Microbial Genetics - Chapter 7 Key Terms

Clostridium difficile overview

  • Antibiotics are used (context for CDI).

  • Spores are present in the colon.

  • Normal microbiota protective measure helps prevent infection.

  • Toxins produced by C. difficile kill tissue inside the colon.

  • Tissue injury plus white blood cells (WBC) lead to lesion formation.

Terminology

  • Genetics: the study of what genes are, how they carry information, how information is expressed, and how genes are replicated.

  • Gene: a segment of DNA that encodes a functional product, usually a protein.

  • Chromosome: a structure containing DNA that physically carries hereditary information; chromosomes contain the genes.

  • Genome: all the genetic information in a cell.

Physical structure of DNA and basic genetic elements

  • Four bases in DNA:

    • Purines: Adenine (A), Guanine (G)

    • Pyrimidines: Cytosine (C), Thymine (T)

  • In RNA, Uracil (U) substitutes for T.

  • Base pairing:

    • G–C

    • A–T (DNA) or A–U (RNA)

  • DNA is a double helix with complementary base pairing.

  • Plasmids:

    • Small DNA molecules that replicate independently.

    • Not essential for normal metabolism, growth, or reproduction.

    • Can confer survival advantages.

    • Types include:

    • Fertility factors

    • Resistance factors

    • Bacteriocin factors

    • Virulence plasmids

DNA replication: general concepts

  • Replication is semiconservative: new DNA contains one original strand and one daughter strand.

  • Initiation occurs at the origin in bacteria.

  • DNA polymerase replicates DNA 5' → 3'.

  • Strands are antiparallel; leading strand is synthesized continuously, lagging strand discontinuously via Okazaki fragments.

  • Figure references illustrate semiconservative replication and initial replication processes.

Characteristics of bacterial DNA replication

  • Topoisomerases remove supercoils in the DNA molecule.

  • DNA is methylated, involved in:

    • Control of genetic expression

    • Initiation of DNA replication

    • Protection against viral infection

    • Repair of DNA

  • End terminology:

    • 3' end is the OH group (3′ end).

    • 5' end is the phosphate group (5′ end).

  • DNA strands are antiparallel; typically two forks moving in opposite directions (bidirectional replication).

DNA replication in eukaryotes (comparison)

  • Similar overall process but with differences:

    • Uses 5 DNA polymerases.

    • Thousands of replication origins.

    • Shorter Okazaki fragments.

    • Methylation in plant and animal cells mainly on cytosine bases (context-dependent).

Information transfer and the Central Dogma

  • DNA (gene) stores hereditary information.

  • Transcription: information in DNA is copied into mRNA.

  • Translation: mRNA is translated into protein.

  • Central Dogma of Genetics: DNA → RNA → protein.

Transcription: events and components

  • Types of RNA transcribed from DNA (as listed):

    • RNA primers (note: RNA primers are involved in DNA replication, not transcription; transcription produces mRNA, rRNA, tRNA)

    • mRNA

    • rRNA

    • tRNA

  • Three steps of transcription: Initiation, Elongation, Termination.

  • Transcription begins when RNA polymerase binds to the promoter sequence, then polymerizes the new chain using complementary bases.

  • Transcription proceeds 5' → 3'.

  • Transcription uses ribonucleotides with A–U pairing (A pairs with U in RNA).

  • Transcription stops at the terminator sequence.

  • Figure illustrating concurrent RNA transcription (for prokaryotes) is referenced.

Differences in Eukaryotic transcription and translation

  • Transcription occurs in the nucleus (also mitochondria and chloroplasts).

  • Multiple RNA polymerases (three main types in nucleus).

  • Numerous transcription factors are required.

  • mRNA processing occurs before translation (e.g., 5' cap, poly-A tail, intron splicing).

  • Translation specifics:

    • mRNA is translated in codons (three nucleotides).

    • There are 64 sense codons encoding 20 amino acids.

    • Start codon is AUG, which codes for methionine; all proteins typically begin with methionine.

    • Stop codons: UAA, UAG, UGA terminate translation.

    • Genetic code is degenerate (multiple codons can encode the same amino acid).

    • tRNA carries complementary anticodons to mRNA codons.

Let’s translate a protein: genetic code example

  • Given mRNA sequence (example):

    • AAAAAAAAAUGCGUUGGUGUGGUGGCGAUGCAGUAUGUUACUCAU AACCUAA AAAAGU AUG CGU UGG UGU GGU GGC GAU GCA GUA UGU UAC UCA UAA CCU

  • From the START codon (AUG), break into codons and translate until a STOP codon is reached:

    • Met – Arg – Trp – Cys – Gly – Gly – Asp – Ala – Val – Cys – Tyr – Ser – STOP

tRNA and the translation apparatus

  • tRNA structure:

    • The 3' acceptor end (the aminoacyl end) holds a specific amino acid; it is "charged" when attached to its amino acid.

    • The anticodon end complementary to the mRNA codon participates in decoding.

  • Process of translation has three stages:

    • Initiation

    • Elongation

    • Termination

  • All stages require additional protein factors.

  • Initiation and elongation require energy in the form of GTP.

  • Visual cue: a polyribosome in a prokaryotic cell illustrates multiple ribosomes translating a single mRNA simultaneously.

Regulation of gene expression

  • Constitutive genes: expressed at a fixed rate; always turned on.

  • Gene regulation is necessary; promoter region or "switch" at the gene's start controls RNA polymerase binding.

  • Other genes are expressed only as needed:

    • Repressible genes: transcription tends to be ON but can be downregulated (repressed).

    • Inducible genes: transcription can be turned on by removal of a repressor.

    • Catabolite repression: regulation based on available carbon source.

Regulating groups of genes: operons

  • Operons: a group of genes located together in the DNA and regulated together.

  • Prokaryotic operons can be:

    • Inducible operons (e.g., lac operon): typically off and transcribed only when inducer is present (lactose converts to allolactose to inactivate the repressor).

    • Repressible operons (e.g., trp operon): typically on and transcribed until repressed.

  • Lac operon regulation example:

    • Inducible operon: repressor inactive → operon ON; CAP–cAMP complex can enhance lac operon transcription.

  • Trp operon regulation example:

    • Repressor active → operon OFF.

  • Figures referenced illustrate the lac and trp operons and regulatory mechanisms.

Mutation and genetic variation

  • Mutation: a change in the base sequence of DNA.

  • Mutations can be silent (no effect), beneficial, or harmful.

  • Mutagen: an agent that causes mutations.

  • Spontaneous mutations: occur without mutagens.

  • Frameshift mutations: triplets displaced, usually due to insertions or deletions.

  • Base substitution (point mutation): single base substitution.

  • The genetic code table (Figure 7.12) relates codons to amino acids and stops.

  • Types of mutation examples:

    • Silent mutation

    • Missense mutation

    • Nonsense mutation

    • Frameshift mutation

    • Frameshift deletion (example category for disruptions)

Sources of mutations and DNA damage

  • Spontaneous mutation: natural replication errors.

  • Mutagens: chemical agents, UV radiation, and ionizing radiation.

  • UV radiation causes thymine dimers, disrupting DNA structure.

  • Ionizing radiation (X-rays, gamma rays) creates ions that damage the deoxyribose-phosphate backbone.

  • The pyrimidine dimer (e.g., thymine dimer) is a common UV-induced lesion.

DNA repair and mutant testing

  • Mutants: descendants of a cell that fails to repair a mutation.

  • Wild types: cells typically found in nature.

  • Methods to recognize mutants:

    • Positive selection

    • Negative (indirect) selection

  • Ames test: a classic assay to detect mutagenicity.

Gene transfer and recombination

  • Recombination: insertion of new genes into a genome; exchange of genetic material between DNA molecules.

  • Vertical gene transfer: transmission of genetic material during reproduction between generations.

  • Horizontal gene transfer: transfer of genes between cells of the same generation.

  • Modes of horizontal transfer:

    • Transformation: uptake of naked DNA from the environment.

    • Transduction: viral-mediated transfer of DNA; generalize (any gene) vs specialized (specific gene).

    • Generalized transduction: donor DNA fragments packaged randomly by a transducing phage.

    • Specialized transduction: only certain donor DNA sequences transferred because of lysogeny site and prophage integration.

    • Bacterial conjugation: transmission of genetic material via cell-to-cell contact; requires a conjugative plasmid and a sex pilus.

Transposons

  • Transposons: segments of DNA that can move within the genome.

  • Simple transposons: contain insertion sequences with transposase enzymes for cutting and resealing.

  • Complex transposons: carry additional genes not directly connected with transposition.

Bacterial chromosome and genome organization

  • Bacterial chromosome: typically a single, circular chromosome.

  • E. coli chromosome size: about 4 imes 10^6 base pairs (4 imes 10^6 ext{ bp}).

  • Implication: medium-sized bacterial chromosome.

  • Gene density: average gene size ~1000 base pairs; estimate ~4000 genes in E. coli.

  • Visual reference: Figure 8.1b showing the Genetic Map of the E. coli chromosome.

Key notational and directional concepts

  • 5' and 3' ends:

    • 5' end (P end): phosphate group end.

    • 3' end (OH end): hydroxyl group end.

  • DNA strands are antiparallel, meaning one runs 5' to 3' in one direction and the other runs 3' to 5' in the opposite direction.

  • DNA replication often proceeds bidirectionally with two replication forks moving away from the origin.

  • The central dogma and gene expression involve crosstalk between DNA, RNA, and protein synthesis pathways, with regulation at transcriptional and translational levels as described above.