From Gene to Protein

7. Transcription

The discovery that genes are made of DNA left unanswered the question of how the information in DNA is used. How does a string of nucleotides in a spiral molecule determine if you have red hair? We now know that the information in DNA is arrayed in little blocks, like entries in a dictionary, each block a gene specifying a protein. These proteins determine what a particular cell will be like.

Just as an architect protects building plans from loss or damage by keeping them safe in a central place and issuing only blueprint copies to on-site workers, so your cells protect their DNA instructions by keeping them safe within a central DNA storage area, the nucleus. The DNA never leaves the nucleus. Instead, "blueprint" copies of particular genes within the DNA instructions are sent out into the cell to direct the assembly of proteins. These working copies of genes are made of ribonucleic acid (RNA) rather than DNA. Recall that RNA is the same as DNA except that the sugars in RNA have an extra oxygen atom and T is replaced by a similar pyrimidine base called U, uracil. The path of infonnation is thus: DNA -> RNA --> protein

This information path is often called the central dogma, because it describes the key organization used by your cells to express their genes. A cell uses three kinds of RNA in the synthesis of proteins: messenger RNA (mRNA), ribosomal RNA (rRNA), and transfer RNA (tRNA). The use of information in DNA to direct the production of particular proteins is called gene expression. Gene expression occurs in two stages: in the first stage, called transcription, an mRNA molecule is synthesized from a gene within the DNA; in the second stage, called translation, this mRNA is used to direct the production of a protein.

The Transcription Process

The RNA copy of a gene used in the cell to produce a protein is called messenger RNA (mRNA)-it is the messenger that conveys the information from the nucleus to the cytoplasm. The copying process that makes the mRNA is called transcriptionjust as monks in monasteries used to make copies of manuscripts by faithfully transcribing each letter, so enzymes within the nuclei of your cells make mRNA copies of your genes by faithfully complementing each nucleotide.

In your cells, the transcriber is a large and very sophisticated protein called RNA polymerase. It binds to one strand of a DNA double helix at a particular site called a promoter and then moves down the DNA strand like a train on a track. As it goes along the DNA "track," the polymerase pairs each nucleotide with its complernentary RNA version (A with U, G with C), building an mRNA chain behind it as it moves down the DNA strand.
Key concept: Transcription is the production of an mRNA copy of a gene by the enzyme RNA polymerase.

Figure. The central dogma of gene expression. Through the production of mRNA(transcription) and the sysnthesis of proteins(translation), the information contained in DNA is expressed.

8. The Genetic Code

The essence of Mendelian genetics is that information determining hereditary traits, traits passed from parent to child, is encoded information. The information is written within the chromosomes in blocks called genes. Genes affect Mendelian traits by directing the production of particular proteins. The essence of gene expression, of using your genes, is reading the information encoded within DNA and using that information to direct the production of the correct protein.

To correctly read a gene, a cell must translate the information encoded in DNA into the language of proteinsthat is, it must convert the order of the ,gene's nucleotides into the order of amino acids in a protein. The rules that govern this translation are called the genetic code. They are very simple:

1. Each gene is read from a fixed starting position, a nucleotide sequence at its beginning called a promoter site where the RNA polymerase first binds to the DNA.
2. RNA polymerase moves down the DNA in steps that are three nucleotides long.
3. Each three-nucleotide block in the gene corresponds to a particular amino acid.

Special three-nucleotide sequences located at the end of genes say "stop." A three-nucleotide sequence on mRNA that corresponds to an amino acid is called a codon. Biologists worked out which codons correspond to which amino acids by trial-and-error experiments carried out in test tubes. In these experiments, investigators used artificial mRNAs to direct the synthesis of proteins in the tube and then looked to see the sequence of amino acids in the proteins. An mRNA that was a string of UUUUUU . . . , for example, produced a protein that was a string of phenylalanine (PHE) amino acids, telling investigators that the codon UUU corresponded to the amino acid PHE. The entire genetic code dictionary is presented in figure 8.13. Because at each position of a three-letter codon any of the four different nucleotides (A, U, G, C) may be used, there are 64 different possible three-letter codons (4 x 4 x 4 = 64) in the genetic code.

The genetic code is universal, the same in practically all organisms. GUC codes for valine in bacteria, in fruit flies, in eagles, and in your own cells. The only exceptions biologists have ever found to this rule s in the way in which cell or,ganelles that contain DNA (rnitochondria and chloroplasts) and a few microscopic protists read the "stop" codons. In every other instance, the same genetic code is employed by all living things.
Key concepts: The genetic code dictates how a particular nucleoticle sequence specifies a particular amino acid sequence. The nucleotide sequence is read in three-base increments called codons. The genetic code determines which amino acid is associated with which codon.

9. Translation

The final result of the transcription process is the production of an mRNA copy of a gene. Like a photocopy, the mRNA can be used without damage or wear and tear on the original. After transcription of a gene is finished, the mRNA passes out of the nucleus into the cytoplasm through pores in the nuclear membrane. There, translation of the genetic message occurs. In translation, organelles called ribosomes use the mRNA produced by transcription to direct the synthesis of a protein.

The Protein-Making Factory

Ribosomes are the factories of the cell. Each is very complex, containing over 50 different proteins and several segments of ribosornal RNA (rRNA). Ribosomes use mRNA, the "blueprint" copies of nuclear genes, to direct the assembly of a protein.

Ribosomes are composed of two pieces, or subunits, one nested into the other like a fist in the palm of your hand. The "fist" is the smaller of the two subunits. Its rRNA has a short nucleotide sequence exposed on the surface of the subunit. This exposed sequence is identical to a sequence called the leader region that occurs at the beginning of all genes. Because of this, an mRNA molecule binds to the exposed rRNA of the small subunit like a fly sticking to flypaper.

The Key Role of tRNA

Directly adjacent to the exposed rRNA sequence are three small pockets or dents, called the A, P, and E sites, in the surface of the ribosome that have just the right shape to bind yet a third kind of RNA molecule, transfer RNA (tRNA). It is tRNA molecules that bring amino acids to the ribosome to use in making proteins. tRNA molecules are chains about 80 nucleotides long, folded into a compact shape, with a three-nucleotide sequence jutting out at one end and an amino acid attachment site to the other.

The three-nucleotide sequence is very important-called the anticodon, it is the complementary sequence to I of the 64 codons of the genetic code! Special enzymes match amino acids with their proper tRNAs, with the anticodon determining which amino acid will attach to a particular tRNA.

Because the first dent in the ribosome (the A site) is directly adjacent to where the mRNA binds to the rRNA, three nucleotides of the mRNA are positioned directly facing the jutting anticodon of the tRNA. Like the address on a letter, the anticodon ensures that an amino acid is delivered to its correct "address" on the mRNA where the ribosome is assembling the protein.

A ribosome is composed of two subunits. The smaller subunit fits into a depression on the surface of the larger one. The A, P, and E sites on the ribosome play key roles in protein synthesis.

How translation works.

The mRNA strand acts as a template for tRNA molecules. The appropriate tRNA is selected and positioned by the ribosome, which moves along the mRNA in three-nucleoticle steps. tRNAs bring amino acids into the ribosome at the A site. A peptide bond is formed between the incoming am i no acid and the growing polypepticle chain at the P site, and the empty tRNAs leave the ribosome at the E site. to Making the Protein

Making the Protein

Once an mRNA molecule has bound to the small ribosomal subunit, the other larger ribosomal subunit binds as well, forming a complete ribosome. The ribosome then begins the process of translation. The mRNA begins to thread through the ribosome like a string passing through the hole in a donut. The mRNA passes through in short spurts, three nucleotides at a time, and at each burst of movement a new threenucleotide codon is positioned opposite the A site in the ribosome, where tRNA molecules first bind.

As each new tRNA brings in an amino acid to each new codon presented at the A site, the old tRNA paired with the previous codon is passed over to the P site and eventually to the E site, as the amino acid it carried is attached to the end of a growing amino acid chain. So as the ribosome proceeds down the mRNA, one tRNA after another is selected to match the sequence of mRNA codons, until the end of the mRNA sequence is reached. At this point, a codon is encountered for which there is no anticodon on any tRNA molecule. With nothing to fit into the tRNA site, the ribosome complex falls apart, and the newly made protein is released into the cell.
Key concepts: Translation is the reading of mRNA by a ribosome, the sequence of mRNA codons dictating the assembly of a corresponding sequence of amino acids in a growing protein chain.

10. Architecture of the Gene

Introns

While it is tempting to think of a gene as simply the nucleotide version of a protein-an uninterrupted stretch of nucleotides that is read three at a time to make a chain of amino acids-this actually occurs only in bacteria. In all organisms that evolved later (that is, the eukaryotes), genes are fragmented. In these more complex genes, the DNA nucleotide sequences encoding the amino acid sequence of the protein (called exons) are interrupted frequently by extraneous "extra stuff" called introns. In most human genes, the intron material far outweighs the protein-encoding exons. Usually less than 10% of a human gene is exons, and all the rest are noncoding introns. Like cars on a rural highway, exons are scattered here and there within genes, in the correct order but not near one another.

How does the cell manage to make a protein from such a mess? After the mRNA has been copied from the DNA, special enzymes attack it and chop out all the introns! The exons are then stitched together to make the mRNA that would have been transcribed if the introns had never existed, and it is this "processed" mRNA that the ribosomes translate into protein. In addition, enzymes add a structure called a 5' cap that protects the 5' end of the RNA template from being degraded during its long Journey through the cytoplasm. Another enzyme adds about 250 A nucleotides to the 3' end of the transcript. Called a 3' poly-A tail, this long string of A nucleotides also protects the transcript frorn degradation and appears to make the transcript a better template for protein synthesis.

Why this crazy organization? It appears that each exon of a gene encodes a different functional part of the protein. One exon may influence which molecules an enzyme is able to recognize and bind, while a different exon may determine what chemical reaction then takes place. Yet another exon may specify which signal molecules the protein will respond to. Biologists believe that evolution has favored this intron/ exon mode of gene organization because it has permitted cells to shuffle exons between genes. This sort of shuffling is a very powerful way to create "new" genes-in one step producing a new kind of functional protein, instead of having to invent each new protein from scratch. Like building models with Tinkertoys or Legos, many different enzymes can be created. Imagine the number of outfits two friends can create by exchanging sweaters, pants, belts. shoes, and hats. The many millions of proteins in human cells appear to have arisen froin only a few thousand exons!

Gene Families

Introns are not the only surprise awaiting you in studying eukaryotic genes. Everything we have said in this chapter assumes that chromosomes each carry one copy of each kind of gene-one to make the enzyme that breaks down lactose, for example, and one to encode the protein hemoglobin, which carries oxygen in your blood from lungs to tissues. In fact, this is far from true. Most genes in your cells exist in multiple copies, clusters of almost identical sequences called multigene families. Multigene families may contain as few as three or as many as several hundred versions of a gene.

Transposons: Jumping Genes

Other genes are very unusual in that they are repeated hundreds of thousands of times, scattered randomly about on the chromosomes. These genes, called transposons, have the remarkable ability to move about from one chromosomal location to another. Once every few thousand cell divisions, a transposon simply picks up and moves elsewhere, jumpinz-, at random to a new location on the chromosome. Transposons appear to be inolecular parasites. They are actively transcribed by RNA polymerase but play no functional role in the life of the cell that bears them.
The coding portions of most eukaryotic genes are embedded as exons within long sequences of noncoding introns. Many eukaryotic genes exist in multiple copies, some of which appear to have moved from one chromosomal location to another.