Chapter 16: How Genes Work

Genetic Information Flow

• Genetic information flows from DNA to RNA and then from RNA to proteins.

Central Dogma of Molecular Biology

• The chapter will cover the central dogma of molecular biology.

The Genetic Code

• The genetic code, with its 3-letter "words," will be analyzed.

Mutations

• The chapter will explain how mutations can modify genes and genomes.

Introduction to How Genes Work

• Early 1900s: Biologists knew about DNA, heredity, chromosomes, and genes.
• It took much longer to understand gene expression, which is the process of converting the information in DNA into functioning molecules within the cell.

What Do Genes Do?

• George Beadle and Edward Tatum proposed discovering what genes do by making them defective.
• First, damage a gene ("knock out" function).
• Second, observe the effect on the phenotype.
• Null or loss-of-function alleles are nonfunctioning alleles.
• Strategy: Irradiated bread mold (N. crassa) and looked for mutants that could not make certain compounds.
• They found mutants with defects in single genes for them to study.

Arginine Synthesis Pathway

• Srb and Horowitz tested the hypothesis that one gene codes for the production of one enzyme by studying a three-step metabolic pathway that produces arginine.
• They performed a genetic screen by growing mold cells on medium that lacked arginine (WT mold makes own Arg).
• Cells that died came from a colony that was missing an enzyme from the arginine pathway: “mutants of interest”.

Beadle and Tatum’s Screen Strategy

• Irradiated bread mold (N. crassa) to generate a bunch of random mutants.
• Grew these on media that has arginine so they can all grow.
• Tested individual mutant colonies to find ones that CANNOT grow without arginine.
• These are the ones they want to do experiments with next.

Experiment

• Isolate mutant N. crassa that cannot synthesize arginine.
• Grow each type of mutant on growth medium that is:
  • Not supplemented (no ornithine, citrulline, or arginine)
  • Supplemented with ornithine only (no citrulline or arginine)
  • Supplemented with citrulline only (no ornithine or arginine)
  • Supplemented with arginine only (no ornithine or citrulline)
• Test to see if each mutant also lacks one of the enzymes required for different steps in the pathway for synthesizing arginine.

Predictions

• Prediction: There will be three distinct types of mutants, corresponding to defects in enzyme 1, enzyme 2, and enzyme 3 in the pathway for synthesizing arginine. Each type of mutant will be able to grow on different combinations of the four types of media.
• Prediction of null hypothesis: There will not be a simple correspondence between a particular mutation and a particular enzyme.

Metabolic Pathway for Arginine Synthesis

• Precursor is converted to Ornithine by Enzyme 1.
• Ornithine is converted to Citrulline by Enzyme 2.
• Citrulline is converted to Arginine by Enzyme 3.

Results

• There are three distinct types of mutants: arg1, arg2, and arg3.
• arg1 cells lack enzyme 1.
• arg2 cells lack enzyme 2.
• arg3 cells lack enzyme 3.
• The one-gene, one-enzyme hypothesis is supported: Each gene contains the information to make a single enzyme.

Expanding the One Gene, One Enzyme Hypothesis

• Work by other scientists ultimately showed that genes contain the information for all proteins, not just enzymes.
• The hypothesis is now called the one-gene, one polypeptide hypothesis.
• Scientists began to form the central dogma of molecular biology.

The Genetic Code Hypothesis

• The question of how we get proteins from DNA was initially unknown.
• Francis Crick proposed that the sequence of bases in DNA acted as a code.
• DNA is an information storage molecule.
• Different combinations of bases specify the 20 amino acids.
• A particular stretch of DNA (a gene) specifies the amino acid sequence of one protein.
• The information in the sequence of DNA is not directly translated into the amino acid sequence of proteins.

RNA as Intermediary

• Jacob and Monod suggested that RNA links genes in the nucleus to protein synthesis in the cytoplasm.
• Messenger RNA (mRNA) was found to carry information from DNA to the site of protein synthesis.

RNA Polymerase

• RNA polymerase synthesizes RNA using a DNA strand as a template.
• It copies the code by matching complementary nucleotides.
• No primer is needed.

Central Dogma

• Summarizes the flow of information in cells:
  • Genes are stretches of DNA that ultimately code for proteins.
  • The DNA sequence codes for the RNA sequence.
  • The RNA sequence codes for the sequence of amino acids in a protein.
• DNA \rightarrow RNA \rightarrow proteins

Roles of Transcription and Translation

• Transcription: Process of using a DNA template to make a complementary RNA.
• Translation: Process of using the information in mRNA to synthesize proteins; interprets nucleotide “language” to amino acids.
• DNA is for information storage.
• mRNA is the information carrier.
• Proteins are the active cell machinery.
• Transcription: DNA \rightarrow mRNA
• Translation: mRNA \rightarrow Proteins

Linking Genotypes to Phenotypes

• Genotype is determined by DNA sequence.
• Phenotype is the product of the proteins produced.
• Alleles of the same gene differ in their DNA sequence.
• Proteins produced by different alleles of the same gene frequently differ in AA sequence.
• Coat color in mice is partly dependent on the melanocortin receptor; melanocortin = more dark pigment.

Exceptions to Central Dogma

• Many genes code for RNAs that do not function as mRNAs and are not translated into proteins.
• May form parts of the ribosome.
• May function as ribozymes to fold or process RNA.
• May regulate gene expression.
• Here the gene flow is DNA \rightarrow RNA (no proteins).

Reverse Transcription

• Sometimes information flows “backwards” from RNA to DNA.
• Some viruses contain reverse transcriptase.
• Synthesizes DNA from an RNA template.
• Here the gene flow is RNA \rightarrow DNA.

The Genetic Code

• Specifies how a sequence of nucleotides codes for a sequence of amino acids.
• George Gamow predicted that each “word” contains three bases.
• 3-base code is the least needed to specify the 20 amino acids.
• A three-base code is known as a triplet code.

Codons

• Codon: Group of 3 bases that specifies an amino acid.
• Francis Crick and Sydney Brenner confirmed that codons are triplets.
• Used mutagenic chemicals that caused an occasional addition or deletion of a base pair in DNA.
• The reading frame (sequence of codons) of a gene could be destroyed by adding or subtracting one or two bases.
• The reading frame was not destroyed by adding or subtracting multiples of three bases.

Genetic Code Example

• Consider: “The fat cat ate the rat”
• If you delete the f in “fat” you get: (deletion of one base) “The atc ata tet her at”
• BUT if you add the word “fat” before rat, you get: “The fat cat ate the fat rat” (addition of 3 bases)
• The words still make sense.
• Likewise, removing “fat” before cat: “The cat ate the rat”.

Codon-Amino Acid Mapping

• Marshall Nirenberg, Heinrich Matthaei, and Philip Leder worked out which codon coded for each amino acid.
• Made nucleic acid chains of specific codons and analyzed the protein that was produced.
• There is one start codon (AUG): codes for methionine and signals where protein synthesis starts.
• There are three stop codons (UGA, UAA, and UAG): signal the end of the protein-coding sequence.
• The other 60 codons code for amino acids.

Features of the Genetic Code

• Redundant: all but 2 AAs are encoded by >1 codon.
• Unambiguous: 1 codon never codes for >1 amino acid.
• Non-overlapping: Codons are read one at a time.
• Nearly universal: All codons specify the same amino acids in all organisms (with a few minor exceptions).
• Conservative: If several codons specify the same amino acid, the first two bases are usually identical.

Value of Knowing the Code

• Predict the codons and amino acid sequence encoded by a particular DNA sequence.
• Approximate the mRNA and DNA sequence that would code for a particular sequence of amino acids.

Types and Consequences of Mutation

• Mutation: Any permanent change in an organism’s DNA.
• Results in a change in its genotype and allows for the creation of new alleles.
• Point mutation: A change in a single base.
• Insertions and deletions (from one to many nucleotides).
• Chromosome-level mutations are larger in scale.

Point Mutations

• Silent: Change in nucleotide sequence that does not change the amino acid specified by a codon.
• Missense: Change in nucleotide sequence that changes the amino acid specified by codon.
• Nonsense: Change in nucleotide sequence that results in an early stop codon.
• Frameshift: Addition or deletion of a nucleotide.

Impacts on Fitness from Mutations

• Beneficial mutations increase fitness. Fitness = ability to survive and reproduce.
• Neutral mutations do not affect fitness.
• Deleterious mutations decrease fitness. Most point mutations are neutral or deleterious.
• Some mutations are not in coding regions but can still affect phenotype by affecting gene expression.

Chromosome Mutations

• Due to errors with moving chromosomes into daughter cells during meiosis or mitosis.
• May change chromosome number:
  • Polyploidy (>2 of each type)
  • Aneuploidy (loss or gain of a whole chromosome)
• May change chromosome structure:
  • Inversion: A segment of a chromosome breaks off, flips around, and rejoins.
  • Translocation: A section of a chromosome breaks off and becomes attached to another chromosome.
  • Deletion: A segment of a chromosome is lost.
  • Duplication: A segment of a chromosome is present in multiple copies.

Visualizing Chromosome Mutations

• Chromosome mutations can be beneficial, neutral, or deleterious.
• They can be visualized on a karyotype.