Our current understanding of the molecular functioning of eukaryotic cells largely rests on studies of just a few types of organisms, termed model organisms (Figure 1). Because of the evolutionary conservation of genes, proteins, organelles, cell types, and so forth, discoveries about biological structures and functions obtained with one experimental organism often apply to others. Thus researchers generally conduct studies with the organism that is most suitable for rapidly and completely answering the question being posed, knowing that the results obtained in one organism are likely to be broadly applicable. Indeed, many organisms, particularly rats, frogs, sea urchins, chickens, and slime molds, have been and continue to be immensely valuable for cell biology research. As more and more organisms have their entire genomes sequenced, a wide variety of other species are increasingly being used for investigations, especially for studies of the evolution of genes, cells, and organisms and of how organisms become adapted to diverse ecological niches.
Fig1. Each eukaryotic organism used in cell biology has advantages for certain types of studies. The yeast Saccharomyces cerevisiae (a) has the cellular organization of a eukaryote but is a relatively simple single-celled organism that is easy to grow and to manipulate genetically. The green alga Chlamydomonas reinhardtii (b) is widely used to study photosynthesis and the structure and function of flagella. In the roundworm Caenorhabditis elegans (c), which has a small number of cells arranged in a nearly identical way in every worm, the formation of each individual cell can be traced. The fruit fly Drosophila melanogaster (d), first used to discover the properties of chromosomes, has been especially valuable in identifying genes that control embryonic development. Many of these genes are evolutionarily conserved in humans. Planaria (e) are flatworms that can regenerate any part of the body that is cut off, including the head and the photoreceptors. The stem cells that give rise to their new cells and tissues are widely studied. The zebrafish Danio rerio (f) is used for rapid genetic screens to identify genes that control vertebrate development and organogenesis. Of the experimental animal systems, mice (Mus musculus) (g) are evolutionarily the closest to humans and have thus provided models for studying numerous human genetic and infectious diseases. The mustard-family weed Arabidopsis thaliana (h) has been used for genetic screens to identify genes involved in nearly every aspect of plant life. [Part (a) Sci mat/Photo Researchers, Inc. Part (b) William Dentler University of Kansas. Part (c) Science Source. Part (d) Darwin Dale/Science Source. Part (e) Peter Reddien, MIT Whitehead Institute. Part (f) blickwinkel/Hartl/Alamy. Part (g) J. M. Labat/ Jacana/Photo Researchers, Inc. Part (h) Darwin Dale/Science Source.]
As we have seen, bacteria are excellent models for studies of several cellular functions, but they lack the organelles found in eukaryotes. Unicellular eukaryotes such as yeasts are used to study many fundamental aspects of eukaryotic cell structure and function. Metazoan models such as the roundworm, fruit fly, and mouse are required to study more complex tissue and organ systems and development. As we will see in this section and the next, several eukaryotic model organisms are widely used to understand complex cell systems and mechanisms.
Yeasts Are Used to Study Fundamental Aspects of Eukaryotic Cell Structure and Function
One group of single-celled eukaryotes, the yeasts, has proven exceptionally useful in molecular and genetic analysis of eukaryotic cell formation and function. Yeasts and their multi cellular cousins, the molds, which collectively constitute the fungi, have an important ecological role in breaking down plant and animal remains for reuse. They also make numerous antibiotics and are used in the manufacture of bread, beer, and wine.
The common yeast used to make bread and beer, Saccharomyces cerevisiae, appears frequently in this book because it has proved to be an extremely useful experimental organism. Homologs of many of the approximately 6000 different proteins expressed in an S. cerevisiae cell (see Table 1) are found in most, if not all, eukaryotes and are important for cell division or for the functioning of individual eukaryotic organelles. Much of what we know of the proteins in the endoplasmic reticulum and Golgi complex that promote protein secretion was elucidated first in yeasts. Yeasts were also essential for the identification of many proteins that regulate the cell cycle and catalyze DNA replication and transcription. S. cerevisiae (Figure 2a; see also Figure 1a) and other yeasts offer many advantages to molecular and cellular biologists:
● Vast numbers of yeast cells can be grown easily and cheaply in culture from a single cell; the cells in such clones are genetically identical and have the same biochemical properties. Individual proteins or multiprotein complexes can be purified from large amounts of cells and then studied in detail.
● Yeast cells may be either haploid (containing one copy of each chromosome) or diploid (containing two copies of each chromosome), and both forms can divide by mitosis; this ability makes isolating and characterizing mutations in genes encoding essential yeast cell proteins relatively straightfor ward.
● Yeasts, like many organisms, have a sexual cycle that al lows exchange of genes between cells. Under starvation conditions, diploid cells undergo meiosis to form haploid daughter cells, which are of two types, a and α cells. If haploid a and α cells encounter each other, they can fuse, forming an a/α diploid cell that contains two copies of each chromosome, one from each parent cell (Figure 2b).
Table1. Genome Sizes of Organisms Used in Molecular Cell Biology Research That Have Been Completely Sequenced
Fig2. The yeast Saccharomyces cerevisiae can be haploid or diploid and can reproduce sexually or asexually. (a) Scanning electron micrograph of the budding yeast Saccharomyces cerevisiae. These cells grow by an unusual type of mitosis termed mitotic budding. One daughter nucleus remains in the “mother” cell; the other daughter nucleus is transported into the bud, which grows in size and soon is released as a new cell. After each bud cell breaks free, a scar is left at the budding site, so the number of previous buds on the parent cell can be counted. The orange-colored cells are bacteria. (b) Haploid yeast cells can have different mating types, called a (blue) and α (orange). Both types contain a single copy of each yeast chromosome, half the usual number, and grow by mitotic budding. Two haploid cells that differ in mating type, one a and one α, can fuse together to form an a/α diploid cell that contains two copies of each chromosome; diploid cells can multiply by mitotic budding. Under starvation conditions, a diploid cell can undergo meiosis, a special type of cell division, to form four haploid ascospores. Rupture of an ascus releases four haploid spores, which can germinate into haploid a and α cells. These cells can also multiply asexually. [Part (a) SCIMAT/Science Source.]
With the use of a single species such as S. cerevisiae as a model organism, results from studies carried out by tens of thousands of scientists worldwide, using multiple experimental techniques, can be combined to yield a deeper level of understanding of a single type of cell. As we will see many times in this book, conclusions based on studies of S. cerevisiae have often proved true for all eukaryotes and have formed the basis for exploring the evolution of more com plex processes in multicellular animals and plants.
Mutations in Yeast Led to the Identification of Key Cell Cycle Proteins
Biochemical studies can tell us much about an individual protein, but they cannot prove that it is required for cell di vision or any other cell process. The importance of a protein is demonstrated most firmly if a mutation that prevents its synthesis or makes it nonfunctional adversely affects the process under study.
In a classical genetics approach, scientists isolate and characterize mutants that lack the ability to do something a normal organism can do. Often large genetic “screens” are done to look for many different mutant individuals (e.g., fruit flies, yeast cells) that are unable to complete a certain process, such as cell division or muscle formation. Mutations are usually produced by treatment with a mutagen, a chemical or physical agent that promotes mutations in a largely random fashion. But how can we isolate and maintain mu tant organisms or cells that are defective in some process, such as cell division or protein secretion, that is essential for survival?
One way is to isolate organisms with a temperature sensitive mutation. These mutants are able to grow at the permissive temperature, but not at another, usually higher temperature, the nonpermissive temperature. Normal cells can grow at either temperature. In most cases, a temperature-sensitive mutant produces an altered protein that works at the permissive temperature but unfolds and is nonfunctional at the nonpermissive temperature. Screens for temperature-sensitive mutations are most readily done with haploid organisms such as yeasts because they have only one copy of each gene, and thus a mutation in it will immediately have a consequence.
By analyzing the effects of numerous different temperature-sensitive mutations that altered the division of haploid yeast cells, geneticists discovered most of the genes necessary for cell division without knowing anything, initially, about which proteins they encode or how these proteins participate in the process. In general, the great power of genetics is to reveal the existence and relevance of all proteins re quired for a particular cell function without prior knowledge of their biochemical identity or molecular function. These “mutation-defined” genes can be isolated and replicated (cloned) with recombinant DNA techniques. With the isolated genes in hand, the encoded proteins can be produced in a test tube or in engineered bacteria or cultured cells. In this way, biochemists can investigate whether the genes necessary for cell division encode proteins that associate with other proteins or DNA or catalyze particular chemical reactions during cell division.
Most of these yeast cell cycle genes are found in human cells as well, and the encoded proteins have similar amino acid sequences. Proteins from different organisms, but with similar amino acid sequences, are said to be homologous; such proteins may have the same or similar functions. Remarkably, it has been shown that a human cell cycle protein, when expressed in a mutant yeast defective in the homologous yeast protein, is able to “rescue the defect” of the mu tant yeast (that is, to allow the cell to grow normally), thus demonstrating the protein’s ability to function in a very different type of eukaryotic cell. This experimental result, which garnered a Nobel Prize for Paul Nurse, was especially notable because the common ancestor of present-day yeasts, plants, and humans is thought to have lived over a billion years ago. Clearly the eukaryotic cell cycle and many of the genes and proteins that catalyze and regulate it evolved early in biological evolution and have remained quite constant over a very long period of evolutionary time. Subsequent studies showed that mutations in many yeast cell cycle proteins that allow uncontrolled cell growth also frequently occur in human cancers, again attesting to the important conserved functions of these proteins in all eukaryotes.
Studies in the Alga Chlamydomonas reinhardtii Led to the Development of a Powerful Technique to Study Brain Function
The green unicellular alga Chlamydomonas reinhardtii (Figure 1b), which swims using its two long flagella, is widely used in studies of the structure, function, and assembly of this organelle. In part because of the powerful genetic techniques now available, Chlamydomonas is also used in studies of chloroplast formation and photosynthesis. The Chlamydomonas genome (see Table 1) encodes many more proteins than do those of yeasts, including flagellar proteins and proteins needed to build a chloroplast, organelles not found in yeasts.
One important outcome of the use of this experimental organism came from studies of phototaxis, the behavior in which an organism moves toward or away from a source of light. Chlamydomonas needs to move toward light to undergo photosynthesis and thus generate the energy it needs to grow and divide, but light that is too intense repels it, as it causes damage to the chloroplast. Studies of Chlamydomonas phototaxis led to the discovery of two proteins in its plasma membrane that, when they absorb light, open a “channel” in the membrane that allows ions such as Ca2+ to flow from the extracellular medium into the cytosol, triggering phototactic responses. recombinant DNA techniques have been used to express one such protein in specific neurons in the mouse brain, allowing investigators to activate just one or a few cells in the brain using a point source of light. Thus studies on this humble alga have led to the development of an important experimental system— optogenetics—for the study of brain function.
The Parasite That Causes Malaria Has Novel Organelles That Allow It to Undergo a Remarkable Life Cycle
Whereas yeasts are used in the manufacture of bread, beer, wine, and cheese, some unicellular eukaryotes cause major human diseases and are widely studied in an attempt to develop drugs that will kill them but not injure their human host. Entamoeba histolytica causes dysentery; Trichomonas vaginalis, vaginitis; and Trypanosoma brucei, sleeping sickness. Each year the worst of these protozoans, Plasmodium falciparum and related species, cause more than 300 million new cases of malaria, a disease that kills 1.5 million to 3 million people annually. These protozoans inhabit mammals and mosquitoes alternately, changing their morphology and behavior in response to signals in each of these environments.
The complex life cycle of Plasmodium dramatically il lustrates how a single cell can adapt to multiple challenges (Figure 3a). Additionally, the merozoite form that infects human red blood cells contains several organelles, not found in most eukaryotes, that enable the parasite to invade a red blood cell, including the rhoptry, polar ring, and microneme, as well as a fuzzy surface coat on the outside of the cell (Figure 3b, c). Entry of the parasite into a red blood cell is initiated by binding of certain parasite cell-surface proteins to proteins on the red blood cell surface, followed by the formation of a tight junction between the two plasma membranes, the loss of the “fuzzy coat,” and secretion of proteins stored in the microneme and rhoptry.
Fig3. Plasmodium species, the parasites that cause malaria, are single-celled protozoans with a remarkable life cycle. Many Plasmodium species are known, and they can infect a variety of animals, cycling between insect and vertebrate hosts. The four species that cause malaria in humans undergo several dramatic transformations within their human and mosquito hosts. (a) Diagram of the life cycle. Step 1: Sporozoites enter a human host when an infected Anopheles mosquito bites a person. Step 2: They migrate to the liver, where they develop into merozoites, which are released into the blood. Merozoites differ substantially from sporozoites, so this transformation is a metamorphosis (Greek, “to transform” or “many shapes”). Step 3: Circulating merozoites invade red blood cells (RBCs) and re produce within them. Proteins produced by some Plasmodium species move to the surface of infected RBCs, causing the cells to adhere to the walls of blood vessels. This prevents infected RBCs from circulating to the spleen, where cells of the immune system would destroy the RBCs and the Plasmodium organisms they harbor. Step 4: After growing and reproducing in RBCs for a period of time characteristic of each Plasmodium species, the merozoites suddenly burst forth in synchrony from large numbers of infected cells. It is this event that brings on the fevers and shaking chills that are the well-known symptoms of malaria. Some of the released merozoites infect additional RBCs, creating a cycle of production and infection. Step 5: Eventually, some merozoites undergo meiosis and develop into male and female gametocytes, another metamorphosis. These cells, which contain half the usual number of chromosomes, cannot survive for long unless they are transferred in blood to an Anopheles mosquito. Step 6: In the mosquito’s stomach, the gametocytes are transformed into sperm or eggs (gametes), yet another metamorphosis marked by development of long hairlike flagella on the sperm. Step 7: Fusion of sperm and eggs generates zygotes, which implant into the cells of the stomach wall and grow into oocysts, essentially factories for producing sporozoites. Step 8: Rupture of an oocyst releases thousands of sporozoites, which migrate to the salivary glands, setting the stage for infection of another hu man host. (b) Organelles of the Plasmodium vivax merozoite. Some of these organelles are found only in Plasmodium and related eukaryotic parasitic microorganisms. (c) Section of a Plasmodium vivax merozoite invading a human red blood cell. See A. Cowman and B. Crabb, 2006, Cell 124:755–766. [Part (c) Masamichi Aikawa.]
All the transformations in cell type that occur during the Plasmodium life cycle are governed by instructions encoded in the genetic material of this parasite (see Table 1). The Plasmodium genome has about the same number of protein-coding genes as the yeast Saccharomyces cerevisiae, but about two-thirds of the Plasmodium genes appear to be unique to this and related parasites, attesting to the great evolutionary distance between these parasites, the Apicomplexa, and most other eukaryotes as well as the presence of unusual organelles required for their complex life cycles.
