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Lecture 7 Slides Flashcards

0
Q

What does DNA polymerase require at chromosome ends

A

A primer

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1
Q

What is the main purpose of a telomere?

A

To stabilize chromosome ends

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2
Q

At what end of DNA does replication end? What is incomplete at end of replication?

A

End of replication of DNA is at 5’ end. At the end of replication, replication of lagging strand is incomplete.

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3
Q

Without telomeres, what would happen to 5’ end of DNA?

A

It would continue to shorten over successive rounds of DNA replication

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4
Q

Telomerase has homology to what?

A

Reverse transcriptase

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5
Q

How does telomerase work?

A

It synthesizes DNA strand (5’ to 3’) from RNA template that is part of the enzyme

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6
Q

What does DNA polymerase do after telomerase?

A

It elongates opposite strand

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7
Q

What is human telomere sequence?

A

T2AG3

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8
Q

Steps of telomerase function

A
  1. Telomerase binds to incomplete lagging strand at end of replication
  2. Binds to parental strand at 3’ and adds to it.
  3. Extends three end with RNA templates DNA synthesis (G strand)
  4. DNA polymerase completes lagging strand (C strand)
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9
Q

What is shelterin complex

A

Protects telomere DNA and recruits telomere at S phase

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10
Q

What does shelterin complex regulate

A

Formation of t-loop

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11
Q

Which shelterin complex proteins bind single strand DNA

A

TPP1 and POT1

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12
Q

Which shelterin complex proteins bind double strand DNA

A

RAP1, TIN2, TRF2, TRF1

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13
Q

What is the difference, at end of DNA replication,between parent and lagging strands?

A

3’ overhang

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14
Q

How is t loop formed

A

Overhang loops above parent strand and links into it

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15
Q

In what cells is telomere active

A

In germ cells and some stem cells

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16
Q

Where does telomere DNA gradually shorten? What does that do to its activity?

A

In somatic cells

Activity is low

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17
Q

What do humans with with mutation in one copy of telomerase gene display?

A

Premature aging

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18
Q

When do primary cultures of animal somatic cells cease to divide?

A

After ~50 cell cycles; telomerase shortens at each cycle

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19
Q

What is function of DNA surveillance system

A

Detects chromosome defects, inhibits cell cycle or promotes cell death

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20
Q

If telomerase is active in cells, do they proliferate

A

Yes, indefinitely

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21
Q

What allows transformed cell lines and cancer cells to divide indefinitely

A

Active telomerase

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22
Q

What enzyme is used to make rna from DNA?

A

Rna polymerase

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23
Q

Difference between ribose and deoxyribose

A

Second carbon in deoxyribose has H i/o OH

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24
Difference between uracil and thymine
Uracil carbon double bond has no methylated carbons, but thymine has one methyl attached to carbon
25
Two types of dna dependent rna polymerases
Multi-subunit polymerases and single-subunit polymerases
26
Bacteria multisubunit polymerase
-one polymerase synthesizes all RNAs alphaI, alphaII, beta, beta', + sigma -polymerase binds DNA via alpha subunits, sigma factors -beta subunit has polymerase activity -homology of all five bacterial proteins are found in archaea and eukaryotes
27
Archaea multisubunit polymerases
One polymerase; 11-13 subunits; | Closely related to eukaryotic polymerases
28
Eukaryotes multisubunit polymerases
Three polymerases; Pol I - 14 subunits; Pol II-12 subunits; Pol III - 17 subunits Pol IV and pol V - roles in plant siRNA function
29
Single subunit polymerases
Bacteriophage T7 RNA polymerase Mitochondrial RNA polymerases Some chloroplast RNA polymerases
30
Where can one find RNA dependent RNA polymerases
In some viruses (like polio virus) | In eukaryotes, play a role in mi-RNA and si-RNA pathways
31
A-amantin sensitivity of RNA polymerase I. Genes transcribed
Insensitive | 5.8S, 18S, rRNA genes
32
A-amanitin sensitivity of rna polymerase II? Genes transcribed.
Very sensitive | Protein coding genes, miRNA, siRNA genes
33
A-amanitin sensitivity of RNA polymerase III. Genes transcribed.
Intermediate sensitivity | TRNA genes, 5s rRNA genes, other small RNA genes
34
Which dna strand is used as template
Either. It depends on promoter orientation.
35
Can there be multiple rna polymerases transcribing
Yes
36
What controls rate of transcript accumulation
Rate of initiation
37
Steps in transcription cycle of RNA polymerase II
1. Pre-initiation complex (PIC) formation 2. Pol II recruitment 3. Transition to initiating pol II 4. Transition to elongating pol II
38
What happens during PIC formation?
Assembly of TFIID | TATA-box binding protein (TBP) + 13 other proteins at TATA box
39
What ahopens at pol II recruitment?
Assembly of additional general transcription factors and RNA pol II
40
Transition to initiating POl II
Helicase activity in TFIIH pries apart dna helix Protein kinase activity in TFIIH phosphorylates residues in c terminal domain (CTD). CTD is unique to POL II largest subunit; repeated heptapeptide: Tyr-Ser-pro-Ser-pro-Ser Binds RNA processing factors
41
What happens during transition to elongate pol II
Pol II is released from initiation factors, elongation begins
42
What does preinitiation complex in vivo include
General transcription factors Pol II Mediator- a structurally flexible of about 25 proteins that links transcription factors (activators or repressors) to TNA pol II Proteins for modifying chromatin structure
43
Steps in eukaryotes RNA processing
- DNA is in nucleus - DNA has introns and exons - rna is transcribed from transcription unit into primary rna transcript - primary rna transcript gets 5' capping, RNA splicing, 3'polyadenylation - becomes mRNA, which is then exported from nucleus - mRNA is then translated into protein outside of nucleus
44
Prokaryotic rna processing pathway
DNA is in cytosol ( no nucleus) Transcribed directly to mRNA Translated in cytosol
45
Processing enzymes carried by C-Terminal domain
Capping factors at 5' end of mRNA Splicing proteins 5' cap 3' end processing proteins
46
What makes up eukaryotic mRNA cap?
5'-to-5' triphosphate bridge | 7-methylguanosine
47
Difference between polycistronic and monocistronic mRNA?
Poly has coding for multiple proteins. | Mono has coding just for one protein
48
How is 5' end cap added
Phosphatase takes phosphate Guanyltransferase add guanosine to end Methyl transferase adds methyl group to base On so e caps, add methyl group to ribose
49
Purpose of intron splicing
Facilitates evolution of new proteins Provides opportunity for regulation of gene expression Alternative splicing allows numerous proteins from a single gene
50
Splicing step 1
1. U1 snRNP forms base pairs with the 5' splice junction | 2. BBP (branch point binding protein) and U2AF (U2 auxiliary factor) recognize the branch point site
51
Splicing step 2
U2 snRNP displaces BBP and U2AF and forms base pairs with branch point site consensus sequence.
52
Splicing step three
The U4/U6.U5 "triple" snRNP enters reaction. U4 and U6 snRNAs are held firmly together by base pair interactions. Subsequently, rearrangements break apart the U4/U6 base pairs, allowing U6 to displace U1 at the 59 splice junction. This creates the active site that catalyzes the first phosphoryl-transferase reaction.
53
Splicing step four
Additional RNA-RNA arrangements create the active site for the second phosphoryl-transferase reaction, which then completes the splice
54
Processing at 3' end of transcript
1. Cleavage and poly-A signals encoded in DNA 2. Cleavage stimulation factor (CSF) and cleavage and polyadenylation specificity factor (CPSF) are attached to RNA polymerase 3. RNA is cleaved 4. Poly-A polymerase enters. Rna polymerase eventually terminates 5. Poly-Abonding proteins enter process 6. Poly-A length is regulated
55
What is polyadenylation signal
AAUAAA
56
At what sequence is there cleavage?shat is added at cleavage?
CA. Add hydroxyl group.
57
Where does gene rich dna lie in nucleus
In euchromatic regions
58
FISH stands for
Fluorescein in situ hybridization
59
Where are alu sequences found with FISH
In gene rich dna
60
What does fluorophore link to in fish
Antibodies
61
Transcription factories
Contains 8-10 RNA POL II molecules
62
Nuclear export of pol II transcripts
"Export-ready DNA exits through nuclear pore complex into cytoplasm - on RNA there is a nuclear export receptor, hnRNP proteins, and SR proteins - 5' end exits first - in cytoplasm, there are initiation factors for protein synthesis (eIF4G and eIF4E) - factors binds to 5' end - prepare for translation

Cell Biology (35 decks)

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