MOL2011: Molecular Biology: Genes and their expression

• Convenor: Dr Priscilla Johanesen
• 6 point unit
• Monash handbook

Topics in this unit include the molecular nature of nucleic acids and the mechanism by which they specify proteins; mutations and their consequences; the impact of viruses, mobile genetic elements and extra-nuclear organelle genomes on the genetic material of the nucleus; expression of genetic information and how it is regulated.

Lecturing Staff

Dr Priscilla Johanesen	Dr Nirma Samarawickrema	Dr John Boyce
Dr Dena Lyras	Dr Wan Shoo Cheong	A/Prof Hans Netter
Dr Alan Neale

Organisation of the unit

MOL2011 consists of 3 lectures per week and one 3 hour practical session per week.

The unit is a collaborative course run across three departments: Microbiology, Biochemistry and Molecular Biology, and Biological Sciences. Students will be exposed to laboratory methods in each of the three departments, and classes will include laboratory exercises, computer exercises tutorials, self-directed exercises, oral presentations and written reports. There will be three rounds of practical sessions: the three departments teaching in MOL2011 will each run a four-week practical session

Topics covered

INTRODUCTION TO THE COURSE.

The impact of molecular biology on biology and biomedical sciences. The future of molecular biology.

INFORMATION FLOW IN CELLS

THE STRUCTURAL BASIS OF CELLULAR INFORMATION:

Nucleic Acids: Molecules that carry genetic information; their chemical structures, nitrogen bases, nucleosides and nucleotides; Features of the Watson and Crick DNA double helix and its role in storing and transmitting information; The central dogma of Molecular Biology.

The Organisation of DNA: Structures of DNA (linear, circular, super-coiled DNA and alternative helical structures); Definitions of genes, chromosomes and genomes; Coding and non-coding DNA, repetitive sequences; DNA packaging in the prokaryotic and eukaryotic cells

DUPLICATION OF DNA

Definition of replication, its relevance and features; Enzymes involved in replication; Strand separation; Events at a replication fork; Repair mechanisms; How the process described above differs in the eukaryotic cell

THE EXPRESSION OF PROTEIN ENCODING GENES:

Transcription of Genes: Definition of terms: coding and template strands, cistrons, monocistronic and polycistronic mRNA, open reading frames, promotors; Initiation, elongation and termination of prokaryotic transcription; Transcription in the eukaryotic cell; Post-transcriptional modifications in brief

Proteins: Levels of structural organisation of proteins; Maintenance of the 3-dimensional structure and protein folding; Conjugated proteins (e.g. Nucleoproteins); Cellular functions of proteins and their importance (e.g. DNA binding proteins and restriction enzymes

Protein Synthesis: Decoding the message; The genetic code and its features; Activation of amino acids; The role of the ribosome; The mechanism of protein synthesis in prokaryotic cells; Differences between eukaryotic and prokaryotic protein synthesis; Protein synthesis in other cellular organelles (mitochondria and chloroplasts); Mistranslation and post-translational modifications; Protein targeting and sorting

THE EXPRESSION OF NON-CODING RNA GENES

The different classes of RNA and their functions; RNA processing (post-transcriptional); Biological roles of non-coding RNA

MUTATION

Introduction. Definitions: genes, genome, genotype, phenotype, mutation, wild-type, mutant, strain; Types of mutation: point, transition/transversion, deletion/frameshift; How mutations are inherited. Molecular causes of mutation: base analogues, chemicals which directly alter DNA, Inducers of error prone repair: tautomerisation, slippage, pyrimidine dimers,

Affect of mutations on phenotype. Silent, missense conservative/non conservative, nonsense; Forward/back mutation, polarity, reversion (first site, second site), nonsense suppressor, physiological suppressor.

Detection of mutations. Antibiotic selection, resistance to other chemicals, colony morphology, replica plating.

DNA repair. Proof reading, direct repair, recombination repair, excision repair, postreplication repair.

Large scale rearrangements. Homologous recombination, RecA, SOS response; Bacterial genome rearrangements; Chromosomal mutation, deletion, duplication, inversion, translocation, chromosome number

Mobile DNA as mutagenic agents. Bacteriophage, transposons, retroviruses;

Genetic variation in populations. Pathogenic bacteria/antibiotic resistance; Genetic variation in eukaryotes; somatic mutations, germ line mutations; Measuring mutation rates

The Ames Test

GENETIC ELEMENTS AND THEIR EXCHANGE

Bacterial DNA Elements. Bacterial plasmids: types of plasmids and their biological significance. Transposable elements: insertion sequence, composite transposon, transposition process.

Genetic Exchange Processes in Bacteria. Conjugation: F⁺ x F^- matings; conjugation process, Hfr conjugation, F' conjugation. Transformation: mechanism of transformation. Transduction: generalised and specialised transduction.

PROKARYOTIC GENE EXPRESSION AND REGULATION

Definition of the gene. Compare and contrast prokaryotes and eukaryotes (gene structure - promoter, introns, terminator region, enhancers).

The operon concept in prokaryotes - General description of an operon.

Detailed analysis of an inducible operon. The E.coli lac operon: general description, induction, catabolite repression, coordination of regulation.

Detailed analysis of a repressible operon. The E.coli trp operon: general description, repression, attenuation, coordination of regulation.

Global control systems

VIRUSES

Introduction to Viruses. Size; host range: bacterial, plant, animal, fungal; examples of some agricultural and human viral diseases; structure of viruses.

Viral Genomes. Nucleic acid types and configurations. Baltimore scheme

Bacteriophage replication. Overview of the virus multiplication cycle using selected bacteriophages as examples; attachment to cell surface; entry of viral nucleic acid; transcription to produce viral mRNA, translation to produce structural and non-structural proteins; genome replication; assembly of progeny virions; maturation and release.

Virus-cell interactions. Lysogeny: temperate bacteriophage, prophage, induction; eukaryotic cells: persistent and latent infections, transformation

Retroviruses-HIV. Retrovirus family; HIV: particle morphology; genome structure; basic overview of HIV lifecycle: entry; role of reverse transcriptase; genome integration; gene expression; virion formation and exit

EUKARYOTIC GENE EXPRESSION AND REGULATION

Eucaryotic DNA Elements. Genomes of chloroplasts and mitochondria; Examples of transposons in Drosophila and in humans; extranuclear genetic elements - organelle genomes.

Transcriptional Control. General/basic transcriptional control components in eukaryotes. RNA polymerase II. Cis-acting DNA sequences associated with general transcription components. Trans acting DNA-binding general transcriptional factors Gene-specific components in eukaryotes (developmental and environmental regulation). Response Elements, Enhancers, Repressors.

Control at the post-transcriptional level prior to translation. RNA processing (splicing, capping, cleavage and polyadenylation, transport to cytoplasm. Control of mRNA degradation - general mechanism (applies to most mRNA's) specific mechanisms (sequences in mRNA which promote or delay degradation (in vivo). Deadenylation-independent pathways. Endonucleolytic cleavage sites, ribosome stalling, specific mRNA binding proteins.

Post-translational control, post-translational modifications to proteins and importance in cell signalling. Mechanism for down-regulating general translation [Specific examples: negative (e.g. ferritin) and positive (e.g. yeast GCN4)]. Protein targeting to nuclei, mitochondria, chloroplast, vacuoles, lysosomes, ER. Non-reversible post-translational modifications (e.g. glycosylation). Reversible post-translational modifications and importance in function (phosphorylation, receptor kinases and MAP Kinase cascades).

Control of protein degradation in the cell. General proteolytic machinery of the cell - proteosome and ubiquitin. Cellular processes involving ubiquitin (cell cycle, DNA repair, stress responses etc) Ubiquitin genes. Specialised sequences promoting protein degradation (e.g. N-terminal amino acids and PEST regions). Cell death (apoptosis, oncosis and necrosis).