Long noncoding RNA, greater than 200 nucleotides, gene regulation
What do ribosomal and transfer RNA (rRNA/tRNA) function for?
Protein synthesis.
How are eukaryotic promoters more complicated than bacterial promoters?
Each RNA pol has distinct promoters
Each euk. RNA pol has to find specific promoter in billions of DNA nucleotides
Transcription regulated to produce right amount of RNA at right time and place
How is transcription in eukaryotic RNA polymerase regulated?
Cis acting elements (ex: promoters/enhancers): DNA sequences that control transcription of a gene are are located on the same molecule of DNA
Transcription factors: proteins that regulate gene expression
What types of RNA pol II promoters are there?
TATA containing and TATA-less promoters.
Describe a TATA-less promoter.
Found in RNA pol II, has enhancer upstream, an initiator element (Inr), and either a TATA box or a downstream promoter element (DPE) downstream
What do enhancers do? In what RNA polymerases are they commonly found?
Bind transcription factors to regulate gene expression, separate from promoter regions in RNA polymerase II.
Where is RNA polymerase I found and what does it do?
Located in nucleolus and transcribes genes for rRNA.
What is a key feature about RNA polymerase I?
Large, coordinated quantities of rRNA needed to generate ribosomes
How many arrays do humans have of RNA pol I?
5 arrays of 300-400 rRNA genes
What is the setup of the RNA pol I promoter?
Promoter= UPE + rInr
UPE: upstream promoter element (~150 bp upstream)
rInr: ribosomal initiator element: lies just upstream of initiation site
What is the setup of RNA pol I?
28S-----UPE---rInr-----18S---5.8S--28S
What is the nucleolus?
Ribosome producing factory within the nucleus?
What does RNA pol III do?
Synthesizes RNA and 5S ribosomal RNA
Where are promoters in RNA pol III?
Promoters within the transcribed sequence, downstream of start site
Large quantities of ________ must be produced by eukaryotic cells to generate _________.
5S rRNA, ribosomes
How many tRNA genes do eukaryotes have?
Euk: hundreds of tRNA genes
~500 in humans
Describe the type I RNA pol III promoter.
A block and C block nearby leads to transcription
Describe the type II tRNA RNA pol III promoter.
A block and B block downstream leads to transcription
What binds to promoters to regulate gene expression?
Transcription factors
What is TFII? What are their individual names?
Set of transcription factors for RNA pol II; TFIIA, TFIIB, etc.
What must genes have for there to be transcription factors that bind to the TATA box?
TATA box, TATA box-binding protein (TBP), a component of TFIID
What is the order in which TFII binds to the promoter (TATA box)?
1. TFIID, 2. TFIIA, 3. TFIIB, 4. TFIIF, 5. RNA pol II, 6. TFIIE, 7. TFIIH
DABFpolEH--What does the Canadian bear say to the hiker when he is stuck in the honey pot?
Do A Bear a Favor and Pull, EH?
What does TFIIH do?
Opens DNA double helix in eukaryotes
What does TFIIH do?
Phosphorylates the carboxyl-terminal domain (CTD) of RNA pol II. Allows polymerase to leave the promoter and begin transcription
Why is TFIIH important?
Facilitates the transition from initiation to elongation phase
Where do transcription factors bind?
Enhancers and promoter regions
What do promoter regions often contain?
Transcription factor binding sites
Why do enhancers contain multiple binding sites for transcription factors?
Different transcription factors recognize different sequences
T/F: Specific transcription factors regulate only one target gene.
False. may regulate hundreds of different target genes
What are enhancers?
Cis-acting elements that have no promoter activity, but can stimulate transcription from promoters
Where are enhancers located?
Close or far. Can be thousands of nucleotides from the start site.
Give an example of enhancer regulation.
Hemoglobin regulation--tissue specific expression of 5 beta hemoglobins. Transcription controlled by enhancers in the Locus Control Region (LCR) and promoters
What are transcription factors?
Bind to specific DNA sequences to regulate gene expression
What are coactivators?
Bind to transcription factors to facilitate transcription by RNA polymerase II.
What are mediators?
Large complex of proteins that act as a bridge between enhancer-bound activators and proteins, including polymerase, at the promoter
What are other proteins that play a role in transcription regulation?
1. DNA binding regulatory proteins (activators/repressors)
2. DNA looping
3. Combinatorial control
Describe the DNA binding regulatory proteins: activators and repressors.
Activators: bind enhancers, stimulate transcription of a gene.
Repressors: reduce or prevent transcription of a gene
How does DNA looping regulate transcription?
Allows enhancers to come into contact with promoters
Describe combinatorial control.
Multiple regulators working together. Different effects on transcription depending on components already bound to enhancer and promoter
How can transcription factors control cell identity?
Pluripotent stem cels can differentiate after stimulation
How are induced pluripotent stem cells (iPS) generated from cells?
Insertion of genes for only 4 specific transcription factors--Sox2, Oct4, cMyc, and Klf4
What are other ways gene expression can be regulated?
Steroid hormones
What does estradiol do?
Controls the genes in the development of female secondary sex characteristics and reproductive cycle
How does estradiol exert its effects?
Forms a complex with a specific receptor protein called estrogen receptor
What is the estrogen receptor part of?
Regulatory protein: nuclear hormone receptor, which are activated by binding of small molecules or ligands
What are DNA binding domains?
Zn-finger domains that confer specific DNA binding.
What do nuclear hormone receptors bind to?
Specific regions of DNA called response elements
What does the estrogen receptor bind to?
Estradiol response element (ERE)
What does a ligand binding domain cause?
Structural change that enables the receptor to recruit other proteins to regulate transcription
What does the nuclear hormone receptor act as?
Hormone-responsive transcriptional switch
How do nuclear hormone receptors control transcription?
Recruit corepressors or coactivators
Y/N: Does ligand binding alter the ability of the nuclear hormone receptors to bind DNA?
No
In the unbound form, some receptors bind to corepressors and inhibit transcription--what happens upon hormone ligand binding?
Repression stops and transcription starts. Ligand-bound form of the receptor may recruit a coactivator that stimulates transcription
What are agonists?
Ligands that activate nuclear hormone receptors
What are antagonists?
Ligands that inhibit a nuclear hormone receptor
What are some forms of breast cancer dependent on?
The estradiol-receptor complex
What antagonists can breast cancer? What is the group called?
Antagonists to the estrogen receptor: tamoxifen and raloxifene. Selective estrogen receptors modulators (SERMS)
How does histone modification remodel chromatin?
DNA is packaged by chromatin, which can impede transcription
What can coactivators do to histone modified chromatin?
Stimulate transcription by loosening interactions between histones and DNA. Makes the DNA more accessible to transcriptional machinery
How can histones be modified?
Methylation, phosphorylation, etc.
What do Histone AcetylTransferase enzymes (HATS) do?
Modify histones. Components of coactivators or are recruited by coactivators.
What does acetylation do?
Reduces the affinity of histones for DNA. Generates docking site for proteins with bromodomains.
Recruitment of additional components of transcriptional machinery
Recruitment of chromatin-remodeling machines, ATP powered complexes that make DNA in chromatin more accessible
In what steps does chromatin remodeling facilitate transcription
