5:  5.4 Organelles of the Eukaryotic Cell

Lysosomes Are Acidic Organelles That Contain a Battery of Degradative Enzymes

Lysosomes provide an excellent example of the ability of intracellular membranes to form closed compartments in which the composition of the lumen (the aqueous interior of the compartment) differs substantially from that of the surrounding cytosol. Found in animal cells, lysosomes are bounded by a single membrane and are responsible for degrading certain components that have become obsolete for the cell or organism. In some cases, materials taken into a cell by endocytosis or phagocytosis also are degraded in lysosomes. Endocytosis refers to the process by which extracellular materials are taken up by invagination of a segment of the plasma membrane to form a small membrane-bounded vesicle (endosome). In phagocytosis, relatively large particles are enveloped by the plasma membrane and internalized.

Lysosomes contain a group of enzymes that degrade polymers into their monomeric subunits. For example, nucleases degrade RNA and DNA into their mononucleotide building blocks; proteases degrade a variety of proteins and peptides; phosphatases remove phosphate groups from mononucleotides, phospholipids, and other compounds; still other enzymes degrade complex polysaccharides and lipids into smaller units. Tay-Sachs disease is caused by a defect in one enzyme catalyzing a step in the lysosomal breakdown of certain glycolipids called gangliosides, which are abundant in nerve cells—with devastating consequences. The symptoms of this inherited disease usually are evident before the age of 1. Affected children commonly become demented and blind by age 2, and die before their third birthday. Nerve cells from such children are greatly enlarged with swollen lipid-filled lysosomes.

All the lysosomal enzymes work most efficiently at acid pH values and collectively are termed acid hydrolases. A hydrogen ion pump and a Cl⁻ channel protein in the lysosomal membrane maintain the pH of the interior at ≈4.8. The pump hydrolyzes ATP and uses the released free energy to pump H⁺ ions from the cytosol into the lumen of the lysosome; the Cl⁻ channel allows Cl⁻ ions to enter. Together they transport HCl (Chapter 15). The acid pH helps to denature proteins, making them accessible to the action of the lysosomal hydrolases, which themselves are resistant to acid denaturation. Lysosomal enzymes are poorly active at the neutral pH values of cells and most extracellular fluids. Thus if a lysosome releases its contents into the cytosol, where the pH is between 7.0 and 7.3, little degradation of cytosolic components takes place.

Lysosomes vary in size and shape, and several hundred may be present in a typical animal cell. In effect, they function as sites where various materials to be degraded collect (Figure 5-44a). Primary lysosomes are roughly spherical and do not contain obvious particulate or membrane debris. Secondary lysosomes, which are larger and irregularly shaped, appear to result from the fusion of primary lysosomes with other membrane organelles; they contain particles or membranes in the process of being digested (Figure 5-44b). The process by which an aged organelle is degraded in a lysosome is called autophagy (“eating oneself”).

Plant Vacuoles Store Small Molecules and Enable the Cell to Elongate Rapidly

Most plant cells contain at least one membrane-limited internal vacuole (see Figure 5-43). The number and size of vacuoles depend on both the type of cell and its stage of development; a single vacuole may occupy as much as 80 percent of a mature plant cell. Plant cells store water, ions, and nutrients such as sucrose and amino acids within these vacuoles. We will see in Chapter 15 how such materials are accumulated in vacuoles. Vacuoles also act as receptacles for waste products and excess salts taken up by the plant and may function similarly to lysosomes in animal cells. Like lysosomes, vacuoles have an acidic pH, maintained by a proton pump and a Cl⁻ channel protein in the vacuole membrane, and contain a battery of degradative enzymes. Similar storage vacuoles are found in green algae and many microorganisms such as yeast.

Like most cellular membranes, the vacuolar membrane is permeable to water but is poorly permeable to the small molecules stored within it. Because the solute concentration is much higher in the vacuole lumen than in the cytosol or extracellular fluids, water tends to move by osmotic flow into vacuoles, just as it moves into cells placed in a hypotonic medium (see Figure 5-22). This influx of water causes both the vacuole to expand and water to move into the cell from the wall, creating hydrostatic pressure, or turgor, inside the cell. This pressure is balanced by the mechanical resistance of the cellulose-containing cell wall that surrounds plant cells. Most plant cells have a turgor of 5 – 20 atmospheres (atm); their cell walls must be strong enough to react to this pressure in a controlled way. Unlike animal cells, plant cells can elongate extremely rapidly — at rates of 20 – 75 μm/h. This elongation, which usually accompanies plant growth, occurs when a segment of the somewhat elastic cell wall stretches under the pressure created by water taken into the vacuole.

Peroxisomes Degrade Fatty Acids and Toxic Compounds

All animal cells (except erythrocytes) and many plant cells contain peroxisomes, a class of small organelles (≈0.2 – 1 μm in diameter) bounded by a single membrane (see Figure 5-25c). (Glyoxisomes are similar organelles found in plant seeds that oxidize stored lipids as a source of carbon and energy for growth. They contain many of the same types of enzymes as peroxisomes as well as additional ones used to convert fatty acids to glucose precursors.) Peroxisomes contain several oxidases — enzymes that use molecular oxygen to oxidize organic substances, in the process forming hydrogen peroxide (H₂O₂), a corrosive substance. Peroxisomes also contain copious amounts of the enzyme catalase, which degrades hydrogen peroxide to yield water and oxygen:

In contrast to oxidation of fatty acids in mitochondria, which produces CO₂ and is coupled to generation of ATP, peroxisomal oxidation of fatty acids yields acetyl groups and is not linked to ATP formation. The energy released during peroxisomal oxidation is converted to heat, and the acetyl groups are transported into the cytosol, where they are used in the synthesis of cholesterol and other metabolites. In most eukaryotic cells, the peroxisome is the principal organelle in which fatty acids are oxidized, thereby generating precursors for important biosynthetic pathways. Particularly in liver and kidney cells, various toxic molecules that enter the bloodstream also are degraded in peroxisomes, producing harmless products.

In the human genetic disease X-linked adrenoleukodystrophy (ADL), peroxisomal oxidation of very long chain fatty acids is defective. The ADL gene encodes the peroxisomal membrane protein that transports into peroxisomes an enzyme required for oxidation of these fatty acids. Individuals with the severe form of ADL are unaffected until mid-childhood, when severe neurological disorders appear, followed by death within a few years.

Mitochondria Are the Principal Sites of ATP Production in Aerobic Cells

Most eukaryotic cells contain many mitochondria, which occupy up to 25 percent of the volume of the cytoplasm. These complex organelles, the main sites of ATP production during aerobic metabolism, are among the largest organelles, generally exceeded in size only by the nucleus, vacuoles, and chloroplasts.

Mitochondria contain two very different membranes, an outer one and an inner one, separated by the intermembrane space (Figure 5-45; see also 5-31). The outer membrane, composed of about half lipid and half protein, contains proteins that render the membrane permeable to molecules having molecular weights as high as 10,000. In this respect, the outer membrane is similar to the outer membrane of gram-negative bacteria (see Figure 1-7a). The inner membrane, which is much less permeable, is about 20 percent lipid and 80 percent protein — a higher proportion of protein than occurs in other cellular membranes. The surface area of the inner membrane is greatly increased by a large number of infoldings, or cristae, that protrude into the matrix, or central space.

In nonphotosynthetic cells, the principal fuels for ATP synthesis are fatty acids and glucose. The complete aerobic degradation of glucose to CO₂ and H₂O is coupled to synthesis of as many as 36 molecules of ATP (Chapter 2). In eukaryotic cells, the initial stages of glucose degradation occur in the cytosol, where two ATP molecules per glucose molecule are generated. The terminal stages, including those involving phosphorylation coupled to final oxidation by oxygen, are carried out by enzymes in the mitochondrial matrix and cristae (Chapter 16). As many as 34 ATP molecules per glucose molecule are generated in mitochondria, although this value can vary because much of the energy released in mitochondrial oxidation can be used for other purposes (e.g., heat generation and the transport of molecules into or out of the mitochondrion), making less energy available for ATP synthesis. Similarly, virtually all the ATP formed during the oxidation of fatty acids to CO₂ is generated in the mitochondrion. Thus the mitochondrion can be regarded as the “power plant” of the cell.

Chloroplasts, the Sites of Photosynthesis, Contain Three Membrane-Limited Compartments

Except for vacuoles, chloroplasts are the largest and most characteristic organelles in the cells of plants and green algae. They can be as long as 10 μm and are typically 0.5 – 2 μm thick, but they vary in size and shape in different cells, especially among the algae. Like the mitochondrion, the chloroplast is surrounded by an outer and an inner membrane (see Figure 5-31). Chloroplasts also contain an extensive internal system of interconnected membrane-limited sacs called thylakoids, which are flattened to form disks; these often are grouped in stacks called grana and embedded in a matrix, the stroma (Figure 5-46). The thylakoid membranes contain green pigments (chlorophylls) and other pigments and enzymes that absorb light and generate ATP during photosynthesis. Part of this ATP is used by enzymes located in the stroma to convert CO₂ into three-carbon intermediates; these are then exported to the cytosol and converted to sugars.

Perhaps surprisingly, the molecular mechanisms by which ATP is formed in mitochondria and chloroplasts are very similar. The details of these critical ATP-generating pathways are discussed in Chapter 16. Chloroplasts and mitochondria share other features: Both often migrate from place to place within cells and also contain their own DNA, which encodes some of the key organellar proteins (Chapter 9). The proteins encoded by mitochondrial or chloroplast DNA are synthesized on ribosomes within the organelles. However, most of the proteins in each organelle are encoded in nuclear DNA and are synthesized in the cytosol; these proteins then are incorporated into the organelles by processes described in Chapter 17.

The Endoplasmic Reticulum Is a Network of Interconnected Internal Membranes

Generally, the largest membrane in a eukaryotic cell encloses the endoplasmic reticulum (ER) — a compartment comprising a network of interconnected, closed, membrane-bounded vesicles (Figure 5-47). The endoplasmic reticulum has a number of functions in the cell but is particularly important in the synthesis of many membrane lipids and proteins. The smooth endoplasmic reticulum is smooth because it lacks ribosomes; regions of the rough endoplasmic reticulum are studded with ribosomes.

Golgi Vesicles Process and Sort Secretory and Membrane Proteins

Several minutes after proteins are synthesized in the rough ER, most of them leave the organelle within small membrane-bounded transport vesicles. These vesicles, which bud off from regions of the rough ER not coated with ribosomes, carry the proteins to the luminal cavity of another membrane-limited organelle, the Golgi complex, a series of flattened sacs located near the nucleus in many cells (see Figure 5-48a).

Three-dimensional reconstructions from serial sections of a Golgi complex reveal a series of flattened membrane vesicles or sacs, surrounded by a number of more or less spherical membrane vesicles (Figure 5-49). The stack of flattened Golgi sacs has three defined regions — the cis, the medial, and the trans. Transfer vesicles from the rough ER fuse with the cis region of the Golgi complex, where they deposit their proteins. As detailed in Chapter 17, these proteins then progress from the cis to the medial to the trans region. Within each region are different enzymes that modify secretory and membrane proteins differently, depending on their structures and their final destinations.

After secretory proteins are modified in the Golgi sacs, they are transported out of the complex by a second set of transport vesicles, which seem to bud off the trans side of the Golgi complex. Some of these transport vesicles, termed coated vesicles, are surrounded by an outer protein cage composed primarily of the fibrous protein clathrin (see Figure 5-26). Some vesicles contain membrane proteins destined for the plasma membrane; others, proteins for lysosomes or for other organelles. How intracellular transport vesicles “know” which membranes to fuse with and where to deliver their contents is also discussed in Chapter 17.

The Double-Membraned Nucleus Contains the Nucleolus and a Fibrous Matrix

The nucleus, the largest organelle in eukaryotic cells, is surrounded by two membranes, each one a phospholipid bilayer containing many different types of proteins. The inner nuclear membrane defines the nucleus itself. In many cells, the outer nuclear membrane is continuous with the rough endoplasmic reticulum, and the space between the inner and outer nuclear membranes is continuous with the lumen of the rough endoplasmic reticulum (see Figure 5-31).

The two nuclear membranes appear to fuse at the nuclear pores (see Figure 5-42b). The distribution of nuclear pores is particularly vivid when the nucleus is viewed by the freeze-fracture technique described earlier (Figure 5-50). Constructed of a specific set of membrane proteins, these ringlike pores function as channels that regulate the movement of material between the nucleus and the cytosol (see Figure 11-28).

In a growing or differentiating cell, the nucleus is metabolically active, producing DNA and RNA. The latter is exported through nuclear pores to the cytoplasm for use in protein synthesis (Chapter 4). In mature erythrocytes from nonmammalian vertebrates and other types of “resting” cells, the nucleus is inactive or dormant and minimal synthesis of DNA and RNA takes place.

How nuclear DNA is packaged into chromosomes is described in Chapter 9. In a nucleus that is not dividing, the chromosomes are dispersed and not thick enough to be observed in the light microscope. Only during cell division are chromosomes visible by light microscopy (see Figure 1-8). However, a suborganelle of the nucleus, the nucleolus, is easily recognized under the light microscope. Most of the cell’s ribosomal RNA is synthesized in the nucleolus; some ribosomal proteins are added to ribosomal RNAs within the nucleolus as well (Chapter 11). The finished or partly finished ribosomal subunit passes through a nuclear pore into the cytosol.

In the electron microscope, the nonnucleolar regions of the nucleus, called the nucleoplasm, can be seen to have areas of high DNA concentration, often closely associated with the nuclear membrane. Fibrous proteins called lamins form a two-dimensional network along the inner surface of the inner membrane, giving it shape and apparently binding DNA to it. The breakdown of this network occurs early in cell division, as we detail in Chapter 13.

The Cytosol Contains Many Particles and Cytoskeletal Fibers

Because sections for standard electron microscopy must be thinner than 0.1 μm, fibers in the cytosol, which may be several microns in length, appear as long elements only in sections that by chance happen to be in the plane of the fiber bundles (Figure 5-51). Serial sectioning of a tissue sample can compensate for these shortcomings by tracing a fiber from one image into the next to reconstruct its threedimensional architecture. Because 200 sections are needed to examine a cell 20 μm thick, serial sectioning is a tedious technique. However, sections up to 1 μm thick can be viewed in high-voltage electron microscopes, considerably reducing the number of sections needed to reconstruct three-dimensional images.

Transmission electron micrographs of cytosolic fibers obtained from unsectioned cells reveal an extensive network of microfilaments, microtubules, and intermediate filaments (Figure 5-52). These cytoskeletal fibers crisscross one another in complex patterns so that different types of fibers contact one another at many points. In cultured cells, actin microfilaments often occur in bundles of long fibers that appear to be connected by small fibrous proteins (see Figure 5-6).

The cytosol of many cells also contains inclusion bodies, granules that are not bounded by a membrane. For instance, muscle cells and hepatocytes contain cytosolic granules of glycogen (see Figure 5-47), a glucose polymer that functions as a storage form of usable cellular energy. In well-fed animals, glycogen can account for as much as 10 percent of the wet weight of the liver. The cytosol of the specialized fat cells in adipose tissue contains large droplets of almost pure triacylglycerols, a storage form of fatty acids (see Figure 5-13a and b).

In addition, the cytosol is a major site of cellular metabolism and contains a large number of different enzymes. About 20 – 30 percent of the cytosol is protein, and from a quarter to half of the total protein within cells is in the cytosol. Because of the high concentration of cytosolic proteins, organized complexes of proteins can form even if the energy that stabilizes them is weak. Many investigators believe that the cytosol is highly organized, with most proteins either bound to fibers or otherwise localized in specific regions.

SUMMARY

• The internal architecture of cells and central metabolic pathways are similar in all plants, animals, and unicellular eukaryotic organisms (e.g., yeast). All eukaryotic cells contain a membrane-limited nucleus and numerous other organelles in their cytosol.

• Lysosomes, which are found only in animal cells, have an acidic interior and contain various hydrolases. These degradative enzymes break down some cellular components that are no longer functional or needed by the cell and some ingested materials (see Figure 5-44).

• Plant cells contain one or more large vacuoles, which often fill much of the cell. The vacuole is a storage site for ions and nutrients. Osmotic flow of water into the vacuole generates turgor pressure that pushes the plasma membrane against the cell wall.

• Peroxisomes are small organelles containing enzymes that oxidize various organic compounds, generating hydrogen peroxide. This toxic substance is converted to water and oxygen by catalase, also present in large amounts in these organelles. Oxidation of fatty acids in peroxisomes produces acetyl groups, used in biosynthetic reactions, but no ATP.

• The mitochondrion is bounded by two membranes, with the inner one extensively folded. Enzymes in the inner mitochondrial membrane and central matrix carry out the terminal stages of sugar and lipid oxidation coupled to ATP synthesis.

• Chloroplasts, the sites of photosynthesis, are surrounded by an inner and outer membrane; a complex system of thylakoid membranes in their interior contains the pigments and enzymes that absorb light and produce ATP.

• Secretory proteins and membrane proteins are synthesized on the rough endoplasmic reticulum, a network of interconnected membrane vesicles studded with ribosomes. These proteins then move to the Golgi complex, where they are sorted and processed (see Figure 5-48).

• The nucleus is surrounded by an inner and outer membrane. These contain numerous pores through which materials pass between the nucleus and cytosol. The outer nuclear membrane is continuous with the rough endoplasmic reticulum.

• The cytosol, the protein-rich fraction remaining after removal of all organelles, contains numerous soluble enzymes and three major types of protein filaments: actin microfilaments, microtubules, and intermediate filaments. In all animal and plant cells, these filaments form a complex network, the cytoskeleton, that gives the cell structural stability and contributes to cell movement.