Epigenetics & miRNA Flashcards
Epigenetics
Study of heritable changes in gene expression or phenotype caused by mechanisms other than chanes in DNA sequences
Explains how individuals w/the same DNA sequences (e.g., identical twins) have different disease profiles
3 major types of epigenetic modification
- DNA methylation
- Histone Modification
- Microribonucleic Acids (miRNAs or miRs)
DNA methylation
- attachment of methyl group to a cytosine base is followed by a guanine base (CpG dinucleotide)
- silences/transcriptionally inactivates gene (heavily methylated = less likely to be transcribed to RNA)
- Aberrant methylation can lead to silencing of tumor-suppressor genes in development of cancer
- Same process in X inactivation
Histone Modification
(e.g., histone acetylation and deacetylation, alterations in chromatin)
Chromatin compaction & organization help to regulate gene expression
Microribonucleic Acids (miRNAs or miRs)
- RNAs encoded by short DNA sequences (~22 nucleotides)
- located on introns of genes or transcribed from noncoding DNA located btwn genes
- untranslated
Functions of noncoding RNA/miRNA?
- noncoding RNAs (ncRNAs) have been shown to regulate gene expressions: e.g., RNA interference, gene cosupression, gene silencing, imprinting, DNA methylation
- MiRNA perform critical functions during development & cell differentiation
- MiRNA regulate diverse signalling pathways - e.g., oncomirs stimulate cancer development and progression
oncomirs
miRNA that stimulate cancer development and progression
Why are miRNAs linked to carcinogenesis?
b/c can act as either oncogenes or tumor suppressor genes
pluripotent
trait of embryonic stem cells - potential to become any type of cell in fetus or adult
How is eukaryotic DNA compacted 10,000-fold into chromosomes?
Histones!
What are histones
- The proteins that make up a nucleosome - there are four pairs
- histones are packers. They pack the protein. They are the beads – they are able to roll to the right or left, and unpack DNA as they have on them, expose it for whatever is needed – replication, transcription… Wind back in when the job is done.
Types of histones
Histones H2A, H2B, H3 and H4 are known as the core histones, while histones H1 and H5 are known as the linker histones.
Histone octamer
The collective of the four pairs of histones that make up a spool for DNA to coil around (nucleosome)
Histone Structure
- octomeric histone core (8 pieces). Come together to build bead structure.
- Histones contain an N-terminal tail which is subject to covalent modification
- Histones H2A and H2B form a dimer via a handshake
- Histones H3 and H4 form asimilar interaction
- N-terminal ends are different: They fold – when they pair and fold, their tails stick out in a certain way – very important
nucleosome
a basic unit of DNA packaging in eukaryotes, consisting of a segment of DNA wound in sequence around eight histone protein cores.
Chromatin
nucleosomes are organized into chromatin, the repeat building blocks of a chromosome
Nature of movement of nucleosomes
Dynamic!
Note when a part of a nucleosome is exposed, a protein can bind to it
What makes the nucleosome turn?
Chromatin remodeling complexes (ATP dependent!)
How do chromatin remodeling complexes work?
Histone exchange and histone removal are catalyzed by chromatin remodeling complexes
Changes H2A part of H2A-H2B dimer, for a new H2A. These cannot be altered, so histone chaperone protects them during the process.
This uses ATP.
Whole nucleosome can be rebuilt over time – but needs to frequently replace H2A-H2B piece. Once H2A is replaced for different H2A, that nucleosome exposes gene for action.
Who gets the replacement H2A?
Histone Tails!
The way they stick out, communicate message. Signals come from gene, and also from proteins attached to gene, and tails on proteins
Histone Tails and the path of DNA
Tails may attach one nucleosome to another
A single histone H1 binds both the histone octomer and DNA and may change the path of the DNA as it exits from the nucleosome thus allowing binding of nucleosomes to each other
Histone DNA contacts
primarily nonspecific
- hydrogen bonds (142) between basic histone Amino acids and the negatively charged DNA phosphodiester backbone
- hydrophobic and ionic interactions
- certain nucleotides favor staying inside, other favor outside. AA, TT, TA favor wrapping. G-C prefer outside.
Why does DNA need packaging & how is it packaged?
DNA is a long molecule: needs packaging to fit in the tiny nucleus. Packaging involves coiling of DNA in a “left-handed” spiral around spools. These spools/nucleosomes are made of four pairs of proteins known as histones
DNA methylation & tumor cells
Tumor cells usually exhibit hypomethylation, which can increase activity of oncogenes
DNA methylation and promoter regions + examples
often hypermethylated –> decrease in transcription rate & ability to inhibit tumor formation
e.g., RB1 in retinoblastoma & BRCA1 in breast cancer
DNA methylation & hereditary nonpolyposis colorectal cancer [HNPCC]
MLH1, whose protein product repairs damaged DNA, is methylated –> inactive –> damaged DNA accumulates –> colon tumors
mrRNAs and cancer
hypermethylation: miRNA encode small, 22 base pair RNA molecules that bind to mRNAs and degrade them/prevent translation
hypermethylation of specific subgroups of miRNA assoc w/tumorigenesis
methylation –> overexpressed targets –> metastasis
DNA mutations vs epigenetic modifications
DNA sequences can’t be directly altered, but epigenetic modifications can be reversed!
e.g., demethylating agents like 5-azacytidine in Tx of leukemia & myelodysplastic syndrome
treatment of histone modification problems
e.g. histone deacetylase (HDAC) inhibitors
counteract removal of acetyl groups from histone proteins – stop silencing tumor-suppressor genes
T-cell lymphomas
*challenge is targeting only specific cancer-causing genes
Imprinting
- process of gene silencing - in which genes are predictably silenced depending on which parent transmits them
- typically via heavy methylation
- clinical significance: if same group of genes are deleted, may cause different expression in child depending on whether inherited from mother or father - b/c different genes were active in that area based on gender
- Role for miRNA
location of miRNA
in the introns of genes (Intronic) or non-coding DNA between genes (Intergenic), or can code for multiple genes (Polycistronic)
DICER
- also known as endoribonuclease Dicer or helicase with RNase motif (scissor)
- An enzyme that in humans is encoded by the DICER1 gene.
- Dicer cleaves double-stranded RNA (dsRNA) and pre-microRNA (pre-miRNA) into short double-stranded RNA fragments called small interfering RNA and microRNA respectively.
process of miRNA
Drosha cleaves off hairpin piece of miRNA, which leaves nucleus and goes to cytosol. Dicer cuts pre-miRNA into miRNA and unused part, miRNA goes to target mRNA, e.g., to area of gene that should be silenced and blocks –> gene not expressed
Drosha
a Class 2 RNase III enzyme responsible for initiating the processing of microRNA (miRNA)
A microRNA molecule is synthesized as a long RNA primary transcript known as a pri-miRNA
become short segments, like hairpins. As a result of cleaving enzyme (Drosha), primitive RNA is cleaved into a pre-miRNA. At that point, it exits the nucleus, goes to cytosol, where part is used and other is cut off (by dicer enzyme). It is now available – role in RNA interference, gene code suppression, gene silencing, imprinting
Clinical significance of miRNA
Don’t get translated but signals that help development, metabolism, fate of cell, most importantly cell death.
cancer cells – we want that signal and why has cancer cell managed to escape it? Why have these cells altered miRNA, how can you restore the death signal in these cells?
Where does methylation occur?
At the 5 position of cytosine
when chromatin is open/active, genes are…
expressed
when chromatin is condensed b/c of methylation or histone modification, genes are…
inactivated
Where does histone acetylation happen?
At histone tails
more acetylation (by HAT): open chromatin/gene expression
deacetylation (by HDAC): closed (e.g., overdone in cancers)
HAT & HDAC are chromatin remodeling complexes
