Gene Regulation

Question 1

Two types of gene expression

Answer 1

• Constitutive: protein product of a housekeeper gene is always needed, though different cells can have different basal levels of that protein
• Regulatated: basal level of a protein product can rise or fall depending on the cell, the environment, and the signaling

Question 2

Where transcription begins is defined as

Answer 2

+1

Question 3

3 elements of the prokaryotic promoter

Answer 3

• UP: A-T rich region that allows the DNA to more easily unwind, promoting transcription
• -35 element may ehlp or hinder transcriptional machinery to bind
• -10 element (TATAAT) binds factors that allow the transcriptional machinery to bind and start transcription at +1
  
  • Determines the +1 site AND which strand will be transcribed

Question 4

Explain how promoters, operators, activators, repressors, and effector signals cause negative and positive gene regulation in prokaryotes

Answer 4

• Polycistronic genes are all operated by one operon.
• Specificity factors will bind the -10 and -35 regions to promote transcriptional initiation
• Repressors can bind to the operator sequence in the promoter; Activators bind adjacent to the promoter.
  
  • Both are modified by an effector signaling molecule that either helps it bind or causes it to fall off

Question 5

Lactose operon when there’s no lactose and glucose (preferred) is present

Answer 5

Don’t need the lactose operon, so cAMP is low and the repressor is bound to the operator

Question 6

Lactose operon when lactose is present but there’s still glucose (preferred) present

Answer 6

Want to transcribe a low level of lactose

• Lactose (effector signaling molecule) takes the repressor off
• cAMP (effector signaling molecule) is still LOW
  
  • –> low CAP (activator) binding at the CAP site
    
    • –> low levels of s70 polymerase bind

Question 7

Lac operon when lactose is present, but glucose is not

Answer 7

Want to transcribe high level sof lactose

• Lactose (effector signaling molecule) takes off the repressor
• cAMP (effector signaling molecule) is high, helping CAP bind to the CAP site
  
  • –> Stimulates s70 Polymerase to bind

Question 8

2 big differences between prokaryotic and eukaryotic promoter

Answer 8

Eukaryotic promoter is much larger and not based on operons, so the promoter is very close to the transcription start site

Question 9

Elements of the eukaryotic proximal promoter

Answer 9

• Initiator (Inr) at -10: contains the +1 site and is where DNA unwinds for transcription
• TATA box at -30: binds TATA binding proteins to form the transcription preinitiation complex (PIC)
• Binding sites for specific transcription factors

Inr & TATA define strand usage and transcription start

Question 10

TBP in preinitiation complex (PIC) assembly

Answer 10

TBP is a TATA-binding protein that orients PIC and identifies strand usage

Question 11

TFIIH in PIC assembly

Answer 11

Two functions:

• Helicase activity
• Serine kinase activity to phoshporylate the carboxyl-termianl domain (CTD) of RNA poly II activates the PIC and transcription

Question 12

How do eukaryotic activators work?

Answer 12

Activators

1. Bind enhancer sequences and upstream activating sequences (UAS) and bends the promoter with HMG proteins to interact with the Mediator complex​
2. Interacts with histone acetyltransferases (HATs) to open the chromatin

Bending the promoter & remodeling the DNA into a more active structure facilitates assembly/activation of PIC

Question 13

How do eukaryotic repressors work?

Answer 13

Repressors

• Bind UAS and enhancers to displace the activators –> prevents bending to interact with Mediator
• Attracts histone deacetylases (HDACs) and histone methyltransferases (HMTs)

Question 14

Relate gene expression to Wilms tumors/nephroblastomas

Answer 14

Loss of function mutation in WT1: transcriptional repressor for proto-oncogenes (EGF family member amphiregulin, BCL-2 anti-apoptosis)

–> Oncogenic expression causes unilateral kidney tumors; most common abdominal tumor in children

Question 15

Relate gene expression to Pit-1 and growth hormone expression

Answer 15

Pit-1: transcriptional activator for growth hormone and prolactin; has 2 types of DNA-binding regions and a transactivation region

E174G mutation of Pit 1 prevents it from binding the GH promoter

–> low GH expression, affecting long bone growth, IGF-1 production, and nitrogen uptake