Exam #4: Transcription, RNA Processing & Control of Gene Expression Flashcards
3 Ways RNA Differs from DNA
1) Ribose sugar instead of Deoxyribose
2) No thyamine, Uracil instead
3) Single stranded instead of double stranded
What is the significance of the DNA template in transcription?
DNA is the template for the synthesis of the complimentary RNA molecule
2 Differences Between DNA & RNA Polymerase
1) RNA Polymerase initiates RNA synthesis without a primer
2) RNA Polymerase is much more error prone
RNA Class Translated into Protein
mRNA
DNA Sequence that Specifies Start
- Promoter
- TATA box, CAAT box, & GC rich regions
DNA Sequence that Specifies End
- Terminator
- Polyadenylation
Typical “Layout of a Gene”
- Enhancer elements far upstream of start site
- Gene-specific (regulatory elements)
- CAAT or GC rich region
- TATA box
- Transcription start point
Alpha-Amanitin
- Toxin found in the “Death Cap” Mushroom
- inhibits Pol II & blocks synthesis of mRNA
- Results in liver failure
How does Rifampicin kill bacteria and not harm human cells?
- Potent antibiotic
- Inhibits bacterial RNA polymerase but not human Pol II
Capping
- 5’ methylguanosine cap is put on pre-mRNA
- Distinguishes mRNA from other forms of RNA
Exon
Gene region that codes for protein
Intron
Gene region that is non-coding & spliced out
pre-mRNA splicing
- Removing introns from pre-mRNA
- Spliceosomes: recognize boundaries, cleave, & rejoin
Outline the Process of Splicing
- Adenine residue attacks the 5’ splice site & cuts backbone
- 5’ end becomes covalently attached to adenine, forming a loop
- Newly generated free 3’ end reacts with the next 5’ end to join the exons together
- Lariat intron structure is released
How are the boundaries of introns and exons defined?
- 5’ splice site
- 3’ splice site
- branch point in middle
- AGG consensus sequence
Alternative Splicing
- Splicing different combinations of exons together to generate different proteins
Polyadenylation
- Tail of roughly 200 adenine nucleotides added to mRNA
- unique to mRNA
B-Thalassemia & Splice Errors
- Incorrect splicing leads to reduced synthesis of the B-chain of Hb
- Less B chain= less Hb & profound anemia
PKU & Splice Errors
- Lack of phenylalanine hydroxylase activity results in an inability to convert phenylalanine to tyrosine
- Arises from a single base change in 5’ splice site of phenylalanine hydroxylase gene
HAT
- Histone acetyltranferases= acetylate lysine residues in histones
- decreases net positive charge of histone & loosens interaction with negatively charged DNA
- ACTIVATE transcription
HDAC
- Histone deacetylases
- Remove acetyl grouops
- INHIBIT transcription
DNA Methylation
- DNA can be methylated on cytosine
- Attracts HDACs
- INHIBITS transcription
Helix-turn-Helix Protein
- Alpha helix, short amino acid chain, alpha-helix
- Side chains close to the C-terminal of helices binds DNA
- E.g. Homeobox proteins
Zinc Finger Protein
- Contain zinc as a structural element
- Alpha-helices bind DNA
- E.g. glucocorticoid receptor
Leucine Zipper Proteins
- Form dimers with two alpha-helices joined to form a coiled coil
- E.g. Fos & Jun
How do DNA-binding proteins regulate transcription?
- Act as nucleation sites for more proteins
- Recruit HATs, HDACs or chromatin remodeling complexes to DNA
How is transcription of the LDL-receptor gene enhanced in response to low cholesterol?
- Promoter region for LDL receptor gene contains SRE-1 (sterol responsive element-1)
- low cholesterol, SREBP-1a (sterol responsive element binding protein-1a) enter the nucleus and binds SRE-1
- Recurits HAT
- Chromatin relaxes and transcription of LDL-receptor increased
How does cortisol regulate transcription?
See Diagram
How do the thyroid hormone receptor and retinoid X receptor (RXR) act together to regulate transcription in response to thyroid hormomne?
See Diagram
What are the roles of CREB & CBP in protein kinase A mediated transcriptional control?
See Diagram
Rubenstein-Taybi Syndrome
- Intellectual Disability
- Caused by mutations in CBP or EP300
Tamoxifen
- Competitive inhibitor of the estrogen receptor
- Tamoxifen + ER bind DNA but don’t recruit proteins to activate transcription
Why is it acceptable for RNA polymerase to be much more error prone than DNA polymerase?
RNA is not the permanent repository of genetic information; DNA is
Pol I
- rRNA (ribosomal)
- 28S, 18S, 5.8S, all come from one large rRNA
- NOT 5S rRNA
Pol III
- tRNA(adapter between mRNA & amino acid)
- 5S rRNA
Pol II
mRNA (messenger)
Consensus sequence
Similar sequences of DNA found between different genes examined (promoters & terminators)
General Transcription Factors
- Required by Pol II to start transcription
- TFIID Complex (Transcription Factor for Pol II)
TBP
- Major component of TFIID complex
- Directs TFIIE, F, H & Pol II to promoter
Transcriptional Initiation Complex/ Basal Transcription Complex
TFIID, E, F, H & Pol II assembled at the promoter
Why are promoter regions rich in AT bases?
Easier to pull apart b/c they only contain two hydrogen bonds
What catalyzes the separation of DNA for transcription?
DNA Helicase activity of TFIIH
Additional transcription factors for high rate transcription
SP-1 & NF 1
SP-1
- high rate transcription factor
- binds GC-rich sequences
NF1
- high rate transcription factor
- binds CAAT box
mRNA degradation
1) shortening of the poly-A tail
2) removal of the 5’ cap (triggered by shortening of tail)
3) Degradation by ribonucleases
Chromatin Remodeling Complexes
Use the energy of ATP hydrolysis to change the structure of nucleosomes so that DNA is less tightly bound
Where do regulatory proteins typically bind DNA & why?
- Major Groove
- Where the exposed portion of the bases is located
