11. Turning Genes Off and On

Being able to translate a gene into a protein is only part of gene expression. Every cell must also be able to regulate when particular genes are used. Imagine if every instrument in a symphony played at full volume all the time, all the horns blowing full blast and each drum beating as fast and loudly as it could! No symphony plays that way, because music is more than noise-it is the controlled expression of sound. In the same way, growth and development is the controlled expression of genes, each brought into play at the proper moment to achieve precise and delicate effects.
Cells control the expression of their genes by saying when individual genes are to be transcribed. At the beginning of each gene are special regulatory sites that act as points of control. Specific regulatory proteins within the cell bind to these sites, turning transcription of the gene off or on.

Repressors

Many genes are "negatively" controlled: they are turned off except when needed. In these genes, the regulatory site is located between the place where the RNA polymerase binds to the DNA (called the promoter site) and the beginning edge of the gene. When a regulatory protein called a repressor is bound to its regulatory site, the operator, its presence blocks the movement of the polymerase toward the gene. Imagine if you sat down to eat dinner and someone placed a brick wall between your chair and the table-you could not begin your meal until the wall was removed, any more than the polymerase can begin transcribing the gene until the repressor protein is removed.

To turn on a gene whose transcription is blocked by a repressor, all that is required is to remove the repressor. Cells do this by binding special "signal" molecules to the repressor protein; the binding causes the repressor protein to contort into a shape that doesn't fit DNA, and it falls off, removing the barrier to transcription. A specific example demonstrating how repressor proteins work is the set of genes called the lac operon in the bacterium Escherichia coli. An operon is a segment of DNA containing a cluster of genes that are transcribed as a unit. The operon consists of both protein-encoding (structural) genes and associated regulatory elements-the operator and promoter. When an E. coli encounters the sugar lactose, the lactose binds to the repressor protein and induces a twist in its shape that causes it to fall from the DNA. RNA polymerase is no longer blocked, so it starts to transcribe the genes needed to break down the lactose to get energy.

How the lac operon works.

(a) The lac operon is shutdown ("repressed") when the repressor protein is bound to the operator site. Because promoter and operator sites overlap, polymerase and repressor cannot bind at the same time, any more than two people can sit in one chair. (b) The lac operon is transcribed ("induced") when lactose binding to the repressor protein changes its shape so that it can no longer sit on the operator site and block polymerase binding.

Activators

Because RNA polymerase binds to a specific promoter site on one strand of the DNA double helix, it is necessary that the DNA double helix unzip in the vicinity of this site in order for the polymerase protein to be able to sit down properly. In many genes, this unzipping cannot take place without the assistance of a regulatory protein called an activator that binds to the DNA in this region and helps it unwind. Just as in the case of the repressor protein described previously, cells can turn genes on and off by binding "signal" molecules to the activator protein. These molecules prevent it from binding to the DNA or enable it to do so.

Activators enable a cell to carry out a second level of control. When a bacterium encounters the sugar lactose, it may not be low on energy. Imagine if you had to eat every time you encountered food! When a bacterial cell already has lots of energy, levels of a special "I'm hungry" signal molecule fall. Without being prodded by this signal molecule, the activator protein, called CAP, cannot twist into the proper shape to fit the DNA unwinding site in front of the lactose-using genes; as a result, the genes are not transcribed, even though the repressor protein does not block the polymerase.

Enhancers

A third level of control is exercised by expanding access to the gene. To make the promoters of complexly controlled genes accessible to many regulatory proteins simultaneously, many eukaryotic genes possess special associated sequences called enhancers. These enhancer sequences bind specific regulatory proteins that interact with the protein transcription factors that help RNA polymerase find and attach to its binding site at the beginning of the structural gene. Unlike promoters and operators, which butt right up to the start of a gene, enhancers are usually located far away from the start of the gene, often thousands of nucleotides distant. Although enhancers occur in exceptional instances in bacteria, they are the rule rather than the exception in eukaryotes.

How can regulatory proteins affect a promoter when they bind to the DNA at enhancer sites located far from the promoter? Apparently the DNA loops around so that the enhancer is positioned near the promoter. This brings the regulatory protein attached to the enhancer into direct contact with the transcription factor complex attached to the promoter.

The enhancer mode of transcriptional control that has evolved in eukaryotes adds a great deal of flexibility to the control process. The positioning of regulatory sites at a distance permits a large number of different regulatory sequences scattered about the DNA to influence that particular gene.
Key concepts: Cells control the expression of genes by saying when they are transcribed, not how fast. Some regulatory proteins block the binding of the polymerase, and others facilitate it. in eukaryotes these regulatory proteins are often associated with control genes located on the chromosome far from the gene being regulated.

12. Mutation

There are two general ways in which the genetic message is altered: mutation and recombination. A change in the content of the genetic message-the base sequence of one or more genes-is referred to as a mutation. Some mutations alter the identity of a particular nucleotide, while others remove or add nucleotides to a gene. A change in the position of a portion of the genetic message is referred to as recombination. Some recombination events move a gene to a different chromosome; others alter the location of only part of a gene. The cells of eukaryotes contain an enormous amount of DNA, and the mechanisms that protect and proofread the DNA are not perfect. If they were, no variation would be generated.

Mistakes Happen
In fact, cells do make mistakes during replication, often causing a change in a cell's genetic message, or a mutation. However, change is rare. Typically, a particular gene is altered in only one of a million gametes. If changes were common, the genetic instructions encoded in DNA would soon degrade into meaningless gibberish. Limited as it might seem, the steady trickle of change that does occur is the very stuff of evolution. Every difference in the genetic messages that specify different organisms arose as the result of genetic change.

The Importance of Genetic Change
All evolution beams with alterations in the genetic message: mutation creates new alleles, gene transfer and transposition alter gene location, reciprocal recombination shuffles and sorts these changes, and chromosomal rearrangement alters the organization of entire chromosomes. Some changes in germ-line tissue produce alterations that enable an organism to leave more offspring, and those changes tend to be preserved as the genetic endowment of future generations. Other changes reduce the ability of an organism to leave offspring. Those changes tend to be lost, as the organisms that carry them contribute fewer members to future generations.

Evolution can be viewed as the selection of particular combinations of alleles from a pool of alternatives. The rate of evolution is ultimately limited by the rate at which these alternatives are generated. Genetic change through mutation and recombination provides the raw material for evolution.

Genetic changes in somatic cells do not pass on to offspring, and so they have less evolutionary consequence than germ-line change. However, changes in the genes of somatic cells can have an important and immediate impact particularly if the gene affects development or is involved with regulation of cell proliferation. Rare changes in genes, called mutations, can have significant effects on the individual when they occur in somatic tissue, but they are inherited only if they occur in germ-line tissue. Inherited changes provide the raw material for evolution.)

13. Kinds of Mutation

Because mutations can occur randomly in a cell's DNA, most mutations are detrimental, just as making a random change in a computer program usually worsens performance. The consequences of a detrimental mutation may be minor or catastrophic, depending on the function of the altered gene.

Mutations in Germ-Line Tissues
The effect of a mutation depends critically on the identity of the cell in which the mutation occurs. During the embryonic development of all multicellular organisms, there comes a point when cells destined to form gametes (germ-line cells) are segregated from those that will form the other cells of the body (somatic cells). Only when a mutation occurs within a germ-line cell is it passed to subsequent generations as part of the hereditary endowment of the gametes derived from that cell.

Mutations in Somatic Tissues
Mutations in germ-line tissue are of enormous biological importance because they provide the raw material from which natural selection produces evolutionary change. Change can occur only if there are new, different allele combinations available to replace the old. Mutation produces new alleles, and recombination puts the alleles together in different combinations. In animals, it is the occurrence of these two processes in germ-line tissue that is important to evolution, because mutations in somatic cells (somatic mutations) are not passed from one generation to the next. However, a somatic mutation may have drastic effects on the individual organism in which it occurs, because it is passed on to all of the cells that are descended from the original mutant cell. Thus, if a mutant lung cell divides, all cells derived from it will carry the mutation. Somatic mutations of lung cells are, as we shall see, the principal cause of lung cancer in humans.

Point Mutations
One category of mutational changes affects the message itself, producing alterations in the sequence of DNA nucleotides. If alterations involve only one or a few base pairs in the coding sequence, they are called point mutations. Sometimes the identity of a base changes (base substitution), while other times one or a few bases are added (insertion) or lost (deletion). If an insertion or deletion throws the reading of the gene message out of register, a frame-shift mutation results. While some point mutations arise due to spontaneous pairing errors that occur during DNA replication, others result from damage to the DNA caused by mutagens, usually radiation or chemicals. The latter class of mutations is of particular importance because modern industrial societies often release many chemical mutagens into the environment.

Changes in Gene Position
Another category of mutations affects the way the genetic message is organized. In both bacteria and eukaryotes, individual genes may move from one place in the genome to another by transposition. When a particular gene moves to a different location, its expression or the expression of neighboring genes may be altered. In addition, large segments of chromosomes in eukaryotes may change their relative locations or undergo duplication. Such chromosomal rearrangements often have drastic effects on the expression of the genetic message. Table 8.1 reviews the effects of some categories of mutations.
Point mutations are changes in the hereditary message of an organism. They may result from spontaneous errors during DNA replication or from damage to the DNA due to radiation or chemicals.

14. Cancer and Mutation

The search for the cause of cancer has focused in part on environmental factors, including ionizing radiation such as X rays and a variety of chemicals. Agents thought to cause cancer are called carcinogens. The association of particular chemicals with cancer, particularly chemicals that are potent mutagens, led researchers early on to the suspicion that cancer might be caused, at least in part, by chemicals, the so-called chemical carcinogenesis theory.

Early Ideas
The chemical carcinogenesis theory was first advanced over 200 years ago in 1761 by Dr. John Hill, an English physician. Hill noted unusual tumors of the nose in heavy snuff users and suggested tobacco had produced these cancers. In 1775, a London surgeon, Sir Percivall Pott, made a similar observation, noting that men who had been chimney sweeps exhibited frequent cancer of the scrotum. He suggested that soot and tars might be responsible. These and many other observations led to the hypothesis that cancer results from the action of chemicals on the body.

Demonstrating That Chemicals Can Cause Cancer
It was over a century before this hypothesis was directly tested. In 1915, Japanese doctor Katsusaburo Yamagiwa applied extracts of coal tar to the skin of 137 rabbits every two or three days for three months. Then he waited to see what would happen. After a year, cancers appeared at the site of application in seven of the rabbits. Yamagiwa had induced cancer with the coal tar, the first direct demonstration of chemical carcinogenesis. In the decades that followed, this approach demonstrated that many chemicals were capable of causing cancer.

These were lab studies, and many did not accept that they applied to real people. Do tars in fact induce cancer in humans? In 1949, the American physician Ernst Winder and the British epidemiologist Richard Doll independently reported that lung cancer showed a strong link to the smoking of cigarettes, which introduces tars into the lungs. Winder interviewed 684 lung cancer patients and 600 normal controls, asking whether each had ever smoked. Cancer rates were 40 times higher in heavy smokers than in nonsmokers. From these studies, it seemed likely as long as 50 years ago that tars and other chemicals in cigarette smoke induce cancer in the lungs of persistent smokers. While this suggestion was (and is) resisted by the tobacco industry, the evidence that has accumulated since these pioneering studies makes a clear case, and there is no longer any real doubt. Chemicals in cigarette smoke cause cancer.

Carcinogens Are Common
In ongoing investigations over the last 50 years, many hundreds of synthetic chemicals have been shown capable of causing cancer in laboratory animals. Among them are trichloroethylene, asbestos, benzene, vinyl chloride, arsenic, arylamide, and a host of complex petroleum products with chemical structures resembling chicken wire.

In addition to identifying potentially dangerous substances, what have the studies of potential carcinogens told us about the nature of cancer? What do these cancer-causing chemicals have in common? They are all mutagens, each capable of inducing changes in DNA.
Chemicals that produce mutations in DNA, such as tars in cigarette smoke, are often potent carcinogens.