NUCLEUS
- The nucleus is the site of deoxyribonucleic acid (DNA) replication and transcription of DNA into precursor ribonucleic acid (RNA) molecules.
- It contains all of the enzymes required for replication and repair of newly synthesized DNA, as wel as for transcription and processing of precursor RNA molecules.
- It is enclosed by the nuclear envelope and contains the nuclear lamina, nucleolus, and chromatin.
Nuclear Envelope
- The nuclear envelope is a double membrane containing pores that are approximately 90nm in diameter. The outer nuclear membrane is continuous with the endoplasmic reticulum.
Nuclear lamina
- The nuclear lamina is a lattice like network of proteins that include lamins.
- Lamins attach chromatin to the inner membrane of the nuclear envelope and participate in the breakdown and reformation of the nuclear envelope during the cell cycle.
- Phosphorylation of the lamina (by lamin kinase) during prophase of mitosis initiates nuclear disassembly into small vesicles.
Nucleolus
- The nucleolus is responsible for ribosomal RNA (rRNA) synthesis and ribosome assembly.
- It contains three morphologically distinct zones:
- Granular zone-found at the periphery; contains ribosomal precursor particles in various stages of assembly.
- Fibrillar zone-centrally located; contains ribonuclear protein fibrils.
- Fibrillar center-contains DNA that is not being transcribed.
Chromatin
- Chromatin is a complex of DNA, histone proteins, and nonhistone proteins.
- DNA-a double-stranded helical molecule that carries the genetic information of the cell. It exists in three conformations: B DNA,Z DNA, and A DNA.
- Histone proteins-positively charged proteins enriched with lysine and arginine residues. They are important in forming two types of structures in chromatin: nucleosomes and solenoid fibers.
- The nucleosomes are the basic repeating units of the chromatin fiber, having a diameter of approximately 10 nm.
- Nonhistone proteins-include enzymes involved in nuclear functions such as replication, transcription, DNA repair, and regulation of chromatin function. They are acidic or neutral proteins.
Forms of chromatin
- Heterochromatin-highly condensed (30-nm solenoid fibers or higher states of condensation) and transcriptionally inactive. In a typical eukaryotic cell, approximately 10% of the chromatin is heterochromatin. Almost the entire inactive X chromosome (Barr body) in each somatic cell in a woman is condensed into heterochromatin.
- Euchromatin---,a more extended form of DNA,which is potentially transcriptionally active. In a typical cell, euchromatin accounts for approximately 90% of the total chromatin, although only about 10% is being actively transcribed in the 10-nm fiber of nucleosomes.
CYTOPLASM
Ribosomes
- Ribosomesare composed of rRNA and protein. They consist of large (60S)and small (405) subunits.
- Ribosomes are assembled in the nucleus and transported to the cytoplasm through the nuclear pores. The large ribosomal subunits are synthesized in the nucleolus, whereas the small subunits are synthesized in the nucleus.
- Polysomes-Ribosomes often form polysomes, which consist of a single messenger RNA (mRNA) that is being translated by several ribosomes at the same time. The ribosomes move on the mRNA from the 5' end toward the 3' end. The two ribosomal subunits associate on the mRNA, with the small subunit binding first.
Forms of ribosomes
- Ribosomes exist in two forms:
- Free polysomes are the site of synthesis for proteins destined for the nucleus, peroxisomes, or mitochondria.
- Membrane-associated polysomes are the site of synthesis of secretory proteins, membrane proteins, and lysosomal enzymes.
Endoplasmic Reticulum
The endoplasmic reticulum exists in two forms, rough endoplasmic reticulum (RER) and smooth endoplasmic reticulum (SER).
Rough endoplasmic reticulum
RER is a single, lipid bilayer continuous with the outer nuclear membrane. It is organized into stacks of large flattened sacs called cisternae that are studded with ribosomes on the cytoplasmic side.
RER synthesizes proteins that are destined for the Golgi apparatus, secretion, the plasma membrane, and lysosomes. RER is very prominent in cells that are specializedin the synthesis of proteins destined for secretion (e.g., pancreatic acinar cells).
Smooth endoplasmic reticulum
SER is a network of membranous sacs,vesicles, and tubules continuous with the RER, but lacking ribosomes.
SER contains enzymes involved in the biosynthesis of phospholipids, triglycerides, and sterols.
Functions of SER
Detoxification Reactions
These are reactions that make compounds water soluble so that they can be excreted. Two types of reactions that increase solubility are:
Hydroxylation reactions-by way of hydroxylase complexes containing cytochrome P450, a flavoprotein, and a nonheme iron protein
Conjugation reactions-the transfer of polar groups (i.e., glucuronic acid) from the active
carrier UDPglucuronic acid to the toxic water-insoluble molecule
Steroid synthesis
- Glycogen Degradation and Gluconeogenesis
Removal of the phosphate group from glucose-6-phosphate by the enzyme glucose-6 phosphatase, an integral membrane protein of the SER. This controls the formation of free glucose from glycogen and via gluconeogenesis.
- Reactions in Lipid Metabolism
Lipolysis begins in the SER with the release of a fatty acid from triglyceride. The SER is also the site where lipoprotein particles are assembled
- Sequestration and Release of Calcium Ions
In striated muscle the SER is known as the sarcoplasmic reticulum (SR).The sequestration and release of calcium ions takes place in the SR.
Golgi Apparatus
The Golgi apparatus consists of disc-shaped smooth cisternae that are assembled In stacks (dictyosomes), having a diameter of approximately 1micro.m and associated with numerous small membrane-bound vesicles.
The Golgi apparatus has two distinct faces:
The cis (forming) face is associated with the RER.
The trans (maturing) face is often oriented toward the plasma membrane.
The transmost region is a network of tubular structures known asthe trans-Golgi network (TGN)
Functions of the golgi apparatus
Proteins and Lipids
The Golgi apparatus is the site of post translational modification and sorting of newly synthesized proteins and lipids.
Glycoproteins
Further modification of the carbohydrate moiety of glycoproteins produces complex and hybrid oligosaccharide chains. This determines which proteins remain in the Golgi apparatus or leave the Golgi apparatus to become secretory proteins, lysosomal proteins, or part of the plasma membrane. Two diseasesare caused by a breakdown in this process,I-cell disease and
hyperproinsulinemia .
I-Cell Disease
- Phosphorylation of mannose in glycoprotein targets proteins to Iysosome. Phosphate is addedin a two-step sequence of reactions that are catalyzed by N-acetylglucosamine-phosphotransferase and N-acetylglucosaminidases.
- A deficiency in N-acetylglucosamine-phosphotransferases results in I-cell disease(mucolipidosis II),in which a whole family of enzyme is sent to the wrong destination. It is characterized by huge inclusion bodies in cells caused by the accumulation of undegraded glycoconjugates in Iysosomes missing the hydrolase that normally degrade these macromolecules. The missing enzymes are found in the plasma and other body fluids, where they have normal levels of activity. The absenceof the mannose-6-phosphotase on the hydrolase results in their secretion rather than their incorporation into Iysosomes.
- The disease results in skeletal abnormalities,coarse features, restricted joint movements, and psychomotor retardation. Symptoms are generally noted at birth, and the life span is less than 10 years.
- A somewhat less severe form of the disease with a later onset and potential survival into adulthood is called pseudo-Hurler polydystrophy.
- There is no treatment for either disease, but prenatal diagnosis is available.
Hyperproinsulinemisia
Characterized by elevated levels of proinsulin in the
serum resulting from the failure of a peptidase to cleave proinsulin to insulin and C- peptide in the Golgi apparatus.
The clinical manifestations are similar to those seen in patients with noninsulin dependent diabetes.
Lysosomes
Lysosomes are spherical membrane-enclosed organelles that are approximately 0.5 micro.m in diameter and contain enzymes required for intracellular digestion.
Lysosomes consist of two forms:
- Primary Iysosomes have not yet acquired the materials to be digested. They are formed by budding from the trans side of the Golgi apparatus.
- Secondary Iysosomes are formed by the fusion of the primary lysosome with the substrate to be degraded and have contents that are in various stages of degradation.
- Lysosomes contain approximately 60 hydrolytic enzymes. These include nucleases for degrading DNA and RNA, lipases for degrading lipids, glycosidases for degrading glycoconjugates (glycoproteins, proteoglycans, and glycolipids), proteases and peptidases for degrading proteins, and a variety of phosphatases. . All lysosomal enzymes are acid hydrolases, with optimal activity at a pH of approximately 5.0.
- The synthesis of the lysosomal hydrolases occurs in the RER; the hydrolases are transferred to the Golgi apparatus, where they are modified and packaged into lysosomes.
Peroxisomes
- Peroxisomes are a heterogeneous group of small, spherical organelles with a single membrane and a diameter that ranges from approximately 0.15 to 0.5 micro.m.
- Peroxisomes contain a number of enzymes that transfer hydrogen atoms from organic substrates (urate, D-amino acids, and very long chain fatty acids) to molecular oxygen with the formation of hydrogen peroxide. Catalase, the major peroxisomal protein, degrades the hydrogen peroxide to water and oxygen.
- Peroxisomal enzymes are synthesized on free polysomes. After translation, the enzymes are incorporated directly into peroxisomes.
Peroxisomes have several functions:
- Synthesis and degradation of hydrogen peroxide .
- Beta-0xidation of very long chain fatty acids (>C24) starts in the peroxisome and proceeds until the carbon chain has been reduced to a length of approximately 10carbons. Oxidation of the residual 10 carbons is completed in the mitochondria.
- Phospholipid exchange-peroxisomes contain enzymes that convert phosphatidylserineand phosphatidylethanolamine.
- Bile acid synthesis
Peroxisome Deficiency
Several genetic diseases are associated with the impairment or absence of peroxisomes. These patients fail to oxidize very long chain fatty acids and accumulate bile acid precursor.The four most common disorders are:
- Zellweger (cerebrohepatorenal) syndrome
- Neonatal adrenoleukodystrophy
- Infantile Refsum disease
- Hyperpipecolatemia
Mitochondria
Mitochondria have two membranes. They are about 0.5 micro.m in width and vary in length from 1 to 10 micro.m. They synthesize adenosine triphosphate (ATP), contain their own double-stranded circular DNA, and make some of their own proteins.
Mitochondria have several compartments.
Outer membrane
- The outer membrane is smooth, continuous, and highly permeable. It contains an abundance of porin, an integral membrane protein that forms channels in the outer membrane through which molecules of less than 10kD can pass.
Inner membrane
- The inner membrane is impermeable to most small ions (Na+, K+, H+) and small molecules (ATP, adenosine diphosphate, pyruvate). The impermeability is likely related to the high content of the lipid cardiolipin.
- The inner membrane has numerous infoldings, called cristae. The cristae greatly increase the total surface area. They contain the enzymes for electron transport and oxidative phosphorylation.
- The number of mitochondria and the number of cristae per mitochondrion are proportional to the metabolic activity of the cells in which they reside.
Intermembrane compartment
The intermembrane compartment is the space between the inner and outer membranes. It contains enzymes that use ATP to phosphorylate other nucleotides (creatine phosphokinase and adenylate kinase).
Matrix
The matrix is enclosed by the inner membrane and contains:
- Dehydrogenases-oxidize many of the substrates in the cell (pyruvate, amino acids, fatty acids), generating reduced nicotinamide adenine dinucleotide (NADH) and reduced flavin adenine dinucleotide (FADHz) for use by the electron transport chain and energy generation.
- A double-stranded circular DNA genome encodes a few of the mitochondrial proteins. Mitochondrial DNA is always inherited from the mother, resulting in the maternal transmission of diseases of energy metabolism.
- RNA,proteins, and ribosomes-although there is some protein synthesis, most mitochondrial proteins are synthesized in the cytoplasm and are transferred into the mitochondria.
- Intramitochondrial granules-contain calcium and magnesium. Their function is not known, but it is believed that they may represent a storage site for calcium.
Cytoskeleton
The cytoskeleton provides a supportive network of tubules and filaments in the cytoplasm of eukaryotic cells. It is composed of microtubules, intermediate filaments, and microfilaments.
Microtubules
- Microtubules are polymers of tubulin that undergo rapid assembly and disassembly. They are found in the cytoplasmic matrix of all eukaryotic cells.
- Tubulin
- The major component of microtubules is tubulin, a protein dimer composed of two different polypeptides, a-tubulin and l3-tubulin.
- Polymerization of tubulin to form microtubules is accomplished by microtubule organizing centers and two types of accessory proteins, tau proteins and microtubule-associated proteins. Microtubules grow from the organizing centers. Calcium ions can block or reverse polymerization.
- Microtubules playa role in:
- Chromosomal movement during meiosis and mitosis. Microtubule assembly is an
- important event in spindle formation.
- Intracellular vesicle and organelle transport. Two specific microtubule-dependent ATPases, kinesin and dynein, are involved in generating the force that drives transport, with the microtubular structure playing a more passive role in intracellular transport.
- Ciliary and flagellar movement.
Intermediate filaments
- Intermediate filaments are intermediate in thickness (10-nm diameter) between microtubules and micro filaments. They function primarily in structural roles and contain several types of tissue- specific proteins:
- Cytokeratins-found in epithelial tissue .
- Desmin-found in smooth muscle; Z disks of skeletal and cardiac muscle
- Vimentin-found in cells of mesenchymal origin (endothelial cells, fibroblasts, chondroblasts, vascular smooth muscle) .
- NeurofIlaments-found in neurons
- Glial fibrillary acidic protein (GFA)-found in astrocytes
Microfilaments
- Microfilaments have a diameter of 6 nm and are composed of actin. Each actin filament (F-actin) consists of two strands of actin twisted into a helical pattern with 13.5 molecules of globular actin (G-actin) per turn of the helix.
- Two types of movement are associated with microfilaments:
- Local movement takes advantage of the polymerization and depolymerization properties of microfilaments.
- Sliding filament movement is generated by the interaction of actin filaments with myosin filaments.
Clinical correlates
Chediak-Higashi syndrome
- Chediak-Higashi syndrome is characterized by defect in microtubule polymerization.
- This leads to defects in cytoplasmic granules including:
- Delayed fusion of phagosomes with Iysosomes in leukocyte,thus preventing phagocytosis of bacteria.
- Increased fusion of melanosomes in melanocytes, leading to albinism.
- Granular defects in natural killer cells and platelets.
Actin-binding drugs
- Actin-binding drugs(e.g. cytochalasin B) can interfere with the polymerization-depolymerization cycle of microfilaments. Process such as endocytosis, pagocytosis, cytokinesis, cytoplasmic and amoeboid movements are all inhibited by cytochalasin B.
CELL SURFACE
Basement Membrane
- The basement membrane is a sheet like structure that underlies virtually all epithelia. It consists of the following:
- Basal lamina-composed of type IV collagen, glycoproteins (e.g.,laminin), and proteoglycans (e.g., heparan sulfate).
- Reticular lamina-composed of delicate reticular fibers.
Lateral Surface
Tight junction(zonula occludens)
- The tight junction is formed by the fusion of opposed cell membranes. These ridges of fusion present as "sealing strands" seen in freeze-fracture replicas. It extends completely around the apical cell borders to seal the underlying intercellular clefts from contact with the outside environment. It constitutes the anatomic component of many barriers in the body.
Zonula adherens
- A zonula adherens (adherent junction) often lies basal to the zonula occludens.
- It is a band like junction that serves in the attachment of adjacent epithelial cells.
Desmosome
- The desmosome (macula adherens) is formed by the juxtaposition of two disk-shaped plaques
- contained within the cytoplasm of each adjacent cell .
- Intermediate filaments (tonofilaments) radiate away from the plaques . These intermediate filaments are anchored by desmoplakins (plaques)that also bind to trans membrane linker proteins, linking adjacent cells.
- Desmosomes are most common in lining membranes, are subject to wear and tear, and are considered spot welds that hold cells together.
Gap junction
- The gap junction is an area of communication between adjacent cells that allows the passage of very small particles and ions across a small intercellular gap within the junction.
- The gap junction consists of a hexagonal lattice of tubular protein subunits called connexons, which form hydrophilic channels connecting the cytoplasm of adjacent cells.
- This permits the direct passage of ions and small molecules between cells to conduct electrical impulses.
Apical (Free) Surface
Microvilli
- Microvilli are apical cell surface evaginations of cell membranes that function to increase the cell surface area available for absorption. A thick glycocalyx coat covers them. The core of each microvillus contains actin microfilaments. It is anchored in the apical cell cytoplasm to the terminal web, which itself is anchored to the zonula adherens of the cell membrane.
Cilia
- Cilia are apical cell surface projections of cell membrane that contain microtubules. They are inserted on centriole-like basal bodies present below the membrane surface at the apical pole.
- Cilia contain two central microtubules surrounded by a circle of nine peripheral microtubule doublets. The peripheral doublets are fused so that they share a common tubule wall and form two subtubules, A and B. Adjacent doublets are connected to one another by nexin links.
Movement of Cilia
- A pair of dynein arms is attached to each A subtubule. The arms bind to ATP and rearrange themselves so that a binding site for the B subtubule in the tip of the arm is exposed. The B tubule interacts with the binding site, causing the arm to snap back and movement to occur. Each cycle of a single dynein arm slides adjacent doublets 10nm past each other.
- Cilia move back and forth to propel fluid and particles in one direction. They are important in clearing mucus from the respiratory tract.
Clinical correlates:
- Stereocilia are elongated microvilli found at the apices of cells lining the epididymis, ductus deferens, and haircells of the inner ear, where they play a role in auditory sensation.
- Flagella are longer than cilia but have the same microstructure, a prominent example is in the sperm, where the single flagellum provides motility.
Kartagener Syndrome
- Absent or aberrant dynein arms are found in the cilia and flagella of individuals suffering from Kartagener syndrome (a subset of immotile cilia syndrome).
- Such individuals often have chronic sinusitis and bronchiectasis as well as infertility and, in some cases, situsinversus.
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