Genetics 01 Flashcards
What is the difference between euchromatin and heterochromatin?
- Euchromatin is loosely packed, transcriptionally active chromatin where genes are accessible for expression.
- Heterochromatin is tightly packed, transcriptionally inactive chromatin that is harder to access.
What is HP1 (Heterochromatin Protein 1)?
HP1 (Heterochromatin Protein 1) is a key protein involved in maintaining heterochromatin structure and gene silencing. It binds to H3K9me3 (trimethylated histone H3 at lysine 9) and helps keep chromatin in a compact, transcriptionally inactive state.
What are types of Heterochromatine?
- Constitutive heterochromatin (e.g., centromeres & telomeres) is always silent
Always tightly packed and transcriptionally inactive (silent). Found in regions like centromeres and telomeres—these areas are structural and don’t need to be expressed. Enriched in repetitive sequences and has a high level of DNA methylation and histone modifications (like H3K9me3: epigenetic modification associated with heterochromatin and gene silencing) that keep it permanently condensed. - Facultative heterochromatin (e.g., Barr body) can switch between active and inactive states.
Can switch between active (euchromatin-like) and inactive (heterochromatin-like) states. Example: Barr body (inactive X chromosome in females). It’s tightly packed in one cell but can be reactivated in certain conditions. Its regulation depends on epigenetic modifications like histone methylation (H3K27me3 for repression) and demethylation when activation is needed.
What is Barr body?
A Barr body is an inactive X chromosome in female cells, appearing as a dense structure in the nucleus. It ensures dosage compensation, preventing females (XX) from having double the X-linked gene expression compared to males (XY).
How does X-Inactivation Works?
1) Random inactivation of either the maternal or paternal X occurs early in development.
2) XIST RNA coats the chosen X chromosome, triggering silencing.
3) Epigenetic modifications (H3K27me3, H3K9me3, DNA methylation) compact the chromatin.
4) The inactive X becomes a Barr body, remaining silent in all descendant cells.
This process creates mosaic gene expression, as different cells may inactivate different X chromosomes (e.g., in calico cats). The inactivation is stable but reversible in germ cells.
What is XIST RNA?
XIST (X-inactive specific transcript) RNA is a long non-coding RNA (lncRNA) that is responsible for turning off one X chromosome in female cells. It is the master switch for X-inactivation.
How does XIST RNA work?
1) Only the X chromosome that will be inactivated makes XIST RNA.
2) XIST RNA coats the X chromosome, spreading over it like a blanket.
3) It recruits silencing proteins and epigenetic marks like:
H3K9me3 and HP1 → help form heterochromatin
DNA methylation at CpG sites → locks the genes in an “off” state
4) This leads to permanent X-inactivation, turning the chromosome into a Barr body.
What do DNMTs do?
DNMTs (DNA methyltransferases) add or mantain methyl groups (-CH₃) to cytosine bases at CpG sites, forming 5-methylcytosine (5mC). This modification plays a key role in epigenetic gene regulation.
How does DNMT1 maintain methylation during DNA replication?
1) When DNA replicates, each new double strand consists of:
One old (methylated) strand
One new (unmethylated) strand
2) DNMT1 recognizes the hemi-methylated DNA (one methylated strand, one unmethylated strand) and adds a methyl group onto the 5’ position of cytosine (making it 5mC) to the new strand, keeping both strands methylated.
What is the difference between DNMT3 and DNMT1?
DNMT3a & DNMT3b → De novo methylation (adds new 5mC marks, e.g., during development).
DNMT1 → Maintenance methylation (copies existing 5mC marks during DNA replication).
What are 5mC Marks?
5mC (5-methylcytosine) marks are chemical modifications on DNA where a methyl group (-CH₃) is added to a cytosine base at CpG sites. This modification is a key part of epigenetic regulation and affects gene expression.
What are MBD proteins?
MBD (Methyl-CpG Binding Domain) proteins are a family of proteins that bind specifically to 5-methylcytosine (5mC) marks on DNA. Their job is to interpret DNA methylation and help regulate gene expression.
What are CpG Islands?
CpG islands are regions of DNA that have a high density of CpG dinucleotides (where a cytosine is followed by a guanine). These regions are often located near gene promoters and are crucial for regulating gene expression.
Key Features of CpG Islands
1) High CpG Content
CpG islands have a much higher frequency of CpG sites compared to the rest of the genome.
In CpG islands, the CpG content is at least 60%, whereas the rest of the genome only has about 1% CpG frequency.
2) Length
CpG islands are usually between 300 and 3,000 base pairs long.
3) Location
They are often found near or within gene promoters and enhancers, which are regions that control the activity of genes.
Around 40% of mammalian gene promoters contain CpG islands, and about 70% of human gene promoters have a high CpG content.
4) Hypomethylation
CpG islands are typically hypomethylated, meaning they have low levels of DNA methylation.
This hypomethylation is important because it allows for gene activation—methylation at CpG islands generally leads to gene silencing.
What is genetic imprinting?
Genetic imprinting is an epigenetic phenomenon where only one allele (either maternal or paternal) is expressed, while the other is silenced.
How is imprinting different from regular gene expression?
Normally, both copies of a gene (one from mom, one from dad) are expressed. In imprinting, only one parent’s allele is active, while the other is turned off by epigenetic modifications like DNA methylation.
When and how is imprinting established?
- Imprints (epigenetic marks) are added in sperm or egg cells (germline) before fertilization.
- These marks are maintained in the somatic cells of the offspring through mitotic divisions
Does imprinting follow Mendelian inheritance?
No, it’s an exception to classical Mendelian genetics because expression depends on which parent the gene came from, not just dominant or recessive alleles.
Why is imprinting important?
It regulates key genes involved in development, growth, and metabolism—especially during embryonic and early postnatal stages.
What happens if imprinting goes wrong?
Imprinting disorders occur when the wrong allele is silenced or both alleles are active/inactive. Examples:
Angelman syndrome → Maternal allele missing or silenced.
Prader-Willi syndrome → Paternal allele missing or silenced.
Several processes in development involve switching genes on or off via differential and dynamic DNA-methylation. What are some examples?
- Genomic imprinting of specific genes during mammalian embryonic development
- X-chromosome inactivation in female mammals (XX)
- Heterochromatin maintenance in fully differentiated cells
- Regulation of key developmental genes that control cell lineage during development
- Tissue-specific gene expression, where certain genes are active only in specific tissues
What are chromatin remodeling complexes and what do they do?
Chromatin remodeling complexes are protein groups that modify chromatin structure to regulate gene expression. They can either open chromatin (activate genes) or compact chromatin (repress genes) by using enzymes like DNMTs, HDACs, HATs, and HMTs.
How do chromatin remodeling complexes regulate transcription?
Gene silencing (Repressive complexes) → Tighten chromatin to prevent transcription:
DNMTs → Add DNA methylation (shuts genes off).
HDACs → Remove histone acetylation (compacts chromatin).
Repressive HMTs → Add methyl groups (H3K9me3, H3K27me3) to silence genes.
✔ Gene activation (Activating complexes) → Loosen chromatin to allow transcription:
TET enzymes → Remove DNA methylation (reactivates genes).
HATs → Add histone acetylation (opens chromatin).
Activating HMTs → Add methylation marks like H3K4me3 (enhances transcription).
✔ Non-coding RNAs (lncRNAs, miRNAs, snoRNAs) help regulate genes by controlling RNA stability, splicing, and translation.
What are histone N-terminal modifications?
These are covalent biochemical modifications on histone tails that influence chromatin structure and gene expression. Examples include acetylation, methylation, phosphorylation, ubiquitylation, and sumoylation at specific amino acid residues.
How do histone modifications affect gene expression?
Acetylation (HATs) → Activates genes by loosening chromatin.
Deacetylation (HDACs) → Silences genes by tightening chromatin.
Methylation (HMTs & HDMs) → Can activate or repress genes, depending on the location.
Phosphorylation (Kinases & Phosphatases) → Often signals transitions between activation and repression.
What is the “histone code” and why is it complex?
The histone code refers to the combinatorial pattern of post-translational modifications (PTMs) on histone proteins, which regulate chromatin structure and gene expression.
✔ Modifications include acetylation, methylation, phosphorylation, ubiquitylation, and sumoylation at specific histone residues.
✔ These modifications act together to determine whether chromatin is in an active (euchromatin) or repressed (heterochromatin) state.
There are over 120 known histone modification sites, creating a huge number of potential modification combinations.
✔ Context matters—the same modification (e.g., H3K4me3 vs. H3K9me3) can have opposite effects depending on location and interacting proteins.
✔ Histone marks work in networks, meaning that one modification can influence or reinforce others, leading to a layered and dynamic regulatory system.
What are epigenetic writers and erasers?
Epigenetic writers → Add histone modifications (e.g., HATs, HMTs, kinases).
Epigenetic erasers → Remove histone modifications (e.g., HDACs, HDMs, phosphatases).
How do histone modifications influence chromatin remodeling?
Activating marks (e.g., H3K4me3, H3K36me3, acetylation) → Recruit chromatin remodelers like NURF to open chromatin for transcription.
Repressive marks (e.g., H3K9me3, H3K27me3) → Recruit proteins that compact chromatin and silence genes.
What are histone variants and why are they important?
Histone variants are special forms of histones that can replace standard histones in nucleosomes. They influence gene regulation, DNA repair, and chromosome stability.
What are some key histone variants and their functions?
H3.3 → Found in active chromatin; promotes gene expression.
H2A.X → Involved in the DNA damage response (phosphorylated to γH2AX when DNA breaks occur).
How does histone phosphorylation signal DNA damage?
When double-strand breaks (DSBs) occur, H2A.X (histone variant) is phosphorylated at Ser139, forming γH2AX.
γH2AX spreads across chromatin and recruits DNA repair proteins to fix the damage. Facilitates chromatin remodeling to allow repair machinery access. Promotes homologous recombination (HR) and non-homologous end joining (NHEJ) repair pathways.
Why is chromatin structure remodeling important for transcription?
To activate a gene, chromatin must be opened using **histone modifications **and nucleosome remodeling complexes (e.g., NURF).
To turn a gene off, activating marks must be removed, and repressive marks (e.g., H3K27me3) must be added.
What are the two main ways RNA is epigenetically regulated?
Epigenetic control over RNA → Chemical modifications to RNAs (e.g., mRNA, rRNA, tRNA, miRNA, piRNA, lncRNA, snRNA).
Epigenetic control by RNA → Regulation of gene expression by non-coding RNAs (e.g., miRNAs, siRNAs, lncRNAs).
What is N6-Methyladenosine (m6A) and why is it important?
m6A is a chemical modification found in mRNA, tRNA, rRNA, and lncRNAs. It influences:
1) mRNA stability and degradation: m6A accelerates mRNA degradation
2) Translation efficiency: enhances translation by recruiting ribosomes
3) Alternative splicing: m6A-modified exons can be included or skipped
4) Regulation of non-coding RNAs (e.g., miRNAs, lncRNAs)
5) Nuclear Export: Helps transport mRNA from the nucleus to the cytoplasm.
m6A modifications can be added by writers (METTL3/METTL14), removed by erasers (ALKBH5, FTO), and read by reader proteins (YTHDF1, YTHDF2).
How does m6A affect translation under stress conditions?
m6A promotes selective translation of stress-response genes while suppressing non-essential mRNAs by:
✔ Enhancing cap-independent translation (e.g., IRES-dependent).
✔ Storing mRNAs in stress granules via YTHDF2/YTHDC1.
✔ Prioritizing protective proteins (e.g., HSP70, ATF4) for survival.
What are non-coding RNAs (ncRNAs) and their main functions?
ncRNAs do not encode proteins but regulate gene expression in various ways:
1) microRNAs (miRNAs) → Control mRNA degradation and translation repression.
2) Long non-coding RNAs (lncRNAs) → Can act as signals, scaffolds, guides, or decoys for transcriptional regulation.
3) piwi-interacting RNAs (piRNAs) → Protect the genome from transposon activity, mainly in germline cells.
4) Small nuclear RNAs (snRNAs) → Splicing and telomere maintenance.
5) Small nucleolar RNAs (snoRNAs) → Modify rRNAs and other RNAs.
What is the role of microRNAs (miRNAs) in gene regulation?
miRNAs bind complementary mRNA sequences (usually in the 3’-UTR).
If the match is perfect, the mRNA is degraded.
If the match is imperfect, translation is repressed.
miRNAs are processed from pri-miRNA → pre-miRNA → mature miRNA and incorporated into RISC (RNA-induced silencing complex).
What are lncRNAs and how do they regulate gene expression?
Long non-coding RNAs (lncRNAs) regulate genes through various mechanisms:
1) Signaling model → Act as molecular signals to activate/silence genes.
2) Decoy model → Bind and sequester miRNAs or proteins, preventing their function.
3) Guide model → Transport proteins to specific genomic locations.
4) Scaffold model → Bring multiple proteins together to form regulatory complexes.
Example: Xist lncRNA coats the X chromosome to mediate X-inactivation.
What are histone variants and how do they influence gene regulation?
Histone variants replace canonical histones and impact chromatin structure:
H3.3 → Found in active genes, promotes transcription.
H2A.X → Involved in the DNA damage response (phosphorylated form: γH2AX).
How does Fluorescence In Situ Hybridization (FISH) detect imprinting disorders?
FISH uses fluorescent probes to detect chromosomal deletions in Angelman Syndrome & Prader-Willi Syndrome:
✔ Can identify 15q11-q13 deletions but CANNOT detect UPD or imprinting defects.
✔ Works best for large deletions but lacks fine resolution for smaller mutations.
How does Array Comparative Genomic Hybridization (array-CGH) work in imprinting disorder detection?
Array-CGH compares patient DNA to a reference genome to detect copy number variations (CNVs):
✔ Identifies deletions or duplications in the 15q11-q13 region for Angelman & Prader-Willi Syndrome.
✔ CANNOT detect uniparental disomy (UPD) or epigenetic changes.
What is SNP array analysis, and how can it detect Angelman Syndrome?
SNP arrays analyze single nucleotide polymorphisms (SNPs) to identify:
✔ Uniparental disomy (UPD) → Lack of heterozygosity suggests both copies are from one parent.
✔ Deletions in the 15q11-q13 region.
✔ Imprinting defects (if combined with methylation-specific analysis).
What additional test is needed to confirm imprinting defects?
Methylation-Specific PCR (MS-PCR) detects abnormal methylation patterns, confirming imprinting disorders when SNP array or array-CGH results are inconclusive.
What is Chromatin Looping
Chromatin looping brings distant parts of the DNA into close proximity, allowing regulatory elements like enhancers to effectively interact with promoters, even if they are linearly separated by large distances. This spatial organization within the nucleus is critical for the precise regulation of gene expression.
What are Topologically Associating Domains (TADs)?
Topologically Associating Domains (TADs) are 3D chromatin structures where DNA forms loops, bringing distant genomic regions physically close together to regulate gene expression.
🔬 Key Features:
✔ TADs facilitate or inhibit long-range chromatin interactions, influencing enhancer-promoter communication.
✔ Cohesin protein complex forms and holds these loops together.
✔ CTCF (CCCTC-binding factor) helps define TAD boundaries, preventing unwanted interactions between adjacent domains.
✅ Functions:
✔ Regulate gene expression by enhancing or insulating contacts between regulatory elements.
✔ Maintain genome organization within the nucleus.
✔ Disruptions in TAD structure can lead to diseases like cancer or developmental disorders.
