1. Regulation of Gene Expression
Chapter 18

2. How do Organisms Control the Level of Gene Expression?
Cells must only express genes when needed
Gene expression (transcription, translation) takes up large amounts of cellular energy and resources
Cells live frugal lifestyles – they conserve energy and resources
So genes will only be expressed when their products are needed.

3. How Prokaryotes Control Gene Expression
Bacteria control genes at the transcriptional level
In other words, the gene is either transcribed or not, based on certain external stimuli

4. The Operon
An operon is a segment of DNA that consists of the following:
A gene or group of genes
A promoter of the gene/group of genes
A region upstream of the gene called the an operator

5. Operons 5

6. Lac Operon – Inducible operon
The lac operon is always off and is turned on by an inducer molecule (allolactose). This is why it is called an inducible operon 6

7. The trp Operon - repressible operon
The trp operon is always on and is turned off only by end product – tryptophan. This is why it is called a repressible operon.                                                      7

8. trp Operon – repressible operon8

9. How do Eukaryotes Control the Level of Gene Expression?
Cells of more complex organisms turn on and turn off genes based on the functions of the cells – hence cells differentiate
Eukaryotes control genes at almost every level:
Regulation of Chromatin Structure
Regulation at the transcriptional level
Regulation at a post-transcriptional level
Regulation at a translational level
Regulation at a post-translational level

10. Regulation of Chromatin Structure
Histone acetylation prevents DNA from winding tightly around histones, allowing easy access to promoter sites (Deacetylation does the opposite)
DNA methylation causes DNA to wind tightly around histones, preventing easy access to gene promoters (Demethylation does the opposite)

11. Bacteria lack histones, although some archea have them
Histone subunits are:
2 units of H2A
2 units of H2B
2 units of H3
2 units of H4
Histone H1 is not in the core, but acts as a clamp and keeps the linker DNA in place
Histones are positively charged, so DNA which is negatively charged, wraps around them
Linker DNA links one
nucleosome to the next
Bacteria lack histones, although some archea have them

12. Histones have tails that get actylated by HAT enzymes (Histone Acetyl Transferases)
Histone acetylation makes their (+) charge more neutral, so DNA interaction is reduced
Histone
Acetyl group

13. DNA Methylation in conjunction with Histone deacetylation

14. Epigenetic Inheritance
Modification of chromatin does not change the DNA, only its expression.
However, this modification pattern IS inherited (remember genomic imprinting?)
Scientists now believe that certain environmental factors may play a part in promoting chromatin modification that causes expression or suppression of certain genes – e.g. one twin gets schizophrenia and another doesn’t. Certain cancers may also be caused that way

15. Regulation at the transcriptional level
Enhancers (proximal and distal)
Silencers
Transcription factors at promoters
General transcription factors
Specific transcription factors
All these play a role in regulating gene expression.
Enhancers increase the rate of a gene’s expression and silencers decrease it.
Transcription factors are needed if the gene is to be expressed at all.

16. Organization of a typical Eukaryotic Gene

18. Regulation at a post-transcriptional level
RNA processing – alternative splicing allows certain proteins to be made instead of others (all from the same gene)
mRNA Degradation – cytoplasmic nucleases degrade mRNAs so polypeptide synthesis stops. More mRNA is made later, if necessary
5’ caps and 3’ tails can be removed or changed and this will prevent translation

19. Pre-translational Regulation
Certain proteins in the cytoplasm can bind to the mRNA’s 5’ UTR and prevent ribosomes from binding
Any change in mRNA shape will prevent ribosome binding
Decreased length of poly-A tail will prevent translation

20. Ribosome stalls due to hairpin loop in mRNA
mRNA is then degraded by a cluster of exonucleases (exosomes)

21. Post-translational Regulation
Proteins can be ubiquitinzed and degraded in a proteasome

22. Non-protein-coding RNAs and Gene Regulation
MicroRNAs or miRNAs are small non-coding RNAs that were
Transcribed from DNA
Complexed with a number of proteins
These miRNAs have several bases that are complementary to some protein-coding mRNAs
The miRNA-protein complex can bind to these protein-coding mRNAs and prevent them from being translated
Nucleases eventually degrade the dsRNA

23. RNA Interference
A relatively new technology uses the concept of miRNAs to stop the expression of certain genes
This is done by creating small RNAs that have a corresponding sequence (antisense RNA) to mRNAs that will give rise to unwanted proteins
The small interfering RNAs (siRNAs) then bind to these mRNAs and block their translation

24. Sense and Antisense RNA
Messenger RNA (mRNA) is single-stranded. Its sequence of nucleotides is called "sense" because it results in a gene product (protein). Normally, its unpaired nucleotides are "read" by transfer RNA anticodons as the ribosome proceeds to translate the message.

25. RNAi
However, RNA can form duplexes just as DNA does. All that is needed is a second strand of RNA whose sequence of bases is complementary to the first strand; e.g.
5´   C A U G   3´     mRNA 3´   G U A C   5´     Antisense RNA The second strand is called the antisense strand because its sequence of nucleotides is the complement of message sense. When mRNA forms a duplex with a complementary antisense RNA sequence, translation is blocked. This may occur because the ribosome cannot gain access to the nucleotides in the mRNA or
duplex RNA is quickly degraded by ribonucleases in the cell
synthetic genes (DNA) encoding antisense RNA molecules can be introduced into the organism.

27. Flavr Savr tomato
Most tomatoes that have to be shipped to market are harvested before they are ripe. Otherwise, ethylene synthesized by the tomato causes them to ripen and spoil before they reach the customer.
Transgenic tomatoes have been constructed that carry in their genome an artificial gene (DNA) that is transcribed into an antisense RNA complementary to the mRNA for an enzyme involved in ethylene production. These tomatoes make only 10% of the normal amount of the enzyme.
The goal of this work was to provide supermarket tomatoes with something closer to the appearance and taste of tomatoes harvested when ripe. However, these tomatoes often became damaged during shipment and handling and have been taken off the market.

29. Zygote  Embryo  Adult organism
How does a single egg or zygote become a complete organism with many different tissues and differentiated cells?
How can this happen, when the zygote undergoes many rounds of mitosis – mitosis is supposed to produce identical daughter cells?

30. Cytoplasmic Determinants
Certain molecules such as maternal mRNAs, transcription factors and other proteins are localized in specific cytoplasmic regions of the unfertilized egg or zygote
These molecules affect cell fate decisions by segregating into different embryonic cells and controlling distinct gene activities in these cells (specialized transcription factors will only turn on certain genes).
Cytoplasmic determinants are also found in some post-embryonic cells, where they produce cytoplasmic asymmetry.
In dividing cells, this leads to asymmetric cell division in which each of the daughter cells differentiates into a different cell type. Also called localized cytoplasmic determinants or morphogenetic determinants.

31. Cytoplasmic Determinants and Cell Fate

32. THE END