Eukaryotic Chromosome Structure and Function Flashcards Preview

Genes To Genomes > Eukaryotic Chromosome Structure and Function > Flashcards

Flashcards in Eukaryotic Chromosome Structure and Function Deck (114)
Loading flashcards...

What does a nucleosome consist of?

DNA and histone proteins


What are the four different histones that make up the octameric protein core of a nucleosome?

H2A, H2B, H3 and H4


Describe the general structure of a histone protein.

A three alpha-helix "histone fold" and unstructured TAIL regions


The histone protein core is modular - how is this made up?

Two molecules each of H3 and H4 form a tetramer; H2A and H2B molecules form two dimers


During regulation of transcription, eukaryotic chromatin-remodelling co-activators/repressors act at two functionally distinct levels. What are these two levels?

1) Local-level remodelling at gene-regulatory regions (promoter/UPE, enhancers etc.)
2) Domain-level remodelling over large chromosomal regions


What is the major function of co-activator/repressor complexes at gene-regulatory regions?

To manipulate individual nucleosomes to allow TF and RNA pol II access to DNA


What two ways can co-activators and repressors remodel chromatin?

1) Some use energy from ATP hydrolysis to modify nucleosomes
2) Some covalently modify histone protein amino acid residues within nucleosomes

(They are enzymes)


What two ways do co-activators and co-repressors manipulate nucleosomes?

1) Nucleosome positioning
2) Nucleosome structure

(Know this because of chromatin-seq/MNase-seq, a modification of DNase-seq)


What is the key reagent involved in Chromatin-seq/MNase-seq?

Micrococcal nuclease (MNase)


What are the key steps involved in Chromatin-seq/MNase-seq?

1) Living cells + detergent + micrococcal nuclease --> MNase digestion of chromosomes in vivo = fragments of DNA wrapped around a nucleosome protected from digestion
2) The around 150 bp of MNase-resistant DNA fragments from the cells are purified and sequences


Chromatin-seq: what does a peak in sequence read frequency tell you?

A peak = positioned nucleosomes. Chromatin-Seq experiments reveal that regions of genome have nucleosomes accurately positioned relative to underlying DNA sequence according to cell type/condition.


What is a pioneer TF and give some examples?

A pioneer TF, e.g. Oct3/4 and FoxA, is able to bind nucleosome-associated motifs


What is an NFR and when do they form?

Nucleosome-free regions: when a gene is activated, gene-regulatory DNA and promoters become exposed as NFRs to allow access for TF binding


How can we map transcription factors?

Using chromatin-seq: MNase will also not cleave DNA when the DNA is bound to TFs, leaving 20-50 bp of undigested DNA which we can sequence


What drives the creation of NFRs?

The recruitment of chromatin-remodelling ATPase co-activators, energy from ATP hydrolysis drives non-covalent changes in nucleosome structure


What are chromatin-remodelling ATPases?

Complexes that contain a core ATPase sub-unit and other proteins that modulate and target their activity (we have at least 5 classes of chromatin-remodelling ATPase complexes which differ in the structure of the ATPase sub-unit and in the type of nucleosome remodelling events they catalyse)


How does the chromatin-remodelling ATPase SWI/SNF act?

1) SWI/SNF ATPases can evict/displace histones from the DNA to remove nucleosomes from underlying sequence
2) They can also SLIDE histone octamers relative to the underlying DNA to alter the position of nucleosomes during gene activation


What are histone chaperones?

Histone chaperones (e.g. ASF1) are used to carry histones that are evicted during chromatin remodelling and during nucleosome assembly/dis-assembly during DNA replication/transcript elongation.


What effect can SWI/SNF ATPses, such as RSC, have on nucleosome structure?

SWI/SNF ATPases such as RSC can destabilise the overall structure of nucleosomes to make the wrapped DNA more generally accessible to DNA-binding proteins


What are two of the main ways to modify nucleosome structure?

1) Incorporating histone variants e.g. H2A.Z; H3.3, H2ABbd
2) Many chromatin remodelling co-activators/repressors (and some TFs) catalyse addition or removal of specific histone PTMs) - amino acids within the histone tails and cores are subject to more than 60 distinct, mostly reversible, PTMs


Give an example of an ATPase that can exchange a variant histone into nucleosomes.

SWR/INO80 ATPase can exchange variant histone H2A.Z into nucleosomes


Where is the variant histone H2A.Z often found?

Flanking NFR associated with yeast and human promoters - H2A.Z-containing nucleosomes may be more accessible to TFs and RNA pol II


What are HAT co-activators, give an example and explain what do they do.

Histone acetyltransferase (HAT) co-activators, e.g. SAGA, add acetyl groups (from acetyl coA) to multiple lysine residues in the N-terminal tails of histones H2B, H3 and H4 to make them more accessible to TF binding and PIC function.


What are HDAC co-repressors, give an example and explain what do they do.

Histone deacetylase (HDAC) co-repressors, e.g. RPD3, remove acetyl groups from lysine residues in the N-terminal tails of histones.


As well as changing the structure of nucleosomes, histone PTMs also..

...create binding sites for other proteins (often more co-activator/repressor complexes), writing an "epigenetic" code (e.g. bromodomains recognise acetyl-lysines; chromo- and PHD domains recognise methylate lysines)


What two histone PTM marks are generally associated with nucleosomes in non-transcribed/"repressed" regions of the genome?

H3K9me2/3 and H4K20me3


i) What histone PTM mark is generally associated with non-activated/"off' state gene regulatory regions and ii) what other active mark is sometimes found with it?

i) H3K27me3.
ii) The "active" PTM H3K4me3 is sometimes found with H3K27me3 at gene-regulatory regions "poised" between "on" and "off"


Give some examples of PTMs generally associated with activated/"on" state gene regulatory region nucleosomes.

H3K9ac, H3K27ac and H3K4me2/3


How do local chromatin remodelling events at widely-spaced higher eukaryotic gene-regulatory regions communicate?

Activated eukaryotic gene-regulatory regions are often separated in sequence in the nucleus but not in space, they interact with each other


How do we know that activated eukaryotic gene-regulatory regions interact with each other?

Chromosome conformation capture (3C) technologies which use NGS to map interactions between DNA regions