epigenetics- lecture 14 Flashcards

Question

DNA packaging

Answer 1

the process of how accessible the DNA is to transcriptional machinery DNA does not float freely inside the nucleus, but instead is wrapped around histone protein complexes and packaged into repeating units known as nucleosomes. Collectively, these nucleosomes are referred to as chromatin. Each individual nucleosome consists of eight histone proteins around which the DNA wraps 1.65 times. These nucleosomes then coil and fold around one another to produce a 30nm fiber, which subsequently undergoes progressive rounds of coiling and condensation culminating in an extremely condensed and ‘super coiled’ chromosome. Coiling the DNA in this fashion results in the genome being condensed so tightly that it can fit into the tiny nuclear space

Answer 2

the totality of epigenetic modifications on a genome). Epigenetic modifications are made in response to particular stimuli; these modifications in turn specify a particular pattern of gene expression. This gene expression profile dictates the identity of a cell. A stable form of gene regulation is critical to differentiation, as the identity of a cell must be maintained through all subsequent cell divisions

Answer 3

DNA methylation, histone acetylation and long non-coding RNAs (lncRNAs)

Answer 4

refers to the addition of methyl (CH3) groups to the nucleotide bases. The predominant form of DNA methylation is the methylation of the DNA base cytosine. DNA methylation often occurs to cytosines that are immediately adjacent to guanines. This combination is commonly referred to as a CpG dinucleotide (‘di’ meaning two and ‘p’ representing the phosphate group that connects the C and G nucleotides). These CpG dinucleotides are distributed across the genome non-randomly: they tend to cluster in “CpG islands” which are typically within the promoter regions of genes. DNA methylation promotes coiling and condensation of chromatin so methylation of CpG dinucleotides in the promoter region of a gene decreases transcription and reduces expression of the gene. Thus, cells can increase or decrease gene expression by demethylating and methylating DNA, respectively. Enzymes called DNA methyltransferases are responsible for the addition of methyl groups, whereas DNA demethylases are responsible for the removal of methyl groups

Answer 5

genomic DNA is treated with the chemical sodium bisulfite, which chemically converts unmethylated cytosines to uracils, which are then detected as thymines during sequencing. Importantly, only unmethylated cytosines are chemically altered during bisulfite sequencing. As such, methylated cytosines are unchanged (you can think of the methyl group as “protecting” the cytosine from conversion to uracil). By comparing sequencing of DNA that has been treated with sodium bisulfite with untreated DNA, we can determine the locations of all methylated cytosines in the genome

Answer 6

This uses an enzyme Tn5 transposase that evolved to facilitate the movement of segments of DNA around genomes. In essence, the enzyme attacks any piece of accessible DNA (it does not have sequence specificity like we see with CRISPR), cuts, and inserts material at the cut site. Tn5 transposase can only successfully attack chromatin in an ‘open’ configuration; it cannot access tightly packed chromatin. The DNA fragments generated by Tn5 transposase therefore correspond to the parts of the genome with open chromatin

Answer 7

‘writers’ or ‘erasers’. Writers are responsible for the addition of the modification (e.g., DNA methyltransferase) erasers are responsible for the removal of the modification (e.g., DNA demethylases). epigenetic tags are recognized by a third class of enzymes known as ‘readers’, which interpret the suite of epigenetic modifications to loosen or condense the DNA accordingly. When readers encounter DNA methylation they recruit additional proteins that can, for example, act as transcriptional repressors through additional chromatin remodeling

Answer 8

Histone modifications affect chromatin structure by chemically altering the positively charged tails of the histone proteins, which interact with the DNA. The best understood histone modification is the addition of acetyl groups to the amino acids on the histone tails. The writers responsible for acetylation are called histone acetyltransferases and the erasers that remove this modification are histone deacetylases. Whereas DNA methylation results in chromatin condensation and transcriptional repression, histone acetylation relaxes the chromatin, which increases transcriptional machinery accessibility, resulting in increased gene expression

Answer 9

a special type of RNA molecule that is transcribed but not translated. These RNAs are typically greater than 100 nucleotides in length and lack an open reading frame. Instead of coding for a protein, lncRNAs perform their functions as RNA molecules. IncRNAs work by interacting with proteins that regulate transcription such as transcription factors or chromatin binding proteins. Alternatively, they can directly bind to the nucleosomes, often in the process attracting additional proteins involved in epigenetic modification X-inactivation is an epigenetic phenomenon that is mediated by lncRNAs. Many epigenetic changes are dynamic and easily altered, but X-inactivation is remarkably stable. X-inactivation is necessary to ensure XX females express the appropriate amount of X chromosome gene product. The process occurs in two steps: First, the cell ‘counts’ the number of X-chromosomes present to determine whether inactivation is necessary. Second, when two X chromosomes are present, one is randomly selected for inactivation, while the other remains active. There are a number of lncRNAs involved in mediating X-inactivation; Xist (X-inactive specific transcript) is the best characterized. Xist expression is up regulated on the X-chromosome selected for inactivation, and then it coats the chromosome, marking it for inactivation. The coated chromosome is then silenced via high levels of DNA methylation and low levels of histone acetylation. This methylated chromosome condenses into an observable Barr body

Answer 10

in humans, approximately 10-15% of human genes on the X chromosome escape inactivation, but enough genes are inactivated to prevent issues with gene dosage

Answer 11

cellular identity is specified by a specific set of epigenetic modifications,

Answer 12

After semiconservative replication, any cytosine that was methylated prior to replication will now only be methylated on one strand – the template strand. The cytosine on the newly replicated strand will be unmethylated. We refer to this state as hemimethylated. Special methyltransferases recognize hemimethylated CpG dinucleotides and add methyl groups to the unmethylated cytosine bases on the new strand (remember the CpG on one strand is paired with a complementary GC on the other strand), thus restoring the methylation pattern to the same state as that of the parent cell.

Answer 13

Gametes are highly specialized and differentiated cells and, like other cell types, their gametic identity is specified by their unique epigenome. However, upon fertilization most or all of these epigenetic marks are removed to generate a totipotent zygote that can divide and give rise to all tissues of the body. The removal of epigenetic marks is triggered by the cellular environment in the zygote; this process is referred to as reprogramming. As development and differentiation progress, cells acquire distinct epigenetic modifications that specify their cellular identity. Some genes in gametes escape this reprograming. This is often associated with genomic imprinting. In the case of genomic imprinting, the gene’s expression in the individual is mediated by the epigenetic markers set by the parent during the gamete’s formation. Typically, the promoter of the silenced copy (whether maternal or paternal) is methylated

Answer 14

she is genetically indistinguishable from the workers: a queen is “created” by the provisioning of a bee larva with “royal jellyThis inhibits the action of a gene, Dnmt3, that causes genome-wide methylation of CpG sites, in the process reducing or preventing transcription of a large number of genes. These methylation-repressed genes are demethylated in queens, and therefore transcribed. Larvae can be turned into queens without royal jelly by using genetic tools that specifically repress Dnmt3 activity

Answer 15

in primordial germ cells (PGCs), which are the direct progenitors of sperm or oocyte. Epigenetic marks are then laid down, based on the cellular context, to direct these PGC progenitors to develop into a sperm or egg. In each of these two reprogramming windows – in PGCs and at fertilization – a unique set of mechanisms regulate DNA methylation erasure and re-establishment