Supplemental Lecture (97/02/22 update) by Stephen T. Abedon (abedon.1@osu.edu)

  1. Chapter title: Transcription and Translation
    1. A list of vocabulary words is found toward the end of this document
    2. DNA, though an information storage molecule, is not inert. Indeed, one of its key roles in a cell, as the storer of genotype, is as the effector of phenotype. It does this through the creation of gene products: RNA and protein species which themselves are the direct producers of phenotype. The process by which DNA stored information is converted into RNA coded information and then into polypeptide (i.e., protein) begins with a process which is very similar to semiconservative DNA replication. This process is called transcription. In transcription an RNA polymer is synthesized from a DNA template. This RNA either is used as is, or delivers its information to an organelle called a ribosome. Ribosomes are protein-making factories, themselves composed of RNA and protein. Ribosomes take RNA coded information and "translate" that information into strings of amino acids called polypeptides. Thus we have the three main facets (central dogma) of molecular genetics: replication, transcription, and translation.
  2. Gene product
    1. A gene product is literally, the product of a gene though more precisely the final, in some way useful product of a gene.
    2. Proteins and some RNAs:
      1. Thus, proteins are gene products. However, the mRNAs from which proteins translated technically are not.
      2. A number of RNA species exist, however, which are not translated into polypeptides such as tRNAs and rRNAs, and these are considered gene products in the strictest sense.
    3. Expensive synthesis:
      1. The synthesis of gene products, especially polypeptides, is not cheap.
      2. Indeed, much of metabolism is devoted to the synthesis of gene products.
  3. Transcription [RNA polymerase]
    1. Transcription is the first step in the production of gene products.
    2. Gene templation of RNA product:
      1. Transcription is the generation of RNA from genes.
      2. Note that the produced strand of RNA is complementary to (i.e., is templated from) a gene (the latter, in most species, is composed of DNA). The term transcription is derived from the information-transferring (transcribing) nature of templated copying, nucleic acid to nucleic acid.
    3. Transcription is catalyzed by an enzyme called RNA polymerase.
    4. Analogous to DNA polymerization:
      1. It works on the basis of sequence complementarity just as does DNA replication.
      2. In the majority of cases, transcription occurs off of only one of the two DNA molecules rather than both as occurs in DNA replication.
      3. Note that as with DNA replication, RNA polymerase is capable of polymerizing RNA only in the 5' to 3' direction.
    5. mRNA, rRNA, and tRNA:
      1. The immediate products of transcription include various species of RNA including the main players in translation:
        1. mRNA
        2. rRNA
        3. tRNA.
  4. mRNA [a.k.a. messenger RNA]
    1. Polypeptide synthesizing information:
      1. The RNA molecule which holds the information necessary to effect the translation of a polypeptide.
      2. In other words, the mRNA information is translated into a specific sequence of amino acid.
    2. mRNAs are the means by which DNA encoded, polypeptide sequence information is delivered to ribosomes.
  5. rRNA [a.k.a. ribosomal RNA]
    1. Large RNAs that make up a substantial portion of the structure of ribosomes. That is, ribosomes are composed of various species of proteins plus various species of RNA, the latter called rRNA.
    2. rRNA is transcribed in much the same manner as is mRNA.
  6. tRNA [a.k.a. transfer RNA]
    1. The intermediary between mRNA coded information and the amino acid building blocks of proteins.
    2. Within ribosomes tRNAs simultaneously bind to mRNA (through a "codon") and amino acids, lining up the latter in the order they are to be incorporated into a growing polypeptide chain.
    3. cellss tend to contain a large number of tRNA types corresponding to the large variety of codons which exist, as well as the corresponding 20 naturally occurring amino acids which are incorporated directly into polypeptides.
    4. Note that the binding of an amino acid to a tRNA is called activation and that activation is effected through specific amino acid and tRNA recognizing proteins called activating enzymes.
  7. Translation
    1. Templated polypeptide synthesis:
      1. The synthesis of a polypeptide from an mRNA template.
      2. Translation occurs only with the intimate help of ribosomes.
      3. The term translation comes from the idea that the process involves the changing of sequence information from the language of nucleic acids (i.e., codon-based) to the language of polypeptides (i.e., amino acid based).
    2. Translation involves the coordinated action of mRNA, tRNA, soluble (i.e., cytoplasmic) pools of amino acids, and ribosomes.
    3. Example: vitally important to metabolism:
      1. Sequencing of the smallest known free-living organism, Microplasma genitalium (580,070 base pair genome) has been accomplished.
      2. "M. genitalism . . . has one of the smallest known geomes of any free-living organism. It is therefore reasonable to assume that its genome sequence reveals the near-minimal set of genes necessary for independent life" (pp. 445-446, Goffeau, 1995).
      3. Subsequent comparison to a larger bacterium (Haemophilus influenzae, 1,830,000 base pairs, also completely sequenced; compare with Escherichia coli, 4,720,000 base pairs, not completely sequenced as of late 1995) suggests that the degree to which a free-living organism can be reduced in size and complexity is limited by some minimal allowable biochemical complexity (apparently allowed by an increasingly complex growth requirements/environment) which may be reached by its translation apparatus before it is reached by other systems. That is, as complexity is reduced, the translation apparatus apparently becomes increasingly dominant, consisting of greater than 20% ( 100) of presumed genes coded by M. genitalium genome (p. 400, Fraser et al., 1995).
      4. "It is likely that the minimal translation machinery requires nearly 90 different proteins to proceed, whereas the complete DNA replication process requires only about 30 proteins." (p. 446, Goffeau, 1995)
      5. Interestingly, M. genitalium actually has more proteins inserted in its single membrane than it devotes to translation, perhaps reflecting one cost associated with a loss of anabolic capability, i.e., a requirement for a high capacity to acquire the many molecules consequently required for growth from the extracellular environment.
  8. Ribosome
    1. The site (organelle) in a cells where translation occurs.
    2. Overview:
      1. An overview of translation as it occurs in ribosomes is as follows:
        1. An mRNA binds to the ribosome at a specific end corresponding to the start of the information it encodes.
        2. This binding occurs though a specific ribosome binding sequence of nucleotides found on the mRNA.
        3. Aided by the ribosome, specific tRNAs bind at one end to the mRNA (i.e., to a codon) and at the other end to an amino acid.
        4. Again through the action of the ribosome, peptide bonds are formed between amino acids, at which point the tRNA's job is completed and it diffuses away.
        5. This process continues (i.e., the mRNA is ratcheted through the ribosome) until a stop codon is encountered on the mRNA.
        6. The encounter with the stop codon terminates protein synthesis.
        7. Polypeptide and mRNA are then released freeing the ribosome to attach to additional mRNAs.
    3. Note that the ribosomes of procaryotes and eucaryotes, though in many ways similar, differ sufficiently structurally that many antibiotics are capable of inhibiting translation through ribosome binding, of procaryotes but not eucaryotes, thus allowing the differential killing of procaryotic pathogens.
  9. Codon [triplet]
    1. Amino acid coding sequence of three nucleotides:
      1. A codon is a three nucleotide sequence which codes for the insertion of a unique amino acid during translation (e.g., UCU specifies the amino acid serine).
      2. Note that since there are only 4 types of nucleotides (abbreviated A, T, G, and C for DNA or A, U, G, and C for RNA), a three nucleotide sequence may be arranged into a total of 64 unique combinations (64 = 43). Thus, with a three nucleotide sequence all 20 (and them some) amino acids may be specified by a unique codon (a two nucleotide sequence could specify only 42 or 16 amino acids, too few, whereas a four nucleotide sequence could specify 44 or 256, wastefully too many).
    2. No punctuation/no overlap:
      1. Codons as arranged in DNA and mRNA neither overlap nor have punctuation.
      2. Instead, they are arranged end to end such that a given nucleotide sequence will represent a new codon every third base, while each nucleotide in a codon is associated with that codon only (rather than shared with adjacent codons).
    3. Almost universal code:
      1. The sequence of codons and the amino acids they bind appears to be, with only a few extremely minor exceptions, universal to all extant organisms.
      2. This universality of codon usage, combined with the apparent arbitrary correspondence of codon sequence to amino acid, is extremely strong evidence for the existence of a common ancestor to all extant organisms.
  10. Stop [nonsense] codon
    1. Translation termination signals:
      1. Not all codonss specify amino acids during translation. A small subgroup of codonss instead specify the termination of the translation of a given polypeptide.
      2. Such codons are called stop codons (because they stop a polypeptide's synthesis) or nonsense codons (which contrasts with the sense nature of codons which specify amino acids.
    2. Specifically these stop codons are UAA, UAG, and UGA, which are know as ochre, amber, and opal, respectively).
  11. Start codon
    1. AUG, which is the codon specifying the amino acid methionine, is found at the beginning of all reading frames specifying polypeptides.
    2. Its presence signals the translation starting point.
  12. Reading frame
    1. potentially polypeptide coding nucleotide arrays:
      1. Having codons arranged without punctuation or overlaps puts a premium on starting translation at the proper point.
      2. Note that in theory any given nucleotide sequence could represent three overlapping sequences of codons (six if you count the second strand of DNA).
      3. Each of these possible sequences of codons is called a reading frame (note that while codons don't overlap within a given reading frame, reading frames themselves can overlap---make sure this doesn't confuse you; see overlapping genes).
    2. A reading frame starts with a start codon and ends with one or more stop codons. In addition, there must be an even number of intervening codons (in other words, y = x/3 where y is an integer and x is the number of intervening nucleotidess). That is, all of the relevant codons much be in frame.
    3. Cannot necessarily produce proteins:
      1. Not all possible reading frames produce products (indeed, only rarely does more than one reading frame express a product).
      2. This is usually because, in addition to lacking a start codon, these reading frames lack proper nucleotide sequences necessary to initiate transcription (see biol1070.htmcontrol of gene expression) and often contain large numbers of stop codons which can terminate wasteful translation should transcription occur.
  13. Overlapping genes
    1. Same location, different reading frame:
      1. Though two or more reading frames may overlap, their products won't share amino acid sequences because various overlapping reading frames are not in frame (that is, each would be considered a unique gene loci, though clearly nucleotide sequences of such overlapping genes would be constrained evolutionarily by the sequences of the genes they overlap).
      2. That is, two genes may share a substantial number of nucleotides and sequence, but at the same time not share codon sequence.
    2. Overlapping genes are most often observed in organisms having genomes whose size is highly constrained such as within the genome of many viruses.
  14. Intron [exon]
    1. Reading frame intervening sequences:
      1. A further complication to most eucaryote genes, and a few procaryote genes, is the existence of introns.
      2. These are non-coding segments which break up reading frames.
    2. Post-transcriptional mRNA modification:
      1. With introns present a gene/mRNA cannot be translated.
      2. However, prior to translation introns are excised from mRNAs by various mechanisms. The coding region remaining is made up of joined together exons. Thus, a gene which contains introns actually exists as a series of exon sequences separated by intron sequences.
  15. Gene expression
    1. The synthesis of a final product (such as a protein or rRNA or tRNA) initially templated from a gene.
    2. For example, a polypeptide is a product of gene expression, a gene product.
    3. See control of gene expression.
  16. Central dogma [reverse transcription; RNA replication]
    1. Movement of nucleic acid-based information about cells is classically considered to be contrained to the following processes:
      1. DNA --(replication)-- DNA
      2. DNA --(transcription)-- mRNA
      3. mRNA --(translation) -- protein.
    2. In addition to the classically defined aspects defining the central dogma, there exist three additional analogous processes, each of which is common among various types of viruses:
      1. reverse transcription (RNA -- DNA)
      2. transcription directly from RNA (RNA -- mRNA)
      3. replication of RNA to produce progeny RNA genomes (RNA -- RNA)
  17. Links
    1. Transcription of DNA (Molecular Biology for Beginners)
    2. Translation of RNA (Molecular Biology for Beginners)
    3. Molecular Biology for Beginners
    4. Molecular Biology in the News (Molecular Biology for Beginners)
    5. ribosomes
  18. Vocabulary
    1. Central dogma
    2. Codon
    3. Exon
    4. Gene expression
    5. Gene product
    6. Intron
    7. mRNA
    8. Ribosome
    9. RNA polymerase
    10. Reading frame
    11. rRNA
    12. Stop codon
    13. Transcription
    14. Translation
    15. tRNA
  19. Practice questions
    1. What is the site in a eucaryotic cell in which transcription takes place? [PEEK]
      1. endoplasmic reticulum
      2. nucleus
      3. ribosome
      4. plasma membrane
      5. all of the above
      6. none of the above
    2. What enzyme catalyzes transcription? [PEEK]
    3. An example of a DNA sequence, discovered in the late 1970s, which disrupts a reading frame but does not prevent the correct expression of a gene is a(n): [PEEK]
      1. open reading frame
      2. exon
      3. intron
      4. codon
      5. all of the above
      6. none of the above
    4. At the beginning of a reading frame lies the codon AUG which codes for methionine. What is an other name for this codon? [PEEK]
    5. All of the following can directly participate in translation (not complete list, look for members that shouldn't be there): [PEEK]
      1. nucleus, tRNA, ribosomes
      2. tRNA, rRNA, mRNA, RNA polymerase
      3. activating enzymes, ribosomes, introns
      4. ribosomes, rough endoplasmic reticulum, mRNA
      5. all of the above
      6. none of the above
    6. State the central dogma of molecular genetics. Don't worry about exceptions. (<20 word answer) [PEEK]
    7. What is transcription? [PEEK]
    8. Give an example of a gene product? [PEEK]
    9. What is the enzyme which catalyzes transcription? [PEEK]
    10. State the central dogma of molecular genetics, indicating all three major processes, including what macromolecule one starts with and which one one ends up with (example of one of the minor, "exceptional" processes is reverse transcription which may be indicated as RNA --(reverse transcription)-- DNA). [PEEK]
    11. For there to be overlapping genes, minimally __________ reading frames must be employed (ignore any complication due to the presense of introns)? [PEEK]
      1. 0.
      2. 1.
      3. 2.
      4. 3.
      5. 4.
      6. 5.
      7. 6.
      8. more than 6.
    12. UAA, UAG, and UGA all have something in common, something which distinguishes these from, for example, AUG, as well as 60 other three unit arrays of the letters AGCU. Specifically, what is this distinguishing thing? [PEEK]
  20. Practice question answers
    1. Nucleus
    2. RNA polymerase
    3. Intron
    4. start codon
    5. iv, ribosomes, rough endoplasmic reticulum, mRNA. The nucleus, RNA polymerase, and introns do not participate directly in translation.
    6. DNA -- DNA -- RNA -- protein with arrows representing replication, transcription, and translation, respectively.
    7. the template-dependent polymerization of RNA, usually, though not always, from a double-stranded DNA template.
    8. tRNA, rRNA, protein, not mRNA.
    9. RNA polymerase.
    10. DNA --(replication)-- DNA; DNA --(transcription)-- mRNA; mRNA --(translation)-- protein (or polypeptide)
    11. iii, 2.
    12. These are the three stop codons.
  21. References
    1. Fraser et al. (1995). The minimal gene complement of Mycoplasma genitalium. Science 270:397-403.
    2. Goffeau, A. (1995). Life with 482 genes. Science 270:445-446.
    3. Raven, P.H., Johnson, G.B. (1995). Biology (updated version). Third Edition. Wm. C. Brown publishers, Dubuque, Iowa. pp. 300-312.