1. Chapter 18 –Regulation of Gene Expression

2. Operons I
Def: Unit of genetic function consisting of coordinately related clusters of genes with related functions (transcription unit)
Repressible (trp operon):
tryptophan (a.a.) synthesis
promoter: RNA polymerase binding site; begins transcription
operator: controls access of RNA polymerase to genes (tryptophan not present)
repressor: protein that binds to operator and prevents attachment of RNA polymerase ~ coded from a regulatory gene (tryptophan present ~ acts as a corepressor)
transcription is repressed when tryptophan binds to a regulatory protein
animation

3. Operons II Inducible (lac operon): lactose metabolism
Def: Unit of genetic function consisting of coordinately related clusters of genes with related functions (transcription unit)
Inducible (lac operon):
lactose metabolism
lactose not present: repressor active, operon off; no transcription for lactose enzymes
lactose present: repressor inactive, operon on; inducer molecule inactivates protein repressor (allolactose)
transcription is stimulated when inducer binds to a regulatory protein
Animation
Animation 2

4. Operons II (cont)
the lac operon and trp operon are examples of negative control
the lac operon is also under positive control, cAMP receptor protein (CRP)
cAMP binds allostericly to CRP switching it to an active conformation, bins upstream of the promoter
this enhances RNA polymerase’s affinity for the promoter
when glucose levels are high, cAMP is scarce, slowed transcription
when glucose levels are low, cAMP is abundent, efficient transcription
Tutorial
Tutorial 2

5. Chromatin Def: complex of DNA and proteins
DNA Packing – DNA wrapped around histone proteins, forms a:
Nucleosome – ”beads on a string”; basic unit of DNA packing
Heterochromatin – highly condensed interphase DNA (can not be transcribed)
Euchromatin – less compacted interphase DNA (can be transcribed)

6. Chromosome packaging

7. Gene Expression
All organisms must regulate which genes are expressed at any given time.
A typical human cell might express about 20% of its protein coding genes at any given time.
Must locate right genes at right time, a very difficult and precise process
The difference between cell types is not due to different genes, but to differential gene expression (expression of different genes by cells with the same genome.
Diagram shows different stages of gene expression regulation.

8. Histone Modification
Genes with highly packed heterochromatin usually not expressed
Histone acetylation – loosens heterochromatic structure enhancing transcription
DNA methylation – addition of methyl groups to DNA bases can reduce transcription (example the mammalian X chromosome)

9. Typical Eukaryotic Gene
most eukaryotic genes have multiple control elements – segments of noncoding DNA that help regulate transcription
proximal control elements – close to the promoter
distal control elements – usually called enhancers, located further upstream than proximal control elements

10. Transcription Reg. – Activators
an activator – a protein that binds to an enhancer and stimulates gene transcription
there are no operons in eukaryotic organisms, but genes in a group will have the same regulatory sequences to coordinate expression, example heat shock response and steroid hormone action
Animation

11. Transcription Regulation

12. Activators (cont.)
Different combinations of control elements can be used to activate transcription
Liver and lens cells – both have genes for albumin (only expressed in liver cells) and crystallin (only expressed in lens cells), but only certain activators are present in each type of cell.

13. RNA Processing
Different mRNA molecules are produced from the same primary transcript, depending on which RNA segments are treated as exons and which as introns
Controlled by cell specific regulatory proteins

14. RNA Processing

15. mRNA Degredation
longevity of mRNA affects how much protein synthesis it directs
some can last for minutes, some for weeks
degradation could begin with the removal of the 5’ cap or the poly A tail

16. ncRNAs (non-coding RNA)
microRNA (miRNA) – small single-stranded RNA molecules that bind to complementary sequences on mRNA, are endogenous and do not have to match perfectly with target mRNA allowing them to match up with hundreds of different mRNA strands
Will either degrade the target mRNA or block translation
Small interfering RNA (siRNA) – similar in function to miRNA but are exogenous, smaller, and bind perfectly with target mRNA.
Both classes (along with shRNA/smallhairpin and piRNA/piwi-associated) are called RNA interference or RNAi

17. Control of Translation -overview
repression or destruction of mRNA with some form of ncRNAs, specifically RNAi.
repressor proteins that bind to specific sequences or structures within the leader region at the 5’ end preventing ribosome attachment (local control)
inactivate necessary translation factors (global control)

18. Protein Processing
Many polypeptides need to be modified before they can properly function – addition of sugars, lipids, additional amino acids; cleaving chain into two or more pieces
Can alter the targeting of a protein – if the protein cannot reach the target site, it cannot function

19. Protein Degradation
Select a protein for degradation – attach ubiquitin, large proteasomes recognize ubiquitin and degrade the protein (could lead to cancer if cell cycle proteins become impervious to proteasomes)

20. Cancer
Proto-oncogene – normal cellular genes that code for proteins involved in cell growth and differentiation
Oncogene – cancer-causing genes
How can a proto-oncogene become an oncogene?
movement of DNA; chromosome fragments that have rejoined incorrectly
amplification; increases the number of copies of proto-oncogenes
proto-oncogene point mutation; protein product more active or more resistant to degradation
Tumor-suppressor genes – changes in genes that prevent uncontrolled cell growth (cancer growth stimulated by the absence of suppression)

21. Interference with cell signaling
Products of proto-oncogenes and tumor suppressor genes are involved in cell-signaling pathways.
2 examples are the ras proto-oncogene and the p53 tumor suppressor gene.
Mutations in the ras occur in about 30% of human cancers, and mutations in p53 in more than 50%.

22. ras proto-oncogene
Ras protein is a G-protein that relays a signal from a growth factor whose cellular response is the synthesis of a protein that stimulates the cell cycle.
Mutations in the ras gene can lead to production of a hyperactive Ras protein that results in increased cell division and possibly cancer.

23. p53 tumor suppressor gene
Codes for a transcription factor that promotes the synthesis of cell cycle-inhibiting proteins.
If mutation occurs (due to UV light for example) the cell cycle-inhibiting proteins gone, increased cell division and cancer possible.

24. Human Genome
98% of the human genome does not code for protein, rRNA, or tRNA
transposable elements (jumping genes) – transposons (DNA intermediate) and retrotransposons (RNA intermediate)
Alu elements – 300 nucleotides long, do not code for protein, seem to exist only to replicate, can act as retrotransosons, sometimes called “selfish DNA”, thought to be valuable for evolution of primates and are used to show evolutionary relationships amongst primates
simple sequence DNA – repeating units found in centromeres and telomeres

25. Transposable Elements
Transposons – move by “cut and paste” or “copy and paste”
Retrotransposons – use an RNA intermediate, always leave a copy in original location in genome (more common in Eukaryotes than transposons)

26. Genome Evolution
Evolution of TPA gene through exon shuffling and duplication (see figure)
Transposable elements can contribute to evolution