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Chapter 15: Gene Regulation in Eukaryotes I: Transcriptional and…
Chapter 15: Gene Regulation in Eukaryotes I: Transcriptional and Translational Regulation
General transcription factors (GTF's)
Regulatory transcription factors (RTF's)
Domain = specific regions of regulatory proteins that have specific functions
Activators bind to enhancer elements
Repressors bind to silencer elements
α helix =
Secondary structure that binds to the major groove of the DNA double helix
Hydrogen bonding
Positively charged amino acids bind with negatively charged DNA backbone
Types of helixes
1) Helix-turn-helix
2) Helix-loop-helix
3) Zinc finger motif
4) Leucine zipper
Homodimers and Heterodimers
Regulatory elements are bidirectional / orientation-dependent.
5'-GATCCA-3'
3'-CTAGGT-5'
=
5'-TGGATC-3'
3'-ACCTAG-5'
Mediator - Mediates the interaction between RNA polymerase II and regulatory transcription factors
Activators stimulate ability of mediator to cause phosphorylation of the CTD
Repressors inhibit this ability
Control of RTF's:
The binding of a small effector molecule;
Protein-protein interactions;
Covalent modifications.
Glucocorticoid hormones / Steroid hormones
Enter cell
Bind to glucocorticoid receptor, releasing HSP90, exposing nuclear localization signal (NLS)
Two receptors form a homodimer which binds to the glucocorticoid response element (GRE), enhancing transcription
CREB Protein
Adenylyl cyclase catalyzes synthesis of cAMP
cAMP binds to protein kinase A (PKA)
Protein kinase A phosphorylates CREB protein dimer, which is bonded to CRE element, causing it to recruit CBP protein dimer
CREB protein acts as a transcriptional activator
Chromatin
Closed formation = transcription inhibited
Open formation = transcription enhanced
CRC's all have an ATPase subunit called DNA translocase, facilitates remodeling
Nucleosomes in 1) fibroblasts and 2) reticulocytes
=
1) Chromatin compacted
2) Chromatin decompacted from -500 to +200
cenH3 / CENP-A
Variant of histone H3
Found at the centromere of each chromosome and functions in the binding of kinetochore proteins. Necessary for proper segregation of eukaryotic chromosomes
Histone modification takes place at the amino-terminal tail of H2A, H2B, H3, and H4
Mating type SWItching
SWI5P binds to its enhancer
SWI
Nucleosome-Free Region (NFR)
Where the transcriptional start site (TSS) is located in eukaryotes; TSS is flanked by a +1 and -1 nucleosome
Nucleosomes and transcription
Activator protein recruits a chromatin-remodeling complex (ex., SWI) and binds to the enhancer site
RNA Pol II and pre-initiation complex form and bind to core promoter
Histones ahead of open complex are acetylated and evicted, histones are deacetylated and carried by chaperone proteins and reform behind RNA Pol II
DNA Methylation
Occurs via DNA methyltransferase on cytosine bases and inhibits transcription
Occurs on 5 position carbon on cytosine base
Transcriptional silencing: Methyl-CpG binding protein binds to a methylated CpG island to recruit other proteins and cause further compaction by converting chromatin to a closed conformation
1) Full methylation vs. 2) hemimethylation
Methyl group on cytosine protrudes into the major groove of the DNA, thus preventing binding of proteins into that groove
Yeast and drosophila have very little DNA methylation
Abundant DNA methylation in vertebrates and plants
2 to 7% of the DNA is methylated
Methylation is heritable, functions through "maintenance" (if parent strand of DNA is methylated, daughter strand also gets methylated)
CpG island
Unmethylated CpG regions correlated with active genes
Methylated CpG regions correlated with suppressed genes
1000-2000 BP's in length
Methylation of CpG islands may prevent or enhance the binding of regulatory transcription factors to the promoter region
Housekeeping genes vs Tissue-specific genes
Encode proteins required in most cells of a multicellular organism
Are highly regulated and may be expressed only in a particular cell type
mRNA stability
Length of PolyA tail
Destabilizing elements
PolyA tail:
200 BP's long
PolyA protein binds and increases stability
As mRNA ages, its tail is shortened by the action of cellular nucleases
Binding protein cannot bind if tail is less than 10-30 adenosines long. mRNA will then rapidly degrade by exo- and endonuclease
Destablizing elements:
Found in mRNA w/short half lives
Most commonly located at 3' end between stop codon and PolyA tail at the "3' untranslated regions UTR's"
Phosphorylation of eIF2 and eIF4:
Phosphorylation of eIF2 inhibits the rate of translation
Phosphorylation of eIF4 increases the rate of translation
EIF4:
Recognizes 7-methylguanosine cap at the 5' end of mRNA and facilitates the binding of the mRNA to the small subunit
Conditions that increase phosphorylation = signaling molecules that promote proliferation like insulin and growth factors
Conditions that decrease phosphorylation = heat shock and viral infection
Viral infection decreases so that less protein is produced
FE3+ Response Element
Transferrin imports FE3+ into cell
Trasnferrin + FE3+ become an endocytic vesicle
Iron released into the cytosol either becomes available to bind to cellular enzymes, or if FE3+ content is too high, is stored within ferritin
Low iron levels:
IRP (iron regulatory protein) binds to IRE (iron response element), causing a loop to form and inhibiting translation to make the ferritin protein
High iron levels:
IRP binds iron, causing a conformational change that prevents it from binding the IRE. Ferritin protein is synthesized
IRE: Found in 5'-UTR region of ferritin mRNA