Lecture 5- Regulation of Gene Expression in Eukaryotes

Question 1

Do cells have the same genes? If so, what makes different tissue types?

Answer 1

• Genes do have the same genes within them but in any cell only a specific subset of these genes are active.
• The active genes determine which proteins are synthesized and these proteins characterize the cell and its properties.

Question 2

Cell specialization: how do your cells become more specialized?

Answer 2

• Cells become more specialized with every cell division they undergo.

Question 3

How do genes play a role in the individual to individual differences?

Answer 3

• The activity of genes is what plays a major role in our individualism. This plays more of a role than coding sequences do.
• The activity of our genes (high, medium, low) dictates which types of proteins are synthesized and the activity levels of proteins. This varies from individual to individual.

Question 4

What are epigenetic factors and what are the 6 factors?

Answer 4

• Epigenetic factors are those that influence chromatin configuration and influence gene activity without changing the coding sequences within genes themselves.
  1. ) DNA packaging: Chromatin configuration changes to make the promoter region more/less accessible to transcription factors.
  2. ) Facultative Heterochromatin: Some heterochromatin regions in some cells will be packaged as euchromatin regions in other cells; mostly they are heterochromatin regions, but some can decondense which allows for transcription due to molecular signals.
  3. ) Transcription initiation: Interaction between regulatory sequences and transcriptional activators/inhibitors.
  4. ) RNA Processing: Alternate splicing and cleavage of pre-RNAs.
  5. ) RNA Stability: Poly A tail and interfering RNAs regulate RNA longevity.
  6. ) Translation initiation: 5’ to 3’ UTRs and interfering RNAs regulate the rate of translation.

Question 5

What is the core promoter and regulatory promoter? Who binds to which?

Answer 5

• The RNA polymerase and transcription factors bind to the core promoter, provides a minimal transcription.
• Activators and repressors bind to the regulatory promotor where they can either increase transcription and activity of a gene or silence a gene.

Question 6

What are response elements and how are they coordinately regulated?

Answer 6

• Are short sequences found in the promoter region that bind certain transcription factors which then transcribe certain genes.
• They respond to stress and hormones to synthesize certain proteins.
• They are regulated coordinately because they work together. This way one event (heat/stress) can activate/repress certain genes whose proteins help the cell cope with that certain event.

Question 7

What prevents transcription factors to activate a gene? Give an example.

Answer 7

• Insulators block transcription factors.

- T.F. I can help stimulate the transcription of gene A but its effect on gene B is blocked by the insulator,

Question 8

What binds to the promoter region in order for transcription to occur? How does chromatin impact this molecule?

Answer 8

• Transcriptional activators must bind to the appropriate sequence in order for transcription to occur.
• Heterochromatin (tightly packed) does not allow transcription activators or factors to get into the DNA and bind to the promoter region.
• Euchromatin allows for this since it is loosely packed.

Question 9

What is DNAse I and where is most likely to work (condensed or loose chromatin)?

Answer 9

• DNAse I is an enzyme which cuts DNA next to pyrimidine nucleotides.
• Less condensed chromatin is better for DNAse I to work.
• More transcription active sites are more sensitive to this enzyme.

Question 10

What do chromosomal puffs indicate?

Answer 10

• They indicate active transcription sites/active genes.

Question 11

What does the Methylation of Histones do?

Answer 11

• Methylating histone proteins can enable or repress gene activity by regulating transcription.
• Depends on the amino acid being methylated.

Question 12

What does Acetylation of Histones do?

Answer 12

• Lysin in the positively charged tail of histone proteins and the negatively charged phosphate group in DNA interact to disallow transcription factors to bind to DNA.
• Acetylation of the histone tails allows the DNA to loosen from the interaction with the positively charged tail. This allows transcription factors to bind.

Question 13

What are chromatin-remodeling complexes?

Answer 13

• They are complexes of transcription factors and proteins that can move nucleosomes around exposing promoter regions.
• They can slide DNA down exposing promoter regions or they can change the conformation of DNA/nucleosomes to expose promoter regions.
• This affects gene activity and regulation.

Question 14

What happens when DNA gets methylated and where does methylation within the DNA usually occur?

Answer 14

• If DNA gets methylated, the genes in that region do not get translated.
• A most notable area where this occurs is the methylation of cytosines in the promoter region. This results in no transcription because transcription factors are unable to bind to these methylated regions.

Question 15

What is a trinucleotide repeat, where is it located, and what happens if its length increases?

Answer 15

• Trinucleotide repeated sequence is a CGG repeated sequence of 50-58 pairs which is found in the FMR1 gene.
• This sequence can increase in length during meiosis.
• If the sequence becomes more than 200 CGG’s long then there is an increase in methylation, a decrease in acetylation which leads to no transcription of the FMR1 gene.
• This results in Fragile X mental retardation.

Question 16

What happens in mammals if you have more than one X chromosome? Where do you get your X chromosomes from?

Answer 16

• If you have two X chromosomes, one will be inactivated.
• In some individuals, the X chromosome from your mom gets silenced in others the X chromosome from your dad gets silenced.

Question 17

What is the process of X-Inactivation?

Answer 17

• The X chromosome being inactivated produces the XIST (X-inactivation specific transcript) RNA which coats over the chromosome, supercondensed it and methylates the C promotor regions.
• The other X chromosome keeps itself active by producing the TSIX RNA which binds to the XIST RNA and doesn’t let prevents it from inactivating the second X chromosome.

Question 18

What is the X inactive chromosome called and what does it look like under a microscope?

Answer 18

• It is also known as the Barr Body and looks like a crumpled piece of paper underneath a microscope.

Question 19

What are CG islands, where are they located, what happens to them, and why are they important? Think imprinted genes.

Answer 19

• CG islands are found in the promoter regions of genes.
• You have two copies of the same imprinted genes from your mother and father so one needs to be silenced (doesn’t matter which one).
• You can only have one active copy of an imprinted gene.
• The CG islands get methylated in one of the genes which does not allow for transcription to occur (inactivity of the gene).

Question 20

Why do methylation patterns need to be reset and when do they need to be reset?

Answer 20

• Methylation patterns need to be reset during spermatogenesis and oogenesis in order to have one active copy of imprinted genes.
• If this does not occur properly then someone may end up with zero or two active imprinted genes.

Question 21

What are two ways epigenetic factors are influenced?

Answer 21

• Epigenetics can be influenced by early life experience and diet.
• Methylation of certain genes are influenced by these factors

Question 22

Are epigenetic factors heritable?

Answer 22

Yes they are

Question 23

Give an example of how inheritable epigenetics can be gender-dependent?

Answer 23

• Feeding mice a high-fat diet made them insulin resistant and obese. This actually caused their female offspring to be obese, not the male offsprings.
• An obese mother causes consistent obese daughters, while an obese father causes approximately 15% of his daughters to become obese.

Question 24

What is paramutation?

Answer 24

• An interaction between two alleles at a single locus where one allele causes a heritable change in the other allele.

Question 25

What are methyltransferases and what do they do? What process do they go through duting repliaction?

Answer 25

• Before replication, the DNA strand is fully methylated, but during replication, the DNA strands do not result in methylation.
• After replication a hemimethylated DNA strand results ( A DNA strand that has methyl groups on only one side
• Methyl groups attract Methyltransferase which methylates the other strand of DNA.

Question 26

What is the result of RNA Splicing?

Answer 26

• RNA splicing can determine the sex of organisms

- It can also create different proteins in different cells from the same gene.

Question 27

How is the RNA stabilized, why do they need to be stabilized, and what happens if it is now?

Answer 27

• RNAse degrades mRNA beginning at the Poly A tail side.
• mRNA needs to be stabilized so it can stay in the cytoplasm to be translated.
• mRNA is stabilized by Poly-A binding proteins, but once the As get down to about 10-30, the protein cannot bind anymore resulting in RNAse degrading the mRNA strand.

Question 28

What are the two main interfering RNAs?

Answer 28

• Small interfering RNAs (siRNA) and microRNAs.

Question 29

What are the similarities between the two interfering RNAs?

Answer 29

1. ) They are cleaved from a long, double-stranded precursor RNA by the enzyme Dicer in the cytoplasm.
2. ) Both join with proteins to make up the RISC (RNA-induced silencing complex).
3. ) They all rely on having a complementary base sequence with either the mRNA or gene itself.

Question 30

What is the difference between siRNAs and microRNAs.?

Answer 30

• siRNAs are either taken up by the cell or enter via vectors. They are exogenous and double-stranded.
• microRNAs are transcribed from sequences in the cell’s genome. They are endogenous and single-stranded. Can be found in the introns of genes.

Question 31

What are the three mechanisms for miRNAs?

Answer 31

1. ) Inhibition of translation: bind to 5’ end of mRNA which prevents the ribosome from binding
2. ) Cleavage of mRNA: The RISC includes endonucleases which cleave double-stranded RNA. After cleavage, the RNA is degraded.
3. ) They bind to complementary strands of DNA and attract methylating enzymes which methylate the DNA or histone and inhibit transcription.

Question 32

Why is it assumed that miRNAs also bind to genes?

Answer 32

• Fire and Mello experiment showed that two miRNAs per cell completely stopped protein production.
• This concluded that miRNAs needed to be able to bind to genes itself and stop transcription because two RNAs would not be able to bind to all the mRNAs present.

Question 33

What is ubiquitin and what does it do?

Answer 33

• Ubiquitin tags proteins to be degraded.

- It takes proteins to the proteosome to be degraded.