Chapter 1: How Genes Work: The New Genetics – National …

People have known for many years that living things inherit traits from their parents. That common-sense observation led to agriculture, the purposeful breeding and cultivation of animals and plants for desirable characteristics. Firming up the details took quite some time, though. Researchers did not understand exactly how traits were passed to the next generation until the middle of the 20th century.

Now it is clear that genes are what carry our traits through generations and that genes are made of deoxyribonucleic acid (DNA). But genes themselves don't do the actual work. Rather, they serve as instruction books for making functional molecules such as ribonucleic acid (RNA) and proteins, which perform the chemical reactions in our bodies.

Proteins do many other things, too. They provide the body's main building materials, forming the cell's architecture and structural components. But one thing proteins can't do is make copies of themselves. When a cell needs more proteins, it uses the manufacturing instructions coded in DNA.

The DNA code of a genethe sequence of its individual DNA building blocks, labeled A (adenine), T (thymine), C (cytosine) and G (guanine) and collectively called nucleotides spells out the exact order of a protein's building blocks, amino acids.

Occasionally, there is a kind of typographical error in a gene's DNA sequence. This mistake which can be a change, gap or duplicationis called a mutation.

A mutation can cause a gene to encode a protein that works incorrectly or that doesn't work at all. Sometimes, the error means that no protein is made.

But not all DNA changes are harmful. Some mutations have no effect, and others produce new versions of proteins that may give a survival advantage to the organisms that have them. Over time, mutations supply the raw material from which new life forms evolve (see Chapter 3, "Life's Genetic Tree").

The monk Gregor Mendel first described how traits are inherited from one generation to the next.

In 1900, three European scientists independently discovered an obscure research paper that had been published nearly 35 years before. Written by Gregor Mendel, an Austrian monk who was also a scientist, the report described a series of breeding experiments performed with pea plants growing in his abbey garden.

Mendel had studied how pea plants inherited the two variant forms of easy-to-see traits. These included flower color (white or purple) and the texture of the peas (smooth or wrinkled). Mendel counted many generations of pea plant offspring and learned that these characteristics were passed on to the next generation in orderly, predictable ratios.

When he cross-bred purple-flowered pea plants with white-flowered ones, the next generation had only purple flowers. But directions for making white flowers were hidden somewhere in the peas of that generation, because when those purple-flowered plants were bred to each other, some of their offspring had white flowers. What's more, the second-generation plants displayed the colors in a predictable pattern. On average, 75 percent of the second-generation plants had purple flowers and 25 percent of the plants had white flowers. Those same ratios persisted, and were reproduced when the experiment was repeated many times over.

Trying to solve the mystery of the missing color blooms, Mendel imagined that the reproductive cells of his pea plants might contain discrete "factors," each of which specified a particular trait, such as white flowers. Mendel reasoned that the factors, whatever they were, must be physical material because they passed from parent to offspring in a mathematically orderly way. It wasn't until many years later, when the other scientists unearthed Mendel's report, that the factors were named genes.

Early geneticists quickly discovered that Mendel's mathematical rules of inheritance applied not just to peas, but also to all plants, animals and people. The discovery of a quantitative rule for inheritance was momentous. It revealed that a common, general principle governed the growth and development of all life on Earth.

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Up until the 1950s, scientists knew a good deal about heredity, but they didn't have a clue what DNA looked like. In order to learn more about DNA and its structure, some scientists experimented with using X rays as a form of molecular photography.

In 1953, Watson and Crick created their historic model of the shape of DNA: the double helix. COLD SPRING HARBOR LABORATORY ARCHIVES

Rosalind Franklin, a physical chemist working with Maurice Wilkins at King's College in London, was among the first to use this method to analyze genetic material. Her experiments produced what were referred to at the time as "the most beautiful X-ray photographs of any substance ever taken."

Other scientists, including zoologist James Watson and physicist Francis Crick, both working at Cambridge University in the United Kingdom, were trying to determine the shape of DNA too. Ultimately, this line of research revealed one of the most profound scientific discoveries of the 20th century: that DNA exists as a double helix.

The 1962 Nobel Prize in physiology or medicine was awarded to Watson, Crick and Wilkins for this work. Although Franklin did not earn a share of the prize due to her untimely death at age 38, she is widely recognized as having played a significant role in the discovery.

Rosalind Franklin's original X-ray diffraction photo revealed the physical structure of DNA. OREGON STATE UNIVERSITY LIBRARIES SPECIAL COLLECTIONS

The spiral staircase-shaped double helix has attained global status as the symbol for DNA. But what is so beautiful about the discovery of the twisting ladder structure isn't just its good looks. Rather, the structure of DNA taught researchers a fundamental lesson about genetics. It taught them that the two connected strandswinding together like parallel handrailswere complementary to each other, and this unlocked the secret of how genetic information is stored, transferred and copied.

In genetics, complementary means that if you know the sequence of nucleotide building blocks on one strand, you know the sequence of nucleotide building blocks on the other strand: A always matches up with T and C always links to G (see drawing).

Long strings of nucleotides form genes, and groups of genes are packaged tightly into structures called chromosomes. Every cell in your body except for eggs, sperm and red blood cells contains a full set of chromosomes in its nucleus.

If the chromosomes in one of your cells were uncoiled and placed end to end, the DNA would be about 6 feet long. If all the DNA in your body were connected in this way, it would stretch approximately 67 billion miles! That's nearly 150,000 round trips to the Moon.

The long, stringy DNA that makes up genes is spooled within chromosomes inside the nucleus of a cell. (Note that a gene would actually be a much longer stretch of DNA than what is shown here.)

DNA consists of two long, twisted chains made up of nucleotides. Each nucleotide contains one base, one phosphate molecule and the sugar molecule deoxyribose. The bases in DNA nucleotides are adenine, thymine, cytosine and guanine.

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Humans have 23 pairs of chromosomes. Male DNA (pictured here) contains an X and a Y chromosome, whereas female DNA contains two X chromosomes. CYTOGENETICS LABORATORY, BRIGHAM AND WOMEN'S HOSPITAL

It's astounding to think that your body consists of trillions of cells. But what's most amazing is that it all starts with one cell. How does this massive expansion take place?

As an embryo progresses through development, its cells must reproduce. But before a cell divides into two new, nearly identical cells, it must copy its DNA so there will be a complete set of genes to pass on to each of the new cells.

To make a copy of itself, the twisted, compacted double helix of DNA has to unwind and separate its two strands. Each strand becomes a pattern, or template, for making a new strand, so the two new DNA molecules have one new strand and one old strand.

The copy is courtesy of a cellular protein machine called DNA polymerase, which reads the template DNA strand and stitches together the complementary new strand. The process, called replication, is astonishingly fast and accurate, although occasional mistakes, such as deletions or duplications, occur. Fortunately, a cellular spell-checker catches and corrects nearly all of these errors.

When DNA polymerase makes an error while copying a gene's DNA sequence, the mistake is called a mutation. In this example, the nucleotide G has been changed to an A.

During DNA replication, each strand of the original molecule acts as a template for the synthesis of a new, complementary DNA strand.

Mistakes that are not corrected can lead to diseases such as cancer and certain genetic disorders. Some of these include Fanconi anemia, early aging diseases and other conditions in which people are extremely sensitive to sunlight and some chemicals.

DNA copying is not the only time when DNA damage can happen. Prolonged, unprotected sun exposure can cause DNA changes that lead to skin cancer, and toxins in cigarette smoke can cause lung cancer.

It may seem ironic, then, that many drugs used to treat cancer work by attacking DNA. That's because these chemotherapy drugs disrupt the DNA copying process, which goes on much faster in rapidly dividing cancer cells than in other cells of the body. The trouble is that most of these drugs do affect normal cells that grow and divide frequently, such as cells of the immune system and hair cells.

Understanding DNA replication better could be a key to limiting a drug's action to cancer cells only.

After copying its DNA, a cell's next challenge is getting just the right amount of genetic material into each of its two offspring.

Most of your cells are called diploid ("di" means two, and "ploid" refers to sets of chromosomes) because they have two sets of chromosomes (23 pairs). Eggs and sperm are different; these are known as haploid cells. Each haploid cell has only one set of 23 chromosomes so that at fertilization the math will work out: A haploid egg cell will combine with a haploid sperm cell to form a diploid cell with the right number of chromosomes: 46.

Chromosomes are numbered 1 to 22, according to size, with 1 being the largest chromosome. The 23rd pair, known as the sex chromosomes, are called X and Y. In humans, abnormalities of chromosome number usually occur during meiosis, the time when a cell reduces its chromosomes from diploid to haploid in creating eggs or sperm.

What happens if an egg or a sperm cell gets the wrong number of chromosomes, and how often does this happen?

Trisomy, the hallmark of Down syndrome, results when a baby is born with three copies of chromosome 21 instead of the usual two.

Molecular biologist Angelika Amon of the Massachusetts Institute of Technology in Cambridge says that mistakes in dividing DNA between daughter cells during meiosis are the leading cause of human birth defects and miscarriages. Current estimates are that 10 percent of all embryos have an incorrect chromosome number. Most of these don't go to full term and are miscarried.

In women, the likelihood that chromosomes won't be apportioned properly increases with age. One of every 18 babies born to women over 45 has three copies of chromosome 13, 18 or 21 instead of the normal two, and this improper balancing can cause trouble. For example, three copies of chromosome 21 lead to Down syndrome.

To make her work easier, Amonlike many other basic scientistsstudies yeast cells, which separate their chromosomes almost exactly the same way human cells do, except that yeast do it much faster. A yeast cell copies its DNA and produces daughter cells in about 1 1/2 hours, compared to a whole day for human cells.

The yeast cells she uses are the same kind bakeries use to make bread and breweries use to make beer!

Amon has made major progress in understanding the details of meiosis. Her research shows how, in healthy cells, gluelike protein complexes called cohesins release pairs of chromosomes at exactly the right time. This allows the chromosomes to separate properly.

These findings have important implications for understanding and treating infertility, birth defects and cancer.

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So, we've described DNAits basic properties and how our bodies make more of it. But how does DNA serve as the language of life? How do you get a protein from a gene?

There are two major steps in making a protein. The first is transcription, where the information coded in DNA is copied into RNA. The RNA nucleotides are complementary to those on the DNA: a C on the RNA strand matches a G on the DNA strand.

1. RNA polymerase transcribes DNA to make messenger RNA (mRNA). 2. The mRNA sequence (dark red strand) is complementary to the DNA sequence (blue strand). 3. On ribosomes, transfer RNA (tRNA) helps convert mRNA into protein. 4. Amino acids link up to make a protein.

The only difference is that RNA pairs a nucleotide called uracil (U), instead of a T, with an A on the DNA.

A protein machine called RNA polymerase reads the DNA and makes the RNA copy. This copy is called messenger RNA, or mRNA, because it delivers the gene's message to the protein-producing machinery.

At this point you may be wondering why all of the cells in the human body aren't exactly alike, since they all contain the same DNA.What makes a liver cell different from a brain cell? How do the cells in the heart make the organ contract, but those in skin allow us to sweat?

Cells can look and act differently, and do entirely different jobs, because each cell "turns on," or expresses, only the genes appropriate for what it needs to do.

RNA polymerase (green) and one end of a DNA strand (blue) are attached to clear beads pinned down in two optical traps. As RNA polymerase moves along the DNA, it creates an RNA copy of a gene, shown here as a pink strand. STEVEN BLOCK

That's because RNA polymerase does not work alone, but rather functions with the aid of many helper proteins. While the core part of RNA polymerase is the same in all cells, the helpers vary in different cell types throughout the body.

You'd think that for a process so essential to life, researchers would know a lot about how transcription works. While it's true that the basics are clearbiologists have been studying gene transcribing by RNA polymerases since these proteins were first discovered in 1960 some of the details are actually still murky.

The biggest obstacle to learning more has been a lack of tools. Until recently, researchers were unable to get a picture at the atomic level of the giant RNA polymerase protein assemblies inside cells to understand how the many pieces of this amazing, living machine do what they do, and do it so well.

But our understanding is improving fast, thanks to spectacular technological advances. We have new X-ray pictures that are far more sophisticated than those that revealed the structure of DNA. Roger Kornberg of Stanford University in California used such methods to determine the structure of RNA polymerase. This work earned him the 2006 Nobel Prize in chemistry. In addition, very powerful microscopes and other tools that allow us to watch one molecule at a time provide a new look at RNA polymerase while it's at work reading DNA and producing RNA.

For example, Steven Block, also of Stanford, has used a physics technique called optical trapping to track RNA polymerase as it inches along DNA. Block and his team performed this work by designing a specialized microscope sensitive enough to watch the real-time motion of a single polymerase traveling down a gene on one chromosome.

The researchers discovered that molecules of RNA polymerase behave like battery-powered spiders as they crawl along the DNA ladder, adding nucleotides one at a time to the growing RNA strand. The enzyme works much like a motor, Block believes, powered by energy released during the chemical synthesis of RNA.

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Genes are often interrupted by stretches of DNA (introns, blue) that do not contain instructions for making a protein. The DNA segments that do contain protein-making instructions are known as exons (green).

Several types of RNA play key roles in making a protein. The gene transcript (the mRNA) transfers information from DNA in the nucleus to the ribosomes that make protein. Ribosomal RNA forms about 60 percent of the ribosomes. Lastly, transfer RNA carries amino acids to the ribosomes. As you can see, all three types of cellular RNAs come together to produce new proteins.

But the journey from gene to protein isn't quite as simple as we've just made it out to be. After transcription, several things need to happen to mRNA before a protein can be made. For example, the genetic material of humans and other eukaryotes (organisms that have a nucleus) includes a lot of DNA that doesn't encode proteins. Some of this DNA is stuck right in the middle of genes.

To distinguish the two types of DNA, scientists call the coding sequences of genes exons and the pieces in between introns (for intervening sequences).

If RNA polymerase were to transcribe DNA from the start of an intron-containing gene to the end, the RNA would be complementary to the introns as well as the exons.

To get an mRNA molecule that yields a working protein, the cell needs to trim out the intron sections and then stitch only the exon pieces together (see drawing). This process is called RNA splicing.

Arranging exons in different patterns, called alternative splicing, enables cells to make different proteins from a single gene.

Splicing has to be extremely accurate. An error in the splicing process, even one that results in the deletion of just one nucleotide in an exon or the addition of just one nucleotide in an intron, will throw the whole sequence out of alignment. The result is usually an abnormal proteinor no protein at all. One form of Alzheimer's disease, for example, is caused by this kind of splicing error.

Molecular biologist Christine Guthrie of the University of California, San Francisco, wants to understand more fully the mechanism for removing intron RNA and find out how it stays so accurate.

She uses yeast cells for these experiments. Just like human DNA, yeast DNA has introns, but they are fewer and simpler in structure and are therefore easier to study. Guthrie can identify which genes are required for splicing by finding abnormal yeast cells that mangle splicing.

So why do introns exist, if they're just going to be chopped out? Without introns, cells wouldn't need to go through the splicing process and keep monitoring it to be sure it's working right.

As it turns out, splicing also makes it possible for cells to create more proteins.

Think about all the exons in a gene. If a cell stitches together exons 1, 2 and 4, leaving out exon 3, the mRNA will specify the production of a particular protein. But instead, if the cell stitches together exons 1, 2 and 3, this time leaving out exon 4, then the mRNA will be translated into a different protein (see drawing).

By cutting and pasting the exons in different patterns, which scientists call alternative splicing, a cell can create different proteins from a single gene. Alternative splicing is one of the reasons why human cells, which have about 20,000 genes, can make hundreds of thousands of different proteins.

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Until recently, researchers looked at genes, and the proteins they encode, one at a time. Now, they can look at how large numbers of genes and proteins act, as well as how they interact. This gives them a much better picture of what goes on in a living organism.

Already, scientists can identify all of the genes that are transcribed in a cellor in an organ, like the heart. And although researchers can't tell you, right now, what's going on in every cell of your body while you read a book or walk down the street, they can do this sort of "whole-body" scan for simpler, single-celled organisms like yeast.

Using a technique called genome-wide location analysis, Richard Young of the Massachusetts Institute of Technology unraveled a "regulatory code" of living yeast cells, which have more than 6,000 genes in their genome. Young's technique enabled him to determine the exact places where RNA polymerase's helper proteins sit on DNA and tell RNA polymerase to begin transcribing a gene.

Since he did the experiment with the yeast exposed to a variety of different conditions,Young was able to figure out how transcription patterns differ when the yeast cell is under stress (say, in a dry environment) or thriving in a sugary-rich nutrient solution. Done one gene at a time, using methods considered state-of-the-art just a few years ago, this kind of analysis would have taken hundreds of years.

After demonstrating that his technique worked in yeast, Young then took his research a step forward. He used a variation of the yeast method to scan the entire human genome in small samples of cells taken from the pancreases and livers of people with type 2 diabetes. He used the results to identify genes that aren't transcribed correctly in people with the disease.

This information provides researchers with an important tool for understanding how diabetes and other diseases are influenced by defective genes. By building models to predict how genes respond in diverse situations, researchers may be able to learn how to stop or jump-start genes on demand, change the course of a disease or prevent it from ever happening.

While most genetic research uses lab organisms, test tubes and petri dishes, the results have real consequences for people. Your first encounter with genetic analysis probably happened shortly after you were born, when a doctor or nurse took a drop of blood from the heel of your tiny foot.

Lab tests performed with that single drop of blood can diagnose certain rare genetic disorders as well as metabolic problems like phenylketonuria (PKU).

Screening newborns in this way began in the 1960s in Massachusetts with testing for PKU, a disease affecting 1 in 14,000 people. PKU is caused by an enzyme that doesn't work properly due to a genetic mutation. Those born with this disorder cannot metabolize the amino acid phenylalanine, which is present in many foods. Left untreated, PKU can lead to mental retardation and neurological damage, but a special diet can prevent these outcomes. Testing for this condition has made a huge difference in many lives.

Newborn screening is governed by individual states. This means that the state in which a baby is born determines the genetic conditions for which he or she will be screened. Currently, states test for between 28 and 54 conditions. All states test for PKU.

Although expanded screening for genetic diseases in newborns is advocated by some, others question the value of screening for conditions that are currently untreatable. Another issue is that some children with mild versions of certain genetic diseases may be treated needlessly.

In 2006, the Advisory Committee on Heritable Disorders in Newborns and Children, which assists the Secretary of the U.S. Department of Health and Human Services, recommended a standard, national set of newborn tests for 29 conditions, ranging from relatively common hearing problems to very rare metabolic diseases.

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A ribosome consists of large and small protein subunits with transfer RNAs nestled in the middle. RIBOSOME STRUCTURE COURTESY OF JAMIE CATE, MARAT YUSUPOV, GULNARA YUSUPOVA, THOMAS EARNEST AND HARRY NOLLER. GRAPHIC COURTESY OF ALBION BAUCOM, UNIVERSITY OF CALIFORNIA, SANTA CRUZ.

After a gene has been read by RNA polymerase and the RNA is spliced, what happens next in the journey from gene to protein? The next step is reading the RNA information and fitting the building blocks of a protein together. This is called translation, and its principal actors are the ribosome and amino acids.

Ribosomes are among the biggest and most intricate structures in the cell. The ribosomes of bacteria contain not only huge amounts of RNA, but also more than 50 different proteins. Human ribosomes have even more RNA and between 70 and 80 different proteins!

Harry Noller of the University of California, Santa Cruz, has found that a ribosome performs several key jobs when it translates the genetic code of mRNA. As the messenger RNA threads through the ribosome protein machine, the ribosome reads the mRNA sequence and helps recognize and recruit the correct amino acid-carrying transfer RNA to match the mRNA code. The ribosome also links each additional amino acid into a growing protein chain (see drawing).

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