Remember: Final Draft of Posters Due at 10 am on Thursday!

1. Today: • Poster Hints • Microbial Genetics • Gene Regulation • Group Quiz 8 Remember: Final Draft of Posters Due at 10 am on Thursday!

2. An Introduction to Microbial Genetics

3. Bacteria Reproduce Asexually via BINARY FISSION Can this generate genetic diversity?!?

4. But, Bacteria still undergo GENETIC RECOMBINATION (combining DNA from two individuals into the genome of a single individual)

5. Genetic Recombination in bacteria occurs through three distinct processes: 1. TRANSFORMATION 2. TRANSDUCTION 3. CONJUGATION

6. TRANSFORMATION- • Def: The uptake of naked, foreign DNA from the surrounding environment. • The foreign DNA may be incorporated into the bacterial chromosome. • Some bacterial cells have specialized surface proteins for the uptake of DNA.

7. General notes about PLASMIDS… • Plasmids are small, circular, self-replicating DNA molecules separate from the bacterial chromosomes. • Some plasmids, like the F plasmids, can reversibly incorporate into the cell’s chromosome (an episome).

8. 2. TRANSDUCTION Def: Phage (bacterial viruses) carry bacterial genes from one host cell to another. • Generalized Transduction occurs when the phage accidentally transfers random bacterial genes instead of its own. • Specialized Transduction occurs when the phage takes a small adjacent region of the bacterial DNA with it.

10. 3. CONJUGATION Def: The direct transfer of genetic material between two bacterial cells that are temporarily joined. • Transfer is one-way, with the DNA donor (or “male”) attaching to the DNA recipient (“female”) with a sex pilus. • The ability to form sex pili and donate DNA results from the presence of an F factor.

12. R PLASMIDS R plasmids contain genes conferring resistance to antibiotics. R plasmids, like F plasmids, have genes that encode sex pili and enable transfer from one cell to another. Many R plasmids carry multiple antibiotic resistance genes.

13. R Plasmids acquire multiple resistance genes through TRANSPOSONs. Transposon: a piece of DNA that can move from one location to another in a cell’s genome. Transposase catalyzes the movement of the transposon from one location to another

14. Composite Transposons include extra genes sandwiched between two insertion sequences. Composite transposons can add a gene conferring antibiotic resistance to a plasmid already carrying genes for resistance. In an environment high in antibiotics, natural selection will favor multi-drug resistant bacterial clones.

15. Gene Regulation: Individual bacteria cope with environmental fluctuation at several levels:

16. Regulation of Gene Expression occurs through OPERONS. OPERONS utilize a segment of DNA called an OPERATOR. The operator controls the access of RNA polymerase to the genes.

17. OPERATORS function as switches to turn transcription ON or OFF. Operators are ON unless a specific REPRESSOR PROTEIN is bound to it.

18. This operon, the trp operon is an example of a REPRESSIBLE OPERON because transcription is INHIBITED when tryptophan binds to the regulatory protein. • INDUCIBLE OPERONs are STIMULATED when a specific small molecule interacts with a regulatory protein. • Examples of Inducible Operons: • the lac operon • the ara operon

19. INDUCIBLE OPERONS: The lac Operon

20. The lac OPERON is also positively regulated by cyclic AMP (cAMP) and the regulatory protein, cAMP receptor protein (CRP). Activated CRP stimulates transcription.

21. Thinking About Genomes… Understanding Genome Structure and Function! Why is genome structure/ function important?

22. Remembering Structure… Nucleosomes are formed of DNA winding around 8 histone proteins, two each of H2A, H2B, H3, and H4. The N-terminus of each protein extends outward forming a “histone tail”.

23. Remembering Structure… Nucleosomes condense into 30nm fibers due to interactions between the histone tails of one nucleosome, the linker DNA, and the nucleosomes on either side.

24. Remembering Structure… During prophase, chromosomes may condense further!

25. Thinking About Genomes… In metaphase chromosomes, the same genes always end up at the same locations. What does this tell us about chromosome packing?? Photo: V. Miszalok, U. Klingbeil, I. Chudoba, V. Smolej

26. The Importance of Gene Expression Cell Differentiation! Differences in cell types are due to differential gene expression. How might a cell regulate gene expression??

27. Gene Regulation at the Level of Chromatin Structure Heterochromatin vsEuchromatin??

28. Regulating Chromatin Histoneacetylation (-COCH3) prevents adjacent nucleosomes from binding to one another So does this activate or silence the acetylated region?

29. Regulating Chromatin Staining of Acetylated H3 Throughout the Cell Cycle. A field of cells containing interphase, prophase(P), prometaphase (PM) and metaphase (M); Michael J. Hendzel and Michael J. Kruhlak

30. Other Modifications to Histone Tails • Histones may also be Methylated (CH3) • New Model: Histone Code Hypothesis! Figure: two different metaphase spreads (human female) with preferential staining.   Barbara A. Boggs, Peter Cheung, Edith Heard, David L. Spector, A. Craig Chinault & C. David Allis

31. Remembering DNA Methylation Proteins that bind to methylated DNA may recruit histonedeacetylases! What would this enzyme do??

32. New Field: Epigenomics! Figure: Epigenomics.com

33. Next Up: Regulating at the Level of Transcription!

34. Regulating Transcription Regulation is almost always based on initiation!

35. Regulating Transcription: Transcription Factors Transcription factors may be general (required for transcription of all genes) or specific, required for high level expression of particular genes)

36. Regulating Transcription: Enhancers and Activators

37. Evidence for Enhancers Michael R. Botchan and his colleagues have produced visual evidence of this model of enhancer action. They created an artificial DNA molecule and observed how it interacted with enhancers using an electron microscope

38. Regulating Transcription Some specific transcription factors function as repressors! • Common Mechanisms: • Block binding of activators • Bind to their own control elements • Recruit histone deacetylases

39. Combinatorial Control Numbers and diversity of control elements suggest that a particular combination of control elements regulates transcription.

40. Gene Clusters for Coordinated Control • Eukaryotic genes may also be packaged together for regulation. But each has its own promoter and is individually transcribed. • More often, genes in a pathway are scatted on multiple chromosomes, but respond to the same combination of control elements.

41. Post-Transcriptional Regulation

42. Post-Transcription: RNA Processing (This helps explain our embarrassingly small number of genes!)

43. Post-Transcription: mRNA Degradation Shortening the poly-A tail can trigger removal of the 5’ cap, followed by breakdown of the mRNA Figure: Effect of TTP deficiency on the stability of TNF-a mRNA in bone-marrow derived macrophages J. Blackshear, Biochem. Soc. Trans.. (2001) 30, (945–952)

44. Post-Transcription: RNA Interference (RNAi) Experimental Observation: Injecting dsRNA into a cell can silence the corresponding gene! http://www.nature.com/focus/rnai/animations/animation/animation.htm

45. Post-Transcription: Initiation of Translation • Regulatory proteins can block attachment of ribosomes • Lack of poly-A tails blocks translation • Block may be global (all translation; i.e. egg cells, dormant plants)

46. Post-Transcription: Protein Processing and Degradation • Many proteins must be cleaved or modified to become active. • Proteins may also be labeled for destruction by attaching a small protein, ubiquitin, that attracts proteosomes

47. Genome Regulation and Cancer • Viruses may carry oncogenes (genes causing cancer) • Oncogenes are similar to proto-oncogenes (genes involved in normal cell division) in our genome

48. A Multistep Model for Cancer