Cell Bio: Chapter 31- Eukaryotic Transcription

What is a basal level of transcription?

Transcription that occurs using just the general transcription factors and RNA pol 2

What are specific transcription factors?

Help with transcription of specific genes + bind upstream

What are the three different types of regulated gene expression?

• Tissue specific: different genes are expressed at different levels in different cell types (ex: neurons make proteins to make neurotransmitters but other cell types don’t)

• Hormone responsive: different genes expressed at different levels due to hormones

• Developmentally regulated: different genes expressed at different levels at different embryonic levels

What is the proximal promoter sequence?

CAAT box and GC box are roughly 200 bp away from core promoter

Enhancer and Silencer sequences

Specific regions of DNA that increase or decrease transcription

• Can be 100-1000 bp away from core promoter

• Usually upstream but can be downstream

• Activators (transcription factors) can bind to enhancers

• Repressors (transcription factors) can bind to silencers

Why can enhancers and silencers be so far away from the core promoter but still affect transcription?

DNA can bend and twist so transcription factors bound to the enhancer or silencer can interact with transcription factor bound to core promoter or RNA pol.

Transcription factor structure

• TF contain DNA binding domain and either an activator (bind to the enhancer) or repressor (bind to the silencer) domain

• Almost all TF are dimers and most are homodimers but some can be heterodimers

• May also have regulatory site (third domain)

  • Ligand binding site: steroid hormone receptors bind steroids that increase transcription

  • Phosphorylation site

Activators

• Contain activator domain

  • Help recruit RNA Pol. to make it bind better and can bind to general TF

  • Binding of activator to enhancer triggers DNA bending

• May bind coactivator

Coactivators

Binds to activator to increase transcription

• Ex: chromatin remodeling factors (SWI/SWF) or histone modifiers (acetyltransferase or demethylase)

After coactivator has been bound activators bind mediator which triggers the assembly of RNA pol. and general transcription factors at promoter

Repressors

• Bind to general TF or RNA Pol. and sequester them causing decrease in transcription

• May bind to a corepressor

Corepressors

Bind to repressor to decrease transcription

• Ex: Chromatin remodeling factors & histone modifiers (methyltransferase or deacetylase)

What happens if a transcription factor binds to the enhancer but isn’t an activator?

Decrease in transcription because that TF is competing with activator

How is the level of transcription determined?

Determined by the relative strength of the number of transcription factors binding to enhancers v silencers

Structure of DNA binding domain

All contain a pair of recognition helices which contain basic amino acids

1. Leucine zipper: made of two alpha helices coiled together to form a dimerized coiled coil; leucine residues in between helices

   1. Ex: CREB: contains a phosphorylation domain

2. Helix-loop-helix: dimer made of 2 alpha helices, each helix connected via a loop; bind to major groove on DNA

3. Zinc Finger: Zinc ion causes alpha helix and beta pleated sheet to interact and make finger shape; contains 4 recognition helices

   1. Ex: steroid hormone receptor

4. Helix turn helix: dimer made of 1 alpha helix and a turn between them

   1. Proks: lac and trp repressor proteins; Euks: homeodomain proteins

cAMP response element binding protein (CREB)

• Binds to enhancer element (CRE: cAMP response element)

• cAMP activates protein kinase A and PKA phosphorylates CREB

• CREB binds CREB binding protein (CBP)

CREB binding protein (CBP)

• A co-activator

• Binds TF2B

• Histone acetyltransferase activity

Steroid Hormone Receptor

• Transcription factor that is mainly homodimers

• Ligand activated

• Glucocorticoid receptors recruit CBP and chromatin remodeling factors and increase transcription

• Glucocorticoid: DNA site the receptor binds

  • Each end of the glucocorticoid receptor binding site (glucocorticoid) has palindromic site where the bases are identical but inverted

    • 5’ to 3’ = AGAACA and 3’ to 5’ = TCTTGT

  • A palindromic site is a sign that the TF is a homodimer

Homeodomain

• Transcription factors containing a homeodomain bind to the homeotic genes which regulate the development of entire body parts (make homeoproteins)

• Homeotic genes contain highly conserved homeobox

  • Contain a helix-turn-helix DNA-binding domain

How can misexpressing homeotic genes affect gene expression?

Drosophila contain 9 homeotic genes and a mutation in antennapedia (homeotic gene) causes legs to grow from head