate and fatty acids into acetyl CoA - acetyl CoA enters the Krebs cycle and NADH and FADH2are produced; these are important carriers of electrons in catabolic reactions b. Reactions on the inner membrane - electrons from NADH and FADH2 are passed down the respiratory or electron transport chain causing protons (H+) to be pumped across the membrane from the matrix into the intermembrane space generating an electrochemical proton gradient across the inner membrane - the gradient drives protons back through the ATP synthase complex causing ATP synthase to add a phosphate to ADP forming ATP - ATP is transferred to the cell 2. Self-replication- proliferate by division (fission) of pre-existing mitochondria; life span is ~ 10 days 3. Mitochondrial DNA- DNA resembles that of bacteria, i.e., a closed circle. The DNA codes for mitochondrial rRNA and tRNAs but only some of the mitochondria's mRNA. Most of the mitochondrial proteins are encoded by nuclear DNA with mRNA translation occurring on free ribosomes. 4. Initiates apoptosis- Mitochondria initiate apoptosis (cell death) by releasing cytochrome c from the intermembrane space into the cytoplasm. Cytochrome c release initiates the activation of proteolytic enzymes, known as caspases leading to apoptosis.

There are over 200 distinguishable cell types in the body assembled into any of the four basic tissues:

1. epithelia 2. connective tissue 3. muscle 4. nerve

Mitochondria Components:

1. outer membrane a. porins (voltage-dependent anion channels) - proteins that form transmembrane channels freely permeable to small uncharged molecules b. receptors for proteins and polypeptides; several enzymes 2. intermembrane space a. space between the two membranes; chemically equivalent to the cytoplasm with regard to small molecules b. contains kinases and cytochrome c; release of cytochrome c from the intermembrane space into the cytoplasm initiates apoptosis 3. inner membrane - has the highest protein concentration than any membrane of the cell.

Cells common functions

1. similar mechanism to synthesize and degrade protein replicate DNA 2. contract and generate energy 3. move substance into and out of the cell

Cells share common features

1. surrounded by a membrane 2. have zero to many nuclei 3. contain organelles (membrane bound and non-membrane bound) 4. contain inclusions

The Cytoskeleton

A highly dynamic, complex network of protein filaments that organizes the cytoplasm and determines cell shape. Functions include cell motility, organelle and vesicle transport, muscle contraction, cell division, endocytosis/phagocytosis, providing mechanical strength.

3. Sketch the mitotic spindle and label the three types of microtubules, the centrioles, chromosomes, centromere, and kinetochore.

A. Replication of the centrioles - occurs during interphase (S phase); remain together until prophase B. Centriole migration - occurs at the beginning of mitosis - cellular MTs increase their rate of disassembly during late prophase - MTs originating from the centrosome increase C. The mitotic spindle consists of: 1. polar MTs - overlap at the midline of the spindle - responsible for pushing poles apart 2. kinetochore MTs - attach to kinetochore at centromere 3. astral MTs - radiate in all directions from centrosome - contributes to forces that separate the poles - positions poles in the cell

4. Describe cytokinesis.

At the end of mitosis, a contractile ring of actin and myosin II assembles beneath the plasma membrane; it contracts splitting the cell in two

Mitochondria of brown fat

Brown fat cells contain condensed mitochondria characterized by a decrease in the size of the inner compartment, an increase in the matrix density, and enlargement of the intermembrane space. In brown fat cells, there is an uncoupling of oxidation from phosphorylation producing heat instead of ATP.

Formation of ribosomes

Events occurring in the nucleolus: (1) transcription of rDNA to form pre-rRNA (2) association of pre-rRNA with ribosomal proteins to form ribonucleoproteins (3) cleavage of pre-rRNA into the 28S, 18S, and 5.8S rRNAs found in ribosomes Continued formation of ribosomes in the nucleus: (4) 28S and 5.8S rRNA combines with 5S rRNA (5S rRNA is transcribed in the nucleus) to form the large ribosomal subunit (5) active transport of ribosomal subunits to the cytoplasm through nuclear pores Continued formation of ribosomes in the cytoplasm: (6) assembly of ribosomal subunits into ribosomes and polysomes in the presence of mRNA

nuclear lamina

Functions: - gives shape and stability to the nuclear envelope - organizes the interphase nucleus - structural link between chromatin and the nuclear envelope - responsible for the dissolution and reformation of the nuclear envelop during cell division - positions nuclear pore complexes within the nuclear envelope cytoskeleton. doesn't allow for transcription to occur

Barr body

In females, one of the two X chromosomes is inactive and highly coiled. This can be seen in light microscopy as a small drumstick or as a clump of heterochromatin usually located adjacent to the nuclear envelope. -most obvious/ visible in a white blood cell

Phagocytosis

Ingestion of large particles usually >250 nm in diameter Vesicles formed are called phagosomes Phagocytosis is performed by specialized cells originating mostly from monocytes (cells of the mononuclear phagocytic system)

Mitochondria

Location: In all cells but prominent in cells that use large amounts of energy such as striated muscle cells and cells engaged in fluid and electrolyte transport. Hepatocytes also have numerous mitochondria (~1000 per cell). Morphology: · variety of shapes such as spheres and rods; the common rod-shaped forms are approximately 0.4-0.8 µm in diameter and 4-9 µm in length · can be visualized with the light microscope with appropriate stains such as PTAH contain an outer and inner membrane thereby

Cells of the mononuclear phagocytic system differentiate from blood monocytes and migrate to and from connective tissue of the respective organs; examples:

Macrophage (histiocyte): connective tissue proper Kupffer Cell: liver Alveolar Macrophages: lung Osteoclast: bone Microglia: brain Langerhan's cell: skin

Lipid

Morphology - lipid droplets are not surrounded by a membrane - usually seen as round, clear vacuoles in light microscopy because lipids are extracted during tissue fixation. If the tissue is fixed in glutaraldehyde and osmium tetroxide, the lipid is preserved as a black or gray spherical globule - in electron micrographs, lipid droplets are usually associated with or surrounded by mitochondria Location - found predominantly in adipocytes where lipids occur as a single large droplet - abundant in liver and steroid secreting cells Functions - storage form of triglycerides and cholesterol - good energy reserve; twice as many ATP molecules are generated from 1 gram of fat as from 1 gram of glycogen Clinical Aspects Steatosis - abnormal fat accumulation in liver cells due to obesity, metabolic syndrome, alcohol consumption, and other conditions indicative of abnormal metabolism

Glycogen

Morphology a. Light microscopy - can be visualized in light microscopy following staining with the periodic-acid Schiff reaction (PAS stain); however, it is lost during routine processing of tissue b. Electron microscopy - in electron microscopy, glycogen particles appear as 20-30 nm dense granules called beta(b) particleswhich aggregate to form alpha (a) particlesalso known as rosettes Location- found in all cells but particularly in abundance in liver and muscle Function- a polymer of d-glucose that animal cells use to store carbohydrates - the synthesis and degradation of glycogen is accomplished by enzymes (glycogen synthase and glycogen phosphorylase) bound to its surface Clinical Aspect Glycogenated hepatocyte nuclei are commonly found in liver biopsy specimens from patients with a variety of clinical disorders such as diabetes, obesity, metabolic syndrome, and non-alcoholic fatty liver disease, and Wilson's disease.

5. Describe the glycocalyx, its structure, function and localization

On the cell surface, the carbohydrate component contributes to the glycocalyx. The glycocalyx is a carbohydrate-enriched coating that covers the outside of many eukaryotic cells and prokaryotic cells, particularly bacteria . glycocalyx is involve in cell to cell recognition, interaction with immune cells

Endocytosis

Pinocytosis = ingestion of fluid and particles usually smaller than 150 nm in diameter Can be either clathrin-independent (may involve caveolin-coated vesicles) or clathrin-dependent as in receptor-mediated endocytosis

F. Secretory vesicles

The fusion of vesicles with the plasma membrane is known as exocytosis. There are two types of secretory pathways leading to exocytosis: (1) constitutive pathway - continuous delivery of contents, for example, immunoglobulins and tropocollagen (2) regulated secretory pathway - in specialized cells such as endocrine cells and neurons; needs a signal for release and is calcium dependent

1. Sketch the nucleus and label its major components.

Three major components of the nucleus: 1. nuclear envelope a. outer nuclear membrane b. inner nuclear membrane c. nuclear pores d. nuclear lamina 2. chromatin 3. nucleolus

1. Draw a pie diagram of the cell cycle and include the subdivisions of interphase.

a. G1Phase - cell monitors its environment and size; period of cell growth b. G0 - specialized resting state or terminal differentiated state c. S phase - replication of DNA; duplication of centrioles d. G2 - safety gap; cell makes sure DNA replication is complete

2. Discuss the stages of mitosis.

a. Prophase - chromatin starts to condense - dissassembly of cytoplasmic MTs - beginning of mitotic spindle formation b. Prometaphase - breakdown of nuclear envelope -attachment of chromosomes to spindle c. Metaphase - alignment of chromosomes on the metaphase plate d. Anaphase - separation of kinetochores - movement of chromatids to opposite poles e. Telophase - separated chromatids arrive at poles - disappearance of kinetochore MTs - elongation of polar MTs - reformation of nucleus - chromosomes decondense Cytoplasmic division (cytokinesis) begins.

2. Recognize the various shapes and number of nuclei per cell and translate this into the possible functions of a cell

a. Shape: round, ellipsoid, infolded, or lobulated. b. Size: varies c. Number per cell: from none to multi d. Location: central, basal, eccentric -Function: The surface area of a cell dictates how much interaction the cell will have with its environment

Components of the inner membrane:

a. cristae - thin folds formed by the inner membrane that project into the interior of the organelle; number of cristae per mitochondria depends on the cell's energy expenditure b. inner membrane subunits or elementary particles (ATP synthases) - proteins consisting of a globular head connected to the membrane by a slender stem (lollipop-like structures); responsible for the generation of ATP from ADP and inorganic phosphate. c. cardiolipin - a phospholipoprotein that makes the inner membrane impermeable to ions d. enzymes of the respiratory chain - essential for the process of oxidative phosphorylation which generates most of the cell's ATP e. special transport proteins - regulate the passage of metabolites into and out of the matrix

1. Identify the three major components of the cytoskeleton.

a. microfilaments (7 nm thick) b. intermediate filaments (10 nm thick) c. microtubules (25 nm in diameter)

Polyribosomes

clusters of ribosomes around a single strand of mRNA.

b. karyotype

how to it looks during metaphase - standard map of the banding pattern of each chromosome

The Endoplasmic Reticulum (ER)

largest membranous system in the cell consisting of interconnected tubules and vesicles; has two interconnected components: the smooth ER (sER) and the rough ER (rER). The rER membrane is continuous with the outer nuclear membrane

Vesicles

membrane-bound compartments that exchange components among themselves and other membrane-bound compartments

matrix space

occupied by the electron dense mitochondrial matrix a. matrix granules - stores calcium and other ions inside the mitochondria thereby contributing to the regulation of cytoplasmic ion concentrations b. enzymes of the citric acid (Krebs) cycle c. enzymes involved in β-oxidation of fatty acids d. DNA, tRNAs, ribosomes - Mitochondria contain their own DNA, rRNA, tRNA, and mRNA.

2. Understand how the nucleolus is formed; what are the nucleolar organizing regions and what are their functions.

regions of ten interphase chromosomes (pairs of 5 different chromosomes, human) that contain the genes that encode rRNA; help reorganize the nucleoli following cell division

The Nucleolus Function

site of ribosomal RNA (rRNA) transcription and rRNA synthesis - regulation of the cell cycle (nucleostemin) -Composition - rich in rRNA and protein

6. What is a lipid raft?

specialized membrane domains that associate with or segregate different proteins or signaling molecules. Areas in the membrane where you want to have specific proteins.

D. Endosomes

vesicles associated with endocytosis including receptor-mediated endocytosis There are two types of endocytosis: (1) Pinocytosis - ingestion of fluids and solutes via small (< 150 nm) vesicles; includes receptor-mediated endocytosis (2) Phagocytosis - the ingestion of large particles (>250 nm) by specialized cells (discussed later)

Phagosomes

vesicles formed from phagocytosis; phagosomes fuse with lysosomes to degrade phagocytized particles

Pigments- Lipofuscin

yellow-brown pigment representing undigested residues in lysosomes (residual bodies). Mostly present in cells with a long life span, i.e., cardiac muscle and nerve. It is referred to as the age pigment. The composition of lipofuscin can vary between different cell types depending on the autophagocytized material. Disease-related lipofuscin pigment (as occurs in lysosomal storage diseases) is occasionally called 'ceroid'. - lipofuscin is not a harmless bi-product of insufficient degradation; it is now believed to contribute significantly to ageing and age-related pathologies. Examples of the later include: 1. age-related macular degeneration - main cause of blindness in the elderly due to lipofuscin accumulation in RPE and degeneration of photoreceptors 2. Alzheimer's disease - leading to senile dementia, is characterized by intraneuronal aggregates of the protein tau among other characteristics 3. Parkinson's disease - accumulation of α-synuclein aggregates in neurons of the substantia nigra 4. increased incidences of cardiomyopathies and heart failure with ageing 5. atherosclerosis - accumulation of oxidized LDL in long-lived macrophages may reduce their function in clearing of hazardous lipids from blood vessels.

B. Coatomer-Coated Vesicles

· COP I: Retrograde Transport - from Golgi cisternae to the rER - the retrieval pathway · COP II: Anterograde Transport - from the rER to the Golgi

C. Caveolae

· Form due to lipid composition of membranes · Major structural protein is caveolin · Function in clathrin-independent endocytosis · Important for transcytosis

3. Golgi Apparatus

· From the rER the next stop for a newly synthesized protein is the Golgi apparatus. · prominent in protein secreting cells; functions in protein packaging, concentration and sorting · does not stain with H&E but is indicated by a clear area. It does stain with osmium. · with em, seen as stacks of 4-10 parallel cisternae which are often curved. The periphery is dilated and surrounded by vesicles that are either budding off or fusing to the Golgi · exhibits functional polarity. The convex surface (cis face) receives secretory protein from the rER; the opposite, concave side is the trans or exit face · In the Golgi, newly synthesized proteins pass in sequence through four distinct compartments: (1) cis compartment (cis face) and small transfer vesicles (2) intermediate or medial compartment (3) trans (trans or exit face) compartment (4) trans Golgi network (TGN) - where sorting of proteins occur (final destination - lysosomes, plasma membrane, and constitutive or regulated secretion)

Microfilaments

· composed of actin which is the major cytoskeletal protein of most cells (~5-10% of total cell protein). · actin filaments are ~7nm diameter · composed of G-actin (globular protein) that binds head to tail to form the polarized actin protofilament - actin subunit bind to ATP

A. Clathrin-coated vesicles

· functions include (1) transport of newly synthesized proteins from the TGN to lysosomes; (2) transport of newly synthesized proteins from the TGN to granules for regulated secretion; (3) formation of vesicles in receptor -mediated endocytosis (discussed below) · morphology: coated with aggregations of clathrin molecules - consists of 3 large and 3 small polypeptide chains that form a triskelion(3-legged structure). The clathrin triskelion associates to form a polyhedral cage-like network around the vesicle.

(1) Receptor-Mediated Endocytosis

· ingestion of specific molecules from the extracellular environment · begins at specialized regions of the plasma membrane known as coated pits containing clathrin · the coated pits invaginate to form clathrin-coated vesicles · vesicles lose their clathrin coat and fuse with or mature into an endosome

Peroxisomes

· major function: β-oxidation (breakdown) of fatty acid molecules to form acetyl CoA · liver and kidney cell peroxisomes are important for the detoxification of toxins that enter the blood stream; they convert half of ingested alcohol to acetaldehyde. · peroxisomes contain oxidative enzymes such as catalase and urate oxidase, oxidative reactions generate hydrogen peroxide · catalase converts hydrogen peroxide to oxygen and water. · Peroxisomes also function in gluconeogenesis and the metabolism of purines. · Similar to mitochondria, peroxisomes are a major site of oxygen use but do not produce ATP. · In some species, the enzyme urate oxidase forms a crystalloid (nucleoid) distinguishing peroxisomes from other vesicles. · Peroxisomes of different cells in the same organism may have different sets of enzymes. · In nerve tissue, peroxisomes catalyze the first reaction in the formation of plasmalogens, the most abundant class of phospholipids in myelin. · Peroxisomes replicate by growth and fission.

B. Rough ER (rER)- RoughEndoplasmic Reticulum

· membrane contains ribosomes bound on the cytoplasmic side · can be localized in a cell due to the staining of the ribosomes with basic dyes. Cytoplasm that takes up basic stains is known as the ergastoplasm. In neurons, it is known as Nissl substance · prominent in protein secreting cells (that is, cells that secrete enzymes and other proteins such as peptide hormones, collagen and immunoglobulins) · functions include post-translation modification (such as sulfation, glycosylation and folding) of newly synthesized proteins destined for secretion, lysosomes, and the plasma membrane · ribosomes are directed to the rER membrane by the signal peptideon the growing polypeptide chain. · The growing polypeptide chain-ribosome complex binds to a SRP receptor on the rER membrane and is directed to the protein translocator on the rER membrane · The translocator directs the growing peptide chain through the rER membrane and into the lumen of the rER · Cleavage of the signal peptide and initial glycosylation of the protein chain occurs in the rER cisternae

E. Lysosomes

· membrane-limited bodies containing acid hydrolases (enzymes) · heterogeneous in appearance · internal pH of ~ 4.7 maintained by an ATP-driven proton pump · functions in the degradation of endocytosed material · also degrades cellular material, in particular, long-lived proteins, all organelles and other macromolecules including the proteosome (a multicatalytic proteinase complex that also degrades many cellular proteins) · synthesis of the lysosomal acid hydrolases occurs on the rER; the hydrolases are targeted to the lysosomal compartment due to the addition of a mannose-6-phosphate (M6P) group to newly synthesized lysosomal protein.

A. Smooth ER (sER) -Smooth Endoplasmic Reticulum (sER)

· membranous system of interconnected tubules & vesicles · prominent in cells that specialize in lipid metabolism and in steroid synthesis · is responsible for synthesizing cholesterol · synthesizes the lipid portion of lipoproteins · contains enzymes for the degradation of toxins · sequesters calcium from the cytosol, in skeletal muscle sER is known as the sarcoplasmic reticulum In hepatocytes: · The sER undergoes hypertrophy when challenged with alcohol, drugs (phenobarbital, anabolic steroids, progesterone) and some drugs used in chemotherapy · The sER can become the predominant organelle in hepatocytes · The presence of sER makes the hepatocyte cytoplasm display acidophilia (acidic dye = eosin) Degradation of drugs and other xenobiotics

Clinical Aspect: Zellweger Syndrome

· most common inherited disease related to peroxisomes · results from a mutated receptor for the peroxisome targeting signal Individuals with this disorder show accumulations of very long-chain and branched-chain fatty acids in tissues and cells which eventually impair organ function There are also deficient levels of plasmalogens affecting brain and nervous system function. Currently there is no cure for diseases resulting from peroxisome deficiencies.

Microfilament functions

· some functions of actin include cell attachment (focal adhesion), cell-cell adhesion, cell surface modifications (example, microvilli and stereocilia), movement/contraction · drugs affecting actin polymerization include phalloidin, cytochalasin, swinholide, latrunculin

3. Discuss the structure, function and location of intermediate filaments.

· ~10 nm diameter · composed of a variety of proteins whose expression is cell-type specific (see below) · IFs are not polarized · more stable than actin or tubulin · function is primarily structural; provides mechanical strength Types & locations of IFs: Class 1: Keratins - epithelial cells Class 2: Vimentin & vimentin-like - fibroblasts, muscle, glial cells, nerve stem cells Class 3: Neurofilament - neurons Class 4: Lamins - nucleus (in all nucleated cells) Class 5: Beaded filaments - eye lens fiber cells

Cell Death

• Apoptosis • Autophagic Cell Death • Necrosis • Mitotic catastrophe • Anoikis - apoptosis induced by loss of attachment • Excitotoxicity • Wallerian degeneration • Cornification

2. Connective Tissue

• Consists of relatively few cells that are not adherent to each other • Vascular • Abundant matrix • Some functions include cohesion of structural elements, serves as a medium through which blood vessels distribute nutrients and take up metabolic waste, cells are involved in immune & inflammatory responses and in tissue repair following injury

3. Muscle

• Muscle tissue is responsible for movement and changes in the size and shape of body organs. • The elongated muscle cells are usually oriented parallel to each other and organized in bundles. • The muscle cytoplasm (sarcoplasm) is occupied mostly by the myofilaments actin and myosin. • The arrangement of the filaments is highly organized allowing for contraction.

4. Nerve

• The nervous system is responsible for responding to the environment and to maintain coordinated functional activities of organs and organ systems. • There are two main cell types - the neuron and supporting cells.

1. List all the components of the cytoplasm.

•Cytoplasmic matrix •Organelles •Inclusions •Cytoskeleton

1. Epithelia

•Epithelia cover body exterior surfaces and line body cavities including blood vessels. • They also form glands both exocrine and endocrine. • Epithelial cells are tightly adherent to one another. • The shape of epithelial cells can be squamous, cuboidal or columnar. • The functions of epithelial cells include protection, absorption, secretion and excretion, and gas exchange.