Lecture 8 - Regulation of Gene Expression

Question 1

Levels of gene expression control (3)

Answer 1

• Before transcription starts
• After transcription/before translation
• Post translationally

Question 2

Transcriptional regulation has __ regulatory elements.

Answer 2

two

1. Promoter - region upstream of a gene where RNA polymerase binds and regulation of gene expression occurs.
2. Enhancer - sequence of DNA that positively regulates gene expression upon binding of an activator from the distance.

Question 3

Transcriptional regulation involves __ regulatory proteins.

Answer 3

two

1. Activator - protein that binds to a regulatory element to increase gene expression
2. Inhibitor - protein that binds to a regulatory element to decrease gene expression.

Question 4

There are __ types of transcriptional regulation.

Answer 4

two

1. Negative regulation - a repressor protein prevents transcription.
2. Positive regulation - an activator protein stimulates transcription.

Question 5

Transcriptional regulation: Regulation of a gene depends on the recruitment/inhibition of ___ _______ to the ____. Prokaryotes and Eukaryotes differ by…

Answer 5

RNA polymerase; promoter.
Prokaryotes: sigma factor binds to -10 and -35 sites
-10: 5’-TATAAT-3’
-35: 5’-TTGACAT-3’
Eukaryotes: General Transcription Factor II D binds to TATA box.

Question 6

E. coli prefers ___ as a source of energy, but it can also use ___. It regulates genes encoding for proteins involved in the digestion of these energy sources. ___ _____ is used to regulate digestion of lactose.

Answer 6

glucose; lactose

lac operon

Question 7

Operon

Answer 7

a set of genes that are regulated under the same regulatory elements.

Question 8

Lac Operon has ___ genes that code for ___ proteins

Answer 8

three

• B-galactoside permease: carrier protein that moves lactose into the cell
• B-galactosidase: enzyme that hydrolyses lactose
• B-galactoside transacetylase: transfers acetyl groups from acetyl CoA to certain B-galactosides.

Question 9

Lac Operon is an example of _____ regulation.

Answer 9

Negative.

• expression of the lac operon is off in the absence of lactose.
  
  • the lac repressor is a protein that binds to the Lac operon promoter and inhibits its expression.
  • when lactose is absent, the lac repressor is active
  • when lactose is present, the lac repressor is inactive.

Question 10

When lactose is present, _____ is produced. Binding of _____ to the ___ ______ induces a _______ _____ that _____ binding to the promoter.

Answer 10

allolactose

allolactose; lac repressor; conformation change; inhibits

Question 11

Lac operon can also be an example of _____ regulation.

Answer 11

positive

• glucose is still E. coli’s preferred energy source.
• lac operon has a “weak” promoter. The sigma factor binds poorly to the -10 and -35.
• low glucose environment produces an accumulation of cAMP
• cAMP-receptor protein CRP (catabolite activator protein) requires binding of cAMP to be able to bind to the promoter.
• CAP (cAMP-CRP complex) is an activator.

Question 12

Binding of ____ to promoter will recruit ___ _______ and therefore ________ transcription.

Answer 12

CAP; RNA polymerase; enhance

Question 13

Regulation of trp operon: Trp operon codes for enzymes that catalyze the synthesis of ______ (2)

Answer 13

tryptophan

• activated when trp is low
• high levels of trp will induce the trp to bind to and activate repressor.

Question 14

There are ___ levels of gene regulation in eukaryotes

Answer 14

six

1. transcriptional level
2. DNA packing - histone modifications
3. DNA methylation
4. Post transcriptional regulation
5. Translational level
6. Post translational regulation

Question 15

Eukaryotic transcriptional regulation: ___ ________ __ can only bind to the promoter after _____ _________ _____ have assembled on the _________. ____ binds to the ______ box; then other factors bind to form an _______ ______. Assembly of ___ ______ will _____ transcription.

Answer 15

RNA polymerase II; general transcription factors; chromosome.
TFIID; TATA; initiation complex.
core complex; initiate

Question 16

There are ___ regulatory elements for eukaryotic transcriptional regulation.

Answer 16

two

1. Enhancers: regulatory sequences where transcription factors bind to activate transcription or increase rate of transcription.
2. Silencers: sequence of DNA where transcription factors bind to repress transcription.

Question 17

Enhancers/ DNA looping (2)

Answer 17

• enhancer is a regulatory element that act at a distance
  
  • DNA has the capacity to bend >150 bp.
  • this is true for both euks and proks
• an activator can bind and recruit the RNA polymerase.

Question 18

If genes have common regulatory sequences, they can be regulated by the ___ transcription factors

Answer 18

same

Question 19

DNA packing: ____ modifications

Answer 19

histone

- eukaryotic cells have their DNA packed by histones which are proteins that help with DNA packing.

Question 20

Histones (4)

Answer 20

• H2A, H2B, H3 and H4 are known as the core histones
• Nucleosomes form when two of each of the core histones come together in an octamer where DNA can bind.
• Ionic interactions hold the nucleosomes together
  
  • negative charge of phosphates (DNA) interacts with positive charge of amino acids on the histones
  • DNA packing is determined by interactions between histones and the DNA

Question 21

Chromatin remodeling

Answer 21

RNA polymerase needs to be able to access the genes to be able to transcribe them
- RNA polymerase and transcriptional factors cannot transcribe areas of packed DNA.

Question 22

Histone modifications - acetylation (2)

Answer 22

• histone acetyltransferases (HATs) add an acetyl group at the end of a conserved Lys
• acetylation of histones disturbs histone-DNA interaction opening nucleosomes which allow accessibility of DNA. Histone acetylation correlates with active genes.

Question 23

Histones can also be modified by ___ and ____

Answer 23

methylation and phosphorylation

methylation: inactivates genes
phosphorylation: effects depend on which amino acids are involved.

Question 24

DNA methylation

Answer 24

• catalyzed by DNA methyltransferase
• usually occurs in regions rich with C and G doublets (CpG islands): high methylation of CpG islands correlates with regions of low transcription (gene silencing)

Question 25

DNA packing: two types of chromatin

Answer 25

1. euchromatin - diffuse, light-straining; contains DNA that is transcribed.
2. heterochromatin - condensed, dark-staining; contains genes not transcribed.

Question 26

Imprinting (2)

Answer 26

• in mammals, eggs and sperm develop different methylation patterns
• for about 200 genes, offspring inherit and inactive (methylated) copy and an activated (demethylated) one.

Question 27

X-inactivation (4)

Answer 27

• males (XY) and females (XX) contain different numbers of X-linked genes, yet for most genes transcription rates are similar.
• early in development, one of the X chromosomes in females is inactivated. Which X chromosome gets inactivated is random in each cell.
• RNA transcribed from Xist (X inactivation-specific transcript) binds to the chromosome, spreading the inactivation.
• The DNA is heavily methylated, and unavailable for transcription, except for the Xist gene.

Question 28

Post transcriptional regulation: Alternative Splicing (3)

Answer 28

• different mRNAs can be made from the same gene.
• there are many more human mRNAs than there are coding genes.
• specific sequences identify the presence of an intron
  
  • snRNPs bind to these sequences and recruit components of the spliceosome
  • by changing the capacity of snRNPs to recognize an intron, you can alter gene expression.

Question 29

Post transcriptional regulation: MicroRNAs (miRNA)

Answer 29

• microRNAs are small RNAs (~22 nucleotides long) that produce a hairpin
• they can target mRNA for degradation or translational repression
• miRNA’s target complementary sequences
• miRNAs have many targets as complementarity does not have to be perfect.

Question 30

Translation level: Repression

Answer 30

• miRNAs can inhibit translation
• GTP cap on 5’ end of mRNA can be modified since euks need 5’-cap for translational initiation
• repressor proteins can block translation directly

Question 31

Translational level: Repression by ______

Answer 31

Riboswitch
- translational repressor proteins can bind to noncoding regions of mRNA and block translation by preventing it from binding to a ribosome.

Question 32

Post translational regulation: degradation & modification
Proteins can be targeted for destruction when ______ attaches to it and attracts other ______, forming a ______ chain.
This ______ protein is recognized by the ________ and _____.

Answer 32

ubiquitin; ubiquitins; polyubiquitin

polyubiquitin; proteasome; degraded.