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BHCM218 > Module 03 - Section 07 > Flashcards

Module 03 - Section 07 Flashcards

Chromosome Organization and Packaging

1
Q

What are the 3 characteristics of chromosome packaging?

A

(1) Highly organized
(2) Allow access to factors that regulate DNA replication
(3) Allow access to factors that regulate transcription

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2
Q

What are the 6 levels of organization of a chromosome? (smallest to largest)

A

(1) nucleotides
(2) DNA double helix
(3) histones
(4) nucleosomes
(5) Chromatin
(6) Mitotic chromosome

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3
Q

What are histones?

A

Family of basic proteins (+ charged) that associate tightly with DNA in the chromosomes of all eukaryotic cells and help condense DNA

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4
Q

What is chromatin?

A

Filamentous complex of DNA, histones, and other proteins constituting the eukaryotic chromosome

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5
Q

What is the nucleosome?

A

Structural unit for packaging chromatin in eukaryotes, consisting of a DNA strand wound around a histone core

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6
Q

What is the largest protein component of chromatin?

A

Histones

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7
Q

How do histones arrange?

A

Into octamers, each octamer unit contains two copies of the four different histone subunits

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8
Q

What is the first level of Chromosome packaging?

A

Formation of Nucleosome: DNA is wrapped twice around the histone octamers. (positive charge of histone interacts with negative charge of DNA backbone through electrostatic interactions)

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9
Q

What is chemical crosslinking?

A

A chemical with 2 reactive compounds (eg: formaldehyde) is reacted with a protein complex. Because it has 2 reactive groups it can covalently bond with 2 proteins, effectively linking them. The chemical is a small molecule, so it can only react with proteins that are close together.

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10
Q

What does chemical crosslinking reveal?

A

Which proteins are next to each other in an oligomer

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11
Q

What did Kornberg’s chemical crosslinking experiment reveal?

A

H3 and H4 form a heterotetramer - this was brought forth when analyzed with SDS-PAGE

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12
Q

What are the 4 core histones?

A

H2A, H2B, H3 and H4

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13
Q

What are the 2 tetramer that forms the histones octamer?

A

H2A-H2B x 2 (first tetramer)

H3-H4 x2 (2nd tetramer)

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14
Q

What are the functions of H1?

A

(1) protecting the linker DNA from degradation
(2) Provides further compaction 6-7 times more than nucleosome
(3) Stabilizing nucleosomes
(4) enhancing the repression of transcription by nucleosomes
(5) promoting higher order chromosome structure

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15
Q

How many BPs are wrapped around the histones? between the nucleosomes?

A

(a) 146 bp

(b) There is 200bp between nucleosome which means there is about 50bp of linker DNA

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16
Q

What amino acids are histones rich in?

A

Basic amino acids arginine and lysine (25% of histone proteins together)

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17
Q

Which histones are nearly identical in all eukaryotes? (2)

A

H3 & H4

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18
Q

What shape does the histone + DNA structure take?

A

Solenoidal supercoil

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19
Q

What is a solenoidal supercoil?

A

An over-wound DNA strand, forming a tightly packed helical structure

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20
Q

How much compaction does the nucleosome structure provide DNA?

A

6 to 7 fold

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21
Q

What is the characteristic histone fold motif?

A

Globular domain that consists of three alpha-helices linked by two short loops

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22
Q

What does the histone motif allow for?

A

Histone proteins to bind together and form a dimer (H3-H4 or H2A-H2B) Which ultimately allows for the tight wrapping of DNA around the histone core to form a nucleosome

23
Q

What is the basic structural unit of the nucleosome?

A

Head-to-tail dimer of histone-fold motifs

24
Q

Describe the Histone fold dimer structure

A

They form a V-shaped structure that contains three DNA-binding sites.

25
Q

Where does the nucleosome interact with DNA?

A

Via the Minor groove, at all 3 DNA-binding sites

26
Q

What is the basic unit of DNA compaction?

A

nucleosome

27
Q

What do nucleosomes look like?

A

Beads on a string

28
Q

What can influence the ability of DNA to bind the nucleosome?

A

the presence of consecutive A=T or G=_C (3 links) base pairs

29
Q

What effects does a local abundance of A=T base pairs in the minor groove have on the nucleosome?

A

It is in contact with the histones and it facilitates the compression of the minor groove that is needed for tight wrapping of DNA around the histone octamer. Histones assemble particularly well with sequences where 2 or more A=T base pairs are staggered at 10bp intervals

30
Q

What effects does a local abundance of G=_C (3 links) base pairs in the the minor groove have on the nucleosome?

A

Opposite of A=T base pairs, they prevent compression of the minor groove and thus are preferred at position not facing nucleosome

31
Q

What is a 30nm Filament?

A

When the nucleosome further condense into a compact filament with a width of 30nm

32
Q

What are the 2 binding sites of H1?

A

One binds the arm of linker DNA

One bind to the central region of the DNA strand in the nucleosome

33
Q

What is the structure called when H1 is bound to the histone octamer and DNA?

A

Chromatosome

34
Q

How is the binding of the histone H1 to the DNA different from the core histones?

A

Histone H1 bind to one strand of the linker DNA as it comes off the nucleosome and binds a second site in the central region of the DNA helix wrapped around the octamer. The core histones interact with the wrapped DNA in the minor groove

35
Q

What is believed to be involved in higher-order levels of organization?

A

Further coiling and loops

36
Q

Describe the experiment that provided the evidence that suggests DNA is packaged into regularly organized units (by Roger Kornberg)

A

(1) Chromosomal DNA was treated with a nonspecific DNA nuclease called micrococcal nuclease that cuts DNA wherever it is NOT associated with proteins - fragments were separated by size on agarose gel

37
Q

What were the two hypotheses of the experiment that provided the evidence that suggests DNA is packaged into regularly organized units (by Roger Kornberg)

A

(1) if DNA is packaged by proteins into units of a particular size, the nuclease would cleave only the DNA between these units, and the protected DNA segments would migrate in the gel as a ladder of unit-sized band
(2) If there is no regular repeating unit of protein-DNA packaging and proteins were distributed on DNA randomly, then nuclease digestion would produce a smear of DNA fargments with no regular pattern

38
Q

What were the results of the experiment that provided the evidence that suggests DNA is packaged into regularly organized units (by Roger Kornberg)

A

Series of regularly spaced DNA bands about 200 bp apart. Protected DNA segments migrated on the gel as a ladder of regularly-spaced bands

39
Q

If DNA is packed into units of 200 bp, then why are there bands representing lengths longer than this visible when the digested DNA is run on agarose gel?

A

Nuclease does nut cute the DNA at every single possible spot, thus the bands appear as 200 bp or multiples of 200bp depending on the number of nucleosomes in between each cut

40
Q

Why are nucleosomes complexes weakly associated?

A

to facilitate access. There must be a way for the chromatin structure within the promoter region to become accessible

41
Q

What does the fact that interactions holding 30nm filament and nucleosomes together are weak imply?

A

Regions within nucleosome arrays will be dynamic, with hydrogen bonds between the histones and DNA breaking and re-forming quite easily, and allowing access to chromatin for DNA replication and transcription

42
Q

What is chromatin composed of?

A

DNA and Protein

43
Q

What are the 3 considerations that are cirtical to the effectiveness of DNA compaction?

A

(1) Dynamic
(2) Modifiable
(3) Responsive

44
Q

What do we mean by DNA compaction must be dynamic?

A

Changes in the degree of condensation must occur quickly and when needed, as the cell passes through the stages of the cell cycle

45
Q

What do we mean by DNA compaction must be modifiable?

A

DNA compaction must be globally and locally modifiable. Global refers to modifications for processes like mitosis or replication, while local means giving access to specific genes for transcriptional regulation

46
Q

What do we mean by DNA compaction must be responsive?

A

Must be able to respond to modification enzymes that are able to alter the state of DNA condensation. Enzymes can target specific regions for transcription or replication – these regions must be recognizable

47
Q

Which part of the Histone enable organization and compaction of DNA?

A

N-terminal Tails

48
Q

What are N-terminal tails?

A

part of the histone subunits that protrude out from the core particle and are less ordered. They are flexible and therefore, mostly disordered

49
Q

Describe the structure of N-Terminal tails

A

Parts of the histone tails that are visible in the crystal structure appear as irregular chain extending outward from the nucleosome disk. The tails exit the DNA superhelix through channels formed by the alignment of minor grooves of adjacent DNA helices every 20 bp.

50
Q

What is the contribution to DNA binding by the N-terminal tails

A

Do not contribute much strength to DNA binding, but they form intermolecular contacts with adjacent nucleosome particles and organize nucleosomes into a higher-order chromatin structure

51
Q

Is H1 essential to the formation of 30nm filament?

A

No

52
Q

Is the N-terminal tails of the core histones essential to the formation of 30nm filament?

A

Yes

53
Q

Why are N-terminal tails essential to the formation of 30nm filament?

A

Tails provide important nucleosome-nucleosome contacts needed for 30 nm filament formation

54
Q

What regulates the accessibility of DNA?

A

internucleosome connections mediated by histone tails

BHCM218 (24 decks)

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