Lecture 2: Gene Expression: Transcription Flashcards
DNA directed RNA synthesis from a DNA template
by a DNA-dependent RNA polymerase
Transcription
starting point of gene expression
transcription
Eukaryotic cell
– DNA to message is nuclear
– protein synthesis is ___________
_______ mechanism is necessary
cytoplasmic
- Transport mechanism is necessary
- mRNA translocate from nucleus to cytoplasm
Structure and Properties of RNA:
* Unbranched polymer of nucleotide 5’ monophosphates linked by 3’-5’ ____________ bonds
phosphodiester
RNA is generally ______-stranded (ss) except in some viruses.
single
Structure and function of RNA:
Can fold upon itself to form ____________ regions. The filded 3D structure of RNA, like the folded proteins, can have multiple biological
functions.
intrastrand helical
Different Genes for Different RNAs include…
Types of RNA
- Pre-RNA
- mRNA
- tRNA
- rRNA
- snRNA
- snoRNA
- miRNA
snoRNAs and miRNAs are extensively characterized small __________ regulatory
RNAs.
non-coding
snoRNAs and miRNAs have distinct and central ____________ roles in cells.
regulatory
-recent studies reveal that snoRNAs and miRNAs display similarities at numerous levels.
- molecules showing both snoRNA and miRNA characteristics and functionality exist.
- subsets of snoRNAs and miRNAs likely share a functional and ____________
relationship.
evolutionary
Ribosomal RNA (rRNA) ___%
Transfer RNA (tRNA)___%
Messenger RNA (mRNA) __%
80
15
5
Secondary structure of RNA?
Tertiary structure of RNA?
stem loop(double-helical stem region) and hairpin
folding —-> pseudoknot
The ability of RNA to form unique _____ structures that act similarly to proteins is one of the key contributor that functional RNA can be involved in a wide range of cellular processes.
3D
RNA is synthesized on a ______ template by RNA polymerase.
DNA
NMPn + ribonucleoside triphosphate ———RNA polymerase/Mg2+
DNA template—–>
NMP n+1 + PPi
DNA to RNA is _____ polymerase
RNA polymerase
RNA to RNA by ___________
RNA replicase
RNA to DNA by ____________
reverse transcriptase
Prokaryotes:
___ polymerase / ___ subunits
1, 6
a2, b, b’, sigma, omega (alpha-2,beta,beta prime, sigma, omega)
________- subunit recognizes the promotor region of DNA that precedes the structural gene
Sigma (s)
______ subunit binds to promotor region, may serves to control the
efficiency of RNA synthesis.
Omega
But the overall function of omega (w) subunit is not
very clear
Rifampin- inhibits ____ subunit of polymerase
beta
Steps in Transcription:
RNA polymerase binds to promotor region (via specific _____ factor)
sigma
promotor has specific sequences
Steps in Transcription:
DNA coding strand = ______ strand
Non-coding Strand (Template) = _______ strand
sense
anti sense
Steps in Transcription:
Anti-sense is the one used in the _____ direction so RNA
chain is synthesized _____ and makes a coding strand
3’-5’
5’-3’
Steps in Transcription:
Coding strand has same sequence as message, except…
U instead of T
Steps in Prokaryotic Transcription?
- initiation
-elongation
-termination
Steps in Prokaryotic Transcription:
15-20bp bubble formed and only 1 strand is read
Initiation
Steps in Prokaryotic Transcription:
Starts with a purine, then continues
Elongation
Steps in Prokaryotic Transcription:
– Rho independent (intrinsic stem-loop)
– Rho dependent
Termination
Eukaryotic Transcription:
Polymerase ___-nucleolus-synthesizes 45s rRNA
1
Eukaryotic Transcription:
Polymerase ___-nucleoplasm, synthesizes mRNA, and others
II
-recognizes promotor sequence
Eukaryotic Transcription:
Polymerase ____-nucleus, synthesizes tRNA, 5sRNA, miRNA
III
________________ polymerase-synthesizes mitochondrial RNA
Mitochondrial
B-thalasemmia involves a mutation of…
TATA before b-hemoglobin gene
Affects rate of transcription and rate of protein production
Eukaryotic Promotor
What increases the rate and frequency of eukaryotic transcription?
-genes have many regions upstream that have sites for additional elements to
bind
-affects tissue specific expression
-enhancer
-Steroid hormone receptors binding
-Cell signaling factors (100-5000 bp upstream)
site of binding of regulatory proteins such as transcription factors and
hormone receptors
Enhancer
(site of binding of regulatory proteins that stop transcription
Suppressor regions
Hormone binding –> __________ —> enhancer —> promotor —> structural gene
transcription factor
Purpose of Post Transcriptional Modifications to
Eukaryotic mRNA?
– Stability of mRNA
– Determines protein variation from a single message (splicing)
– Guide for the translocation of mature mRNA to cytoplasm
a large RNA-protein complex that catalyses the removal of introns from nuclear pre-mRNA
spliceosome
Types of processing for mRNA?
-5’ cap
-3’ poy-A tail
-splicing
- removal of introns
- alternative splicing
– tissue specific protein
expression( a-
tropomyosin, smooth vs
striated muscle)
Splicing
3’ poy-A tail:
many adenosines for stability
5’ cap:
7-methyl-guanosine
to 5’ end
Inhibitors of RNA Synthesis
-template binding
-polymerase binding
-alpha Amanitin (mushroom toxin)
-inhibitors of reverse transcription
examples of inhibitors of reverse transcription
– AZT (azidothymidine)
– ddI (dideoxyinosine)
a Amanitin (mushroom toxin)
– Inhibitors of euk. pol ___ and ___
II and III
example of polymerase binding
Rifampin (b subunit)
Example of Template binding (at transcription initiation complex to prevent elongation)
Type of substance
Actinomycin D
Human Diseases Related to mRNA Synthesis?
alpha and beta thalassemia
– A form of inherited autosomal recessive blood disorder characterized by abnormal
formation of hemoglobin.
– Also affect rate of globin synthesis
– US carrier rate= 7%; Mediterranean = 20%
Thalassemia
a Thalassemias
– production of the α globin chain is affected
– 4 alleles, mostly deletions
beta Thalassemias
Types of mutation effected
Affects production of what
What is destroyed
– production of the b globin chain is affected
– promotor mutations
– splicing mutations
– capping and tailing mutations
– nonsense and frame shift mutations
– dysfunctional protein is destroyed
