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MCB - Topic 2 > Chromosomes and Replication > Flashcards

Chromosomes and Replication Flashcards

1
Q

Why is it important for DNA to be packaged into chromatin?

A

Chromatin compaction = correlates with expression, and is related to accessibility of transcriptional machinery to the DNA.
DNA compaction level = relevant to DNA recombination and repair.

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2
Q

What is the difference between Heterochromatin and euchromatin?

A

Heterochromatin = closed, inactive marks on the chromatin.
Euchromatin = open, active marks on the chromatin.

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3
Q

What is the nucleosome?

A
  • 146 bp of DNA wrapped around a histone octamer (2 x H2A, 2 x H2B, 2 x HB, 2 x H4) - linked by an exterior H1.
  • Histones = +vely charged (lys and arg rich).
  • +vely charged histones bind to -vely charged DNA.
  • N-ter histone tails protrude from the octamer.
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4
Q

Explain post-translational histone modifications.

A
  • Modifications occur on the N-ter tails that protrude from the nucleosome, accessible to other chromatin proteins.
  • Different histone tail modification = associated with different functions (transcriptional activation, silencing, replication, repair, etc).
  • Providing opportunity to alter chromatin structure by chemically altering the histone/providing docking sites for binding partners.
    More than 50 sites can be modified, some with more than 1 type of tag.
    Acetyl or methyl groups = added to histone tails due to accessibility.
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5
Q

Explain what histone acetylation is and what occurs.

A
  • Acetylation = correlated with gene activity, partly due to reduced positive charge of histones.
  • Acetylation (especially for H3 and H4) = neutralises the charge and loosens the chromatin structure.
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6
Q

Explain what histone methylation is.

A
  • does not alter the charge of the histone.
  • mono, di, and tri-methylation exist (mostly referring to tri-methylation).
  • correlate with either transcriptional activity or inactivity - depending on which histone tail residue is methylated.
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7
Q

Explain the difference between H3K4me, H3K9me, and H3K27me.

A
  • H3K4me – active locus around the promoter.
  • H3K9me – inactive locus spread over the gene, constitutive heterochromatin.
  • H3K27me – inactive locus spread over the gene, facultative heterochromatin.
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8
Q

What are the three parts of a mitotic chromosome.

A
  1. Origins of replication - regions where DNA replication is initiated.
  2. Centromere - point of attachment of microtubules during mitosis.
  3. Telomeres - a protective cap on the ends of linear chromosomes (must be maintained/extended after replication).
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9
Q

What are the four phases of the cell cycle?

A
  1. Interphase.
  2. S-phase.
  3. M phase.
  4. Interphase.
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10
Q

Explain what occurs in interphase (pt 1).

A

Single copy of the chromosomes are unfolded and loose, allowing for transcription to take place.

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11
Q

Explain what occurs in S phase.

A

Copy of the chromatids, packaged up in preparation for mitosis.

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12
Q

Explain what occurs in M phase.

A

Separation of the two copies of DNA.

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13
Q

Explain what occurs in interphase (pt 2).

A

chromosomes unpack again.

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14
Q

What is the Origin of Replication (DNA replication)?

A

Location of a chromosome where DNA replication is initiated.
Eukaryotes have multiple origins of replication on each linear chromosome.

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15
Q

What is the function/feature of the origin of replication?

A
  • AT-rich (easier to separate the two DNA strands).
  • Enriched for H4K20me2 (binding site for ORC) = flag for the origin of replication.
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16
Q

Explain what occurs in the initiation of DNA rep in Eukaryotes.

A
  1. ORC (Origin binding complex) binds to the origin.
  2. DNA helicase binds to complete the pre-replicative complex.
  3. Wait for S-phase to begin (signaled by an activation of s-cdk).
  4. S-cdk phosphorylates many targets associated with DNA synthesis (deactivates the ORC and activates helicase).
  5. Helicase separates the AT-rich area.
    DNA polymerase is loaded and the replication forks head off in opposite directions.
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17
Q

Explain how replication occurs at the end of the chromosome (for the leading strand).

A

Leading strand = can be replicated right to the end (from 3’ to 5’).

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18
Q

Explain how replication occurs at the end of the chromosome (lagging strand).

A
  • Lagging strand = last part of the chromosome (where the final RNA primer was placed) cannot be replicated with DNA (exonucleases will remove this single stranded overhang).
  • Therefore, after each cell cycle, chromosomes lose 50-100 bp. Normal process in somatic cells of the body, which will typically only divide 50-70 before senescing (ceasing division).
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19
Q

What opposes the loss of bp from the lagging stran?

A

Telomeres and telomerase.

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20
Q

What are telomeres?

A

Protective structure at both ends of a linear chromosome.
Consists of a tandem repeat of the sequence GGGTTA, and associated proteins which serves to maintain the integrity of chromosomes.

21
Q

What is the function of telomeres?

A

Telomeres protect the ends of chromosomes from the loss of bp.
Chromosomes lacking telomers = unstable and may be susceptible to chromosomal rearrangements (e.g. fusion).

22
Q

What is telomerase?

A

Ribonucleoprotein (protein + RNA) that elongates telomere sequences.

23
Q

What are the two key features of telomerase?

A
  1. RNA portion of telomerase = sequence CUAACCCUAAC which is complementary to the telomere tandem repeat (TTAGGG).
  2. Telomerase = has protein component (TERT) which has reverse transcriptase activity. Making a DNA copy from the RNA portion of the telomerase enzyme.
24
Q

Explain the process of telomere extension.

A
  1. Start with the 3’ end of the overhanging strand.
  2. Telomerase binds to the 3’ end of the telomere that is complementary to the telomerase RNA to extend the overhanging strand.
  3. The telomerase relocates to continue this process.
  4. DNA polymerase then complements the lagging strand.
25
Q

How do the loss of telomeres/telomerase lead to aging?

A
  • Loss of telomeres = causes cellular senescence.
  • Telomere length correlates with age (shorter = older).
  • Werner syndrome = mutation in the WRN (protein involved in the telomere CAP structure) which causes shorter telomeres and premature aging.
26
Q

What is a centromere?

A
  • A heterochromatic region of a chromosome that joins sister chromatids during mitosis.
  • Region to which microtubules attach (via the kinetochore).
  • Pericentric regions flank the centric region. Pericentric regions = often rich in LINEs and SINEs. The centric region of the centromere consists of a tandem repeat of a 171 bp sequence called the alpha-satellite repeat. Microtubules bind to the centromere via the kinetochore.
27
Q

What are the four types of chromosomal aberrations?

A
  1. Gain or loss of chromosomes.
  2. Duplications/deletions of parts of chromosomes.
  3. Chromosomal fusions (two homologous chromosomes fuse together, especially if there is a loss of telomeres).
  4. Reciprocal translocations (a section switches with another chromosome) – often not an immediate problem for the cell since the same genes are there.
28
Q

What is the relationship between cancer and chromosomal instability?

A

Defining characteristic of cancer = chromosomal instability. A departure from the usual characteristics of a set of chromosomes.

29
Q

What is down syndrome?

A
  • Physical growth delays
  • Characteristic facial features
  • Mild/moderate intellectual disability
  • Occurs in 1/1000 birth and is caused by a 3rd copy of chromosome 21.
30
Q

Why does Down Syndrome Occur?

A
  • Most cases = due to meiotic non-disjunction.
  • 2-3% are due to a Robertsonian Translocation.
31
Q

What is an acrocentric chromosome?

A
  • A chromosome in which the centromere is located close to the end of the chromosome.
32
Q

How many acrocentric chromosomes exist?

A
  • 5 (13, 14, 15, 21, 22)
33
Q

What is a Robertsonian Translocation?

A
  • Two chromosomes joining at the centromere region.
  • Occur in 1/1000 births and are most common between chromosomes 13 and 14.
  • Short p arms of the acrocentric chromosomes 13, 14, 15, 21, 22 = do not contain any essential genes but do contain tRNA and rRNA which are present in multiple copies.
  • These genes = tend to be present at multiple points throughout the genome. As a consequence, if you lose these parts of the chromosome = NOT catastrophic since they don’t contain essential genes.
  • BUT, if there is a chromosomal translocation between the two acrocentric chromosomes = will lose the rRNA and tRNA genes.
34
Q

What is the ‘central dogma’?

A

Information flow from DNA to RNA to amino acid sequence.

35
Q

What is transcription?

A

Copying of one strand of DNA into a complementary RNA sequence by the
enzyme RNA polymerase.

36
Q

What is RNA polymerase?

A

Enzyme that catalyses the synthesis of an RNA molecule on a DNA template from ribonucleoside triphosphate precursors (come in as triphosphates and then they are cleaved).

37
Q

Explain RNA polymerase in Prokaryotes.

A
  • Prokaryotes = have a single RNA pol.
  • Core enzyme = responsible for polymerization and is composed of 5 subunits.
  • Sigma factor is only needed to initiate transcription .
  • When those two sections come together = known as HOLOENZYME.
38
Q

Explain RNA pol in eukaryotes.

A
  • Eukaryotes = have 3 types of RNA.
  • RNA pol 1: ribosomal RNA
  • RNA pol 2: messenger RNA to protein
  • RNA pol 3: transfer RNA
  • Euk RNA pol = have subunits homologous to the prokaryotic RNA pol and have many subunits in common.
39
Q

Explain the process of prokaryotic transcription initiation.

A
  1. The sigma factor binds to binding sites in the promotor region of the
    DNA.
  2. It then recruits the rest of the core enzyme
  3. The core enzyme is able to separate the double stranded DNA
  4. And then begin transcription
40
Q

Explain how transcription initiation for RNA pol 2 occurs.

A
  • Promoter is more variable and complex than in prokaryotes.
  • The initiation complex involves many more proteins/factors.
  • General Transcription Factors: bound to enhancers
  • RNA polymerase 2
  • The mediator complexes
  • Transcriptional activators
  • Chromatin remodelling complexes
  • Histone-modifying enzymes
41
Q

What is the purpose of general transcription factors?

A

Needed to initiate transcription of all genes.

42
Q

What is the GTF for RNA polymerase 2?

A

TFII

43
Q

What is the key recognition region called in eukaryotes? Why is this important?

A

TATA box. AT-rich.

44
Q

Explain the role of the TATA box in allowing transcription to begin. How does this occur?

A
  1. Transcription begins when the TFIID binds to the TATA box via the TBP component (TATA-binding protein). The TFIID complex also has 11 other
    proteins called TAFs (TBP-Associated Factor)
  2. Next, other GTFs (such as TFIIB) and RNA polymerase 2 bind and finally
    there is a pre-initiation complex
  3. Finally, TFIIH activates transcription with its helicase activity, unwinding
    the DNA at the TATA box, and then with its kinase activity it
    phosphorylates the CTD tail at Ser-5 of the RNA polymerase
  4. RNA polymerase 2 is now able to start transcription.
45
Q

What are the two subunits that make up the TFIID? What are their functions?

A
  1. TBP subunit = 1 subunit, recognizes TATA box.
  2. TAF subunit = 11 subunits, recognizes other DNA sequences near the transcription start point; regulates DNA-binding by TBP.
46
Q

What is the C-terminal domain?

A

The CTD = key regulatory region of RNA pol 2.
Consists of 52 tandem repeats of a 7 amino acid motif with ser at positions 2 and 5.

47
Q

Explain why the C-ter domain is important?

A

Phosphorylation of Ser5 affects the conformation and activity of RNA
polymerase 2, releasing it to begin transcription.

48
Q
A

MCB - Topic 2 (5 decks)

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