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Eukaryotic Gene Regulation April 23, 2014 Richard D. Howells, PhD MSB E-643 howells@njms.rutgers.edu. Objectives. During this presentation you may Catch another 40 winks, or Understand key concepts regarding regulation of gene expression in eukaryotes. Eukaryotic RNA Polymerases. 3.

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objectives
Objectives

During this presentation you may

  • Catch another 40 winks, or
  • Understand key concepts regarding regulation of gene expression in eukaryotes
slide4

Eukaryotic RNA polymerases

The poisonous mushroom, Amanita phalloides, contains -amanitin, a cyclic octapeptide that blocks elongation by RNA Pol II. It is lethal at 10 nM. The initial reaction to ingestion of the mushroom is gastrointestinal distress, and 48 h later the subject dies, usually from liver dysfunction.

4

slide10

RNA Pol II

Preinitiation

Complex

10

slide11
General pattern of control elements that regulate gene expression in multicellular eukaryotes and yeast

From Transcriptional Control

Of Gene Expression, Molecular

Cell Biology, Lodish et al., eds.,

6th edition

slide14
Hormones that bind specific receptors that are stimulated to become ligand-activated transcription factors

These and related lipid-soluble hormones bind to receptors located in the nucleus and cytoplasm. The hormone-receptor complex functions as a transcription factor in the nucleus.

select members of the nuclear receptor family
Select members of the nuclear receptor family

The centrally located DNA binding domains each contain 2 copies of the

zinc finger motif. Ligand binding domains are C-terminal, and N-termini

often contain activation domains.

dna response elements that bind nuclear receptors
DNA response elements that bind nuclear receptors

Repeat sequences are indicated by red arrows.

hormone dependent gene regulation by glucocorticoid receptor
Hormone-dependent gene regulation by glucocorticoid receptor

AD: activation domain; DBD: DNA binding domain; LBD: ligand binding domain

activation of gene expression via elevated camp and phosphorylation of creb
Activation of gene expression via elevated cAMP and phosphorylation of CREB

CRE: cAMP response element;

CREB: CRE binding protein; CBP: CREB binding protein

slide20

Mutation of CBP causes pleiotropic developmental abnormalities:Rubinstein-Taybi syndrome

slide21

Regulation of gene expression via

post-translational modification of histone side chains

Acetylated

Lysine

Lysine

HAT: histoneacetyltransferase activity

HDAC: histonedeacetylase activity

histone acetylation deacetylation in transcriptional control of gene expression
Histone acetylation/deacetylation in transcriptional control of gene expression

The transcriptional activator Gcn5 has histoneacetyltransferase activity

histone acetylation deacetylation in transcriptional control of gene expression1
Histone acetylation/deacetylation in transcriptional control of gene expression

The transcriptional repressor Rdp3 has histonedeacetylase activity

slide24

Intron 1

Example: Developmental-Temporal Regulation of Factor IX Transcription

Factor IX Transcribed Region

AR

HNF4

Exon 1

AR binding site

HNF4 binding site

  • Factor IX is a blood clotting factor
  • Mutation at –20 cause Hemophilia B Leyden
    • Reduces HNF4 binding (Hepatocyte Nuclear Factor 4)
    • Factor IX levels increase with onset of puberty due to testosterone production
  • Mutation at –26 causes Hemophilia B Brandenburg
    • Reduces HNF4 binding and androgen receptor (AR) binding
    • Factor IX levels remain low throughout life
slide27

Model for cleavage and polyadenylation of pre-mRNAs in mammalian cells

CPSF- cleavage and

polyadenylation specificity factor

CStF- cleavage stimulatory factor

CFI,CFII- cleavage factors

PAP- poly(A) polymerase

PABP II- poly(A) binding protein

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