Eukaryotic Gene Regulation – Lecture 1

Key vocabulary terms and definitions related to eukaryotic gene regulation, focusing on epigenetics, chromatin, transcription factors, and gene expression.

Epigenetics

A change in the state of expression of a gene that does NOT involve a mutation, but is inherited after cell division.

Chromatin

The complex of DNA and proteins that forms chromosomes; can be modified to regulate gene expression.

Differential Gene Expression

The process by which different cell types express different sets of genes, leading to functional diversity.

Transcription Factors (TFs)

Proteins that bind to specific DNA sequences to regulate the transcription of genes.

DNA Methylation

An epigenetic mechanism that involves adding a methyl group to DNA, typically acting to repress gene expression.

Histone Modifications

Chemical modifications of histones, such as methylation or acetylation, that influence chromatin structure and gene expression.

Nucleosome

The fundamental unit of chromatin, consisting of a segment of DNA wound around a core of histone proteins.

DNase I Hypersensitive Sites

Regions of open chromatin where nucleosomes are absent or displaced, indicating areas active in transcription.

Promoter

A region of DNA that initiates transcription of a particular gene, typically located upstream of the gene.

Gal4–LexA Chimera Experiment

An experiment demonstrating that the DNA-binding domain of transcription factors specifies which promoter is activated.

Epigenetic Memory

The capacity of epigenetic marks to maintain a memory of gene activity/inactivity without altering the DNA sequence.

Tissue-specific Gene Expression

The specific expression of genes in particular tissues, regulated by both transcription factors and epigenetic modifications.

Nucleosome Remodeling Complexes

Enzymatic complexes that alter nucleosome positioning or composition to facilitate access to DNA sequences for transcription.

Cis-acting Sequences

Regulatory DNA sequences located on the same molecule as the gene they regulate.

Trans-acting Factors

Factors (usually proteins) that regulate gene expression but are encoded by genes located elsewhere in the genome.

Histone Acetylation

The addition of an acetyl group to histone proteins, typically associated with transcriptional activation.

Histone Methylation

The addition of methyl groups to histones; depending on the site and context, it can either activate or repress transcription.

Polycomb / Trithorax Complexes

Protein complexes that maintain repressed or active chromatin states via histone methylation.

Differential Gene Expression Mechanisms

Different transcription factors and chromatin modifications that contribute to the variability in gene expression among cell types.

CpG Methylation

The addition of methyl groups to the cytosine base of CpG dinucleotides in DNA, often leading to gene silencing.

DNMT1

DNA methyltransferase responsible for the maintenance of DNA methylation patterns after DNA replication.

Non-Homologous End Joining (NHEJ)

A DNA repair pathway that repairs double-strand breaks without requiring a homologous template.

Homologous Repair (HR)

A DNA repair mechanism that uses a homologous sequence as a template for accurate repair.

Chromatin Structure

The organization of chromatin into nucleosomes and higher-order structures, which impacts gene accessibility and expression.

What is the central question of eukaryotic gene regulation?

How can different cell types in an organism contain the same DNA yet exhibit different behaviors and appearances?

What evidence shows that different cell types express different proteins?

2D gel electrophoresis demonstrates that human brain and liver cells have distinct protein profiles despite identical genomes.

What do cloning experiments reveal about DNA during differentiation?

Cloning experiments, such as frog nuclear transplant, show that DNA is not lost during differentiation.

What are the two main mechanisms of epigenetics?

DNA methylation and certain histone modifications.

What is the definition of epigenetics?

A change in gene expression that does not involve a mutation but is inherited after cell division.

What role do transcription factors (TFs) play in gene expression?

TFs are expressed differently in various cell types and determine which genes are activated.

What is the difference between active and repressed chromatin?

Active chromatin (euchromatin) is open and accessible to transcription factors, while repressed chromatin (heterochromatin) is compact and inaccessible.

What are DNase I hypersensitive sites?

Regions of open chromatin where nucleosomes are absent or displaced, found at promoters and enhancers of actively expressed genes.

What is the function of nucleosome remodeling complexes?

They use ATP hydrolysis to alter histone-DNA contacts, creating accessible sites for transcription factors.

What are the two main mechanisms of nucleosome remodeling?

Sliding (moving the histone octamer along DNA) and displacement (evicting the histone octamer entirely).

What types of modifications can occur on histone tails?

Acetylation, methylation, phosphorylation, and ubiquitylation.

What is the role of histone acetyltransferases (HATs)?

HATs add acetyl groups to histone tails, loosening chromatin structure and facilitating transcription.

What is the significance of the Gal4-LexA chimera experiment?

It demonstrated that the DNA-binding domain of transcription factors determines target gene specificity.

How do transcription factors recruit nucleosome remodeling complexes?

TFs bind first to specific DNA sequences and then recruit remodeling complexes to facilitate transcription.

What is the role of the TATA box in transcription?

The TATA box is a core promoter element that binds the TATA-binding protein (TBP) as part of the transcription initiation complex.

What is the function of chromatin modifications in gene expression?

They regulate access to DNA by transcription factors, influencing gene activity and cellular memory.

What is the relationship between transcription factors and chromatin structure?

Transcription factors can modify chromatin structure, making DNA more or less accessible for transcription.

How does the estrogen receptor illustrate tissue-specific gene expression?

The same estrogen receptor can activate different genes in different tissues due to varying epigenetic landscapes.

What is the significance of cellular memory in gene expression?

Cellular memory allows cells to maintain their identity and function through multiple divisions, even in the absence of specific signals.

What are the roles of acetylation and methylation in histone modifications?

Acetylation generally activates gene expression by loosening DNA-histone interactions, while methylation can either activate or repress gene expression depending on the context.

What is the main purpose of chromatin remodeling during gene activation?

To create DNase I hypersensitive sites that allow transcription factors to bind and initiate transcription.

What is the role of the activation domain in transcription factors?

It recruits co-activators and the basal transcription machinery to facilitate gene activation.

What is the significance of the N-terminal tails of histones?

They are the primary sites for chemical modifications that influence chromatin structure and gene expression.

What is the function of the CAAT box in gene regulation?

The CAAT box is a promoter element that binds general transcription activators to enhance gene expression.

What is the role of the GC box in transcription?

The GC box binds the transcription factor Sp1, which is involved in the regulation of gene expression.

What do undifferentiated fibroblasts NOT convert to in Medium A?

Cartilage

What determines gene expression in cells?

Chromatin state, not just signal environment

What do imaginal disc cells in Drosophila retain?

Their developmental identity even when transplanted

Which histone modifications maintain cellular memory in Drosophila?

Polycomb (repression) and Trithorax (activation)

What occurs to one X chromosome in female mammals during early embryonic development?

It is randomly inactivated

What mediates the stable inheritance of X chromosome inactivation?

Epigenetic marks like DNA methylation and histone modifications

What happens to DNA methylation patterns in identical twins as they age?

They diverge increasingly

What drives epigenetic change throughout life?

Environmental factors

Which statement is INCORRECT regarding DNA sequence during development?

The DNA sequence changes during development (it does NOT change)

What is the role of DNMT1?

Maintenance methyltransferase that restores methylation after DNA replication

What does DNMT3a do?

Establishes new methylation patterns during embryonic development

What is passive demethylation?

Occurs when DNMT1 fails to methylate the hemi-methylated strand after replication

What is active demethylation?

Enzymatic removal of the methyl group that does not require replication

What is the significance of the C→T transition at CpG sites?

It accounts for ~50% of point mutations in human genetic diseases

What does the term 'epigenetics' refer to?

Heritable changes in gene expression not involving DNA sequence changes

What is a DNase I hypersensitive site?

Region of open chromatin lacking nucleosomes, marking active promoters/enhancers

What is the function of HAT (Histone Acetyltransferase)?

Adds acetyl groups to histone tails, promoting open chromatin

What is the role of SWI/SNF complexes?

They are major nucleosome remodelling complexes that contain a Bromo domain

What does ATAC-Seq do?

Maps open chromatin genome-wide using Tn5 transposase

What is the function of Polycomb and Trithorax complexes?

Maintain repressed and active chromatin states via histone methylation

What is the difference between cis-acting and trans-acting factors?

Cis-acting factors are on the same DNA molecule, while trans-acting factors are encoded elsewhere

What is the genomic distribution of CpG dinucleotides in mammals?

70-80% are methylated, but 5-methylcytosine is only ~1% of total DNA bases

What causes the under-representation of CpG dinucleotides in mammalian DNA?

Mutagenic loss over evolutionary time due to spontaneous deamination

What is the role of MBD4?

Contains a methyl-CpG binding domain and a glycosylase domain for DNA repair

What is the preferred substrate for DNMT1?

Hemi-methylated DNA

What is the main function of DNMT2?

Methylates tRNA, not DNA, and has very weak DNMT activity

What happens during fertilization and blastocyst formation regarding methylation?

Global demethylation occurs, protecting imprinted genes and certain repeat elements

What is the significance of the H3K4-unmethylated tails for DNMT3L?

Stimulates DNMT3a/3b activity when bound

What does MBD4 contain?

An MBD (methyl-CpG binding domain) and a C-terminal glycosylase domain.

What type of mismatches does MBD4 bind?

G:T mismatches at CpG sites.

What is the primary function of MBD4?

To remove the T base to enable base excision repair (BER).

Are all 5-meC→T mutations corrected by MBD4?

No, repair is incomplete; not all mutations are corrected.

What are CpG islands?

Regions that are approximately 1 kb in length, GC-rich, and protected from methylation.

How many CpG islands are present in the haploid human genome?

Approximately 25,500.

Where are CpG islands typically found?

At the promoters of all housekeeping genes and about 40% of tissue-specific genes.

What prevents DNMT activity at CpG islands?

Transcription factor binding physically blocks DNMTs.

What is the role of H3K4me3 in relation to CpG islands?

H3K4me3 blocks DNMT3L binding, preventing de novo methylation.

What does bisulphite sequencing do?

Converts unmethylated C to U, allowing distinction between methylated and unmethylated cytosines.

What is the effect of DNA methylation on transcription?

It can cause approximately 50-fold repression of transcription.

What are the two mechanisms by which DNA methylation represses transcription?

Direct mechanism (methyl group prevents TF binding) and indirect mechanism (MBPs block TF access).

What is the primary function of MeCP1?

To mediate transcriptional repression through binding to methylated DNA.

What distinguishes weak CpG islands from strong CpG islands?

Weak CpG islands show differential methylation between tissues.

What happens to H3K4me3 levels when Cfp1 is knocked down?

There is a loss of H3K4me3 at CpG islands.

What is the significance of the Cfp1-SETD1-H3K4me3 axis?

It maintains unmethylated states at CpG islands, promoting transcription.

What is the role of MBD proteins in transcriptional repression?

They bind to methylated CpG and recruit complexes that compact chromatin.

What are the key features of MBD2?

Contains MBD and TRD, binds methylated CpG, and is part of the NuRD complex.

What does the term 'orphan CGIs' refer to?

Intergenic CpG islands that are also sites of transcriptional initiation.

What is the main function of the Kaiso protein?

To bind methylated CGCG sequences as part of MeCP1 activity.

What is the primary role of DNMT1?

To perform maintenance methylation on hemi-methylated DNA after replication.

What is the effect of aberrant de novo methylation on CpG islands?

It leads to transcriptional silencing by recruiting MBPs.

What is the relationship between CpG density and gene regulation?

Higher CpG density correlates with stronger regulation and repression.