Genetics: Chapter 15

Transcriptional silencing via methylation

1. Methylation inhibits the binding of an activator protein -Transcriptional activator binds to unmethylated DNA -This would inhibit the initiation of transcription 2. Methyl-CpG-binding protein recruits other proteins that change the chromatin to a closed conformation

3 common interactions that communicate effects of regulatory transcription factors

1. TFIID 2. Mediator 3. Recruiting proteins that affect nucleosome composition

In vertebrates and plants, many genes contain _____ near their promoters

CpG islands

DNA methylation is carried out by the enzyme ____

DNA methyltransferase

Combinatorial Control

Eukaryotic genes are regulated by a combination of proteins acting like a committee

What are the five histone genes that are moderately repetitive

H1, H2A, H2B, H3 and H4

Transcription Factors

Proteins that influence the ability of RNA polymerase to transcribe a given gene

Control elements

Regulatory transcription factors recognize cis regulatory elements located near the core promoter

The ferritin mRNA in mammals is controlled by the iron regulatory protein (IRP) that binds to an iron response element (IRE) near the 5' end of the mRNA. What do you expect to happen when iron levels are high?

The translation of the mRNA would be high because the IRP would not bind to the IRE.

Glucocorticoid Response Elements (GRE):

These function as enhancers -Located near dozens of different genes, so hormones can activate many genes

Why is eukaryotic transcription more complex than prokaryotic?

You need to open the chromatin

TFIID

a type of general transcription factor in eukaryotes that is needed for RNA polymerase II function. It binds to the TATA box and recruits RNA polymerase II to the core promoter

Transcription factor proteins contain regions called _____ that have specific functions

domains

An activator binds to a sequence called the _____

enhancer

Heterodimers

formed by two different transcription factors

Homodimers

formed by two identical transcription factors

coactivators

proteins that increase the rate of transcription but do not directly bind to the DNA itself

Repressor

regulatory protein that decreases the rate of transcription

An repressor binds to a sequence called the _____

silencer

down regulation

the binding of a transcription factor to a silencer decreases the rate of transcription

upregulation

the binding of a transcription factor to an enhancer increases the rate of transcription

orientation independent/bidirectional

which means that they can function in the forward or reverse orientation

Activator

-A regulatory protein that increases the rate of transcription

ChIP-Seq

-Allows determination of: -Where nucleosomes are located -Where histone variants are found -Where covalent modifications of histones occur

zinc finger motif

-Composed of one alpha helix and two beta sheets held together by Zinc metal icon

The ENCODE Project

-Encyclopedia of DNA Elements Consortium -Isolate and sequence RNA from humans -Identify transcription factor binding sites -Map DNA methylation sites -Identify histone modification sites -Map DNaseI cleavage sites

What are the two main types of transcription factors?

-General transcription factors -Regulatory transcription factors

Action of glucocorticoid hormones promoting glucose utilization, fat mobilization, and protein breakdown

1. Heat shock proteins released when hormone binds 2. Nuclear localization signal is exposed 3. Formation of homodimer 4. Transcription of target gene is activated

ChIP-Seq steps

1. crosslink DNA and protein 2. immunoprecipitate 3. reverse crosslinks and purify DNA 4. mass parallel sequencing and alignment

Methylated DNA sequences are inherited during ______

cell division

histone variants

histone proteins whose amino acid sequences are slightly different from the standard histones. They often play a specialized role in chromatin structure and function

-DNA methylation usually ___ the transcription of eukaryotic genes

inhibits

DNA methylation

is a change in chromatin structure that silences gene expression

How does ATP-dependent chromatin remodeling orchestrate changes in chromatin structure?

-Energy of ATP hydrolysis is used to drive change in location and/or composition of nucleosomes -Makes the DNA more or less amenable to transcription -The energy of ATP is used to alter the structure of nucleosomes and thus make the DNA more accessible

Which of the following event(s) need to happen for the glucocorticoid receptor to activate a gene?

-Protein-protein interaction (e.g., dimer formation) -The binding of the glucocorticoid receptor to a GRE -The binding of a small effector molecule to the glucocorticoid receptor subunits

general transcription factors (GTFs)

-Required for the binding of the RNA pol to the core promoter and its progression to the elongation stage -Are necessary for basal transcription

Regulatory transcription factors

-Serve to regulate the rate of transcription of the target gene -They influence the ability of RNA pol to begin transcription of a particular gene

How did Chromatin Immunoprecipitation Sequencing reveal a common pattern of nucleosome organization

-Showed that the nucleosome-free region (NFR) is found at the beginning and end of many genes -Nucleosomes tend to be precisely position near the beginning and end of the gene, but less regularly distributed elsewhere

How Methylated DNA sequences inherited during cell may explain genomic imprinting

-Specific genes are methylated in gametes from mother or father -Pattern of one copy of the gene being methylated and the other not is maintained in the resulting offspring

CpG islands in housekeeping genes

-The CpG islands are unmethylated -Genes tend to be expressed in most cell types

histone code

-The amino terminal tails of histones can be selectively modified, such as with acetylation, methylation and phosphorylation are common -These modifications affect the level of transcription and May influence interactions between nucleosomes -Occur in patterns that are recognized by proteins

CpG islands in tissue-specific genes

-The expression of these genes may be silenced by the methylation of CpG islands -Methylation may influence the binding of transcription factors -Methyl-CpG-binding proteins may recruit factors that lead to compaction of the chromatin

CREB protein

-another regulatory transcription factor -CRED protein factors become activated in response to cell-signaling molecules that cause an increase in the cytoplasmic concentration of cAMP

steroid receptors

-regulatory transcription factors that respond to steroid hormones -The hormone actually binds to the transcription factor

Insulators

-segments of DNA that insulates a gene from the regulatory effects of other genes -Some act as barriers to chromatin remodeling -Others block the effects of enhancers (May do this by chromosome looping)

Factors contributing to combinatorial control:

1. 1 or more activator proteins may stimulate transcription 2. 1 or more repressor proteins may inhibit transcription 3. Activators and repressors may be modulated by: -Binding of small effector molecules -Protein-protein interactions -Covalent modifications 4. Regulatory proteins may alter nucleosomes near promoter 5. DNA methylation may inhibit transcription -Prevent binding of an activator proteins -recruiting proteins that compact the chromatin

Steps in transcriptional activation

1. Activator binds to enhancer 2. Chromosome remodeling and histone modification: regions open more and recruit histone modifying enzyme 3. Formation of the preinitiation complex 4. Elongation phase

3 ways that function of regulatory transcription factors can be modulated:

1. Binding of a small effector molecule 2. Protein-Protein interactions 3. Covalent modifications

ATP-dependent chromatin remodeling steps

1. Change in nucleosome position: These changes may significantly alter gene expression 2. Histone octamers are removed: Creates a gap with no nucleosomes 3. Replacement with Histone variants: region with variant histones which affect transcription

What is the molecular model for the inheritance of DNA methylation?

1. DNA initially undergoes de novo methylation, a rare highly regulated event 2. DNA replication produces hemimethylated DNA molecules, which are fully methylated by DNA methyltransferase

Gene regulation is necessary to ensure what two things:

1. Expression of genes in an accurate pattern during the various developmental stages of the life cycle -Some genes are only expressed during embryonic stages, whereas others are only expressed in the adult 2. Differences among distinct cell types Nerve and muscle cells look so different because of gene regulation rather than differences in DNA content

Activity of CREB protein

1. Extracellular signaling molecule 2. cAMP 3. Phosphorylated CRED binds to DNA and stimulates transcription -Unphosphorylated CREB can bind to DNA, but cannot activate RNA pol

helix-loop-helix motif

2 alpha helices intertwined due to leucine motifs -Alternating leucine residues in both proteins interact resulting in protein dimerization

CREB protein recognizes a response element with the consensus sequence______________

5' - TGACGTCA - 3' -Termed cAMP response element

Motif

A domain, or a portion of a domain that has very similar structure in many different proteins