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Cell Biology > Lecture 7 Slides > Flashcards

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
Q

Difference between uracil and thymine

A

Uracil carbon double bond has no methylated carbons, but thymine has one methyl attached to carbon

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25
Q

Two types of dna dependent rna polymerases

A

Multi-subunit polymerases and single-subunit polymerases

26
Q

Bacteria multisubunit polymerase

A

-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
Q

Archaea multisubunit polymerases

A

One polymerase; 11-13 subunits;

Closely related to eukaryotic polymerases

28
Q

Eukaryotes multisubunit polymerases

A

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
Q

Single subunit polymerases

A

Bacteriophage T7 RNA polymerase
Mitochondrial RNA polymerases
Some chloroplast RNA polymerases

30
Q

Where can one find RNA dependent RNA polymerases

A

In some viruses (like polio virus)

In eukaryotes, play a role in mi-RNA and si-RNA pathways

31
Q

A-amantin sensitivity of RNA polymerase I. Genes transcribed

A

Insensitive

5.8S, 18S, rRNA genes

32
Q

A-amanitin sensitivity of rna polymerase II? Genes transcribed.

A

Very sensitive

Protein coding genes, miRNA, siRNA genes

33
Q

A-amanitin sensitivity of RNA polymerase III. Genes transcribed.

A

Intermediate sensitivity

TRNA genes, 5s rRNA genes, other small RNA genes

34
Q

Which dna strand is used as template

A

Either. It depends on promoter orientation.

35
Q

Can there be multiple rna polymerases transcribing

A

Yes

36
Q

What controls rate of transcript accumulation

A

Rate of initiation

37
Q

Steps in transcription cycle of RNA polymerase II

A
  1. Pre-initiation complex (PIC) formation
  2. Pol II recruitment
  3. Transition to initiating pol II
  4. Transition to elongating pol II
38
Q

What happens during PIC formation?

A

Assembly of TFIID

TATA-box binding protein (TBP) + 13 other proteins at TATA box

39
Q

What ahopens at pol II recruitment?

A

Assembly of additional general transcription factors and RNA pol II

40
Q

Transition to initiating POl II

A

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
Q

What happens during transition to elongate pol II

A

Pol II is released from initiation factors, elongation begins

42
Q

What does preinitiation complex in vivo include

A

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
Q

Steps in eukaryotes RNA processing

A
  • 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
Q

Prokaryotic rna processing pathway

A

DNA is in cytosol ( no nucleus)
Transcribed directly to mRNA
Translated in cytosol

45
Q

Processing enzymes carried by C-Terminal domain

A

Capping factors at 5’ end of mRNA
Splicing proteins
5’ cap
3’ end processing proteins

46
Q

What makes up eukaryotic mRNA cap?

A

5’-to-5’ triphosphate bridge

7-methylguanosine

47
Q

Difference between polycistronic and monocistronic mRNA?

A

Poly has coding for multiple proteins.

Mono has coding just for one protein

48
Q

How is 5’ end cap added

A

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
Q

Purpose of intron splicing

A

Facilitates evolution of new proteins
Provides opportunity for regulation of gene expression
Alternative splicing allows numerous proteins from a single gene

50
Q

Splicing step 1

A
  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
Q

Splicing step 2

A

U2 snRNP displaces BBP and U2AF and forms base pairs with branch point site consensus sequence.

52
Q

Splicing step three

A

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
Q

Splicing step four

A

Additional RNA-RNA arrangements create the active site for the second phosphoryl-transferase reaction, which then completes the splice

54
Q

Processing at 3’ end of transcript

A
  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
Q

What is polyadenylation signal

A

AAUAAA

56
Q

At what sequence is there cleavage?shat is added at cleavage?

A

CA. Add hydroxyl group.

57
Q

Where does gene rich dna lie in nucleus

A

In euchromatic regions

58
Q

FISH stands for

A

Fluorescein in situ hybridization

59
Q

Where are alu sequences found with FISH

A

In gene rich dna

60
Q

What does fluorophore link to in fish

A

Antibodies

61
Q

Transcription factories

A

Contains 8-10 RNA POL II molecules

62
Q

Nuclear export of pol II transcripts

A

“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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