Chapter 3: Gene expression (book) Flashcards
What are single-nucleotide polymorphisms (SNPs)?
A single-nucleotide polymorphism is a substitution of a single nucleotide at a specific position in the genome that is present in a sufficiently large fraction of the population
True/false: The majority of cancer-related SNPs are located in the coding regions of the genome
False, the majority is located in non-coding regions of the genome. This can be intergenic (regions between genes) or intronic (sequence of introns). Only a small number are found in coding regions
How can gene expression be modulated?
Through the regulation of transcription, chromatin structure and post-transcriptional mechanisms
Where do transcription factors (TF) bind to?
Gene promoters
What are the different independent domains TF can bind to?
- DNA-binding domains
- helix-turn-helix motif
- leucine zipper motif
- helix-loop-helix motif
- zinc finger motif,
- Transcriptional activation domains
- Dimerization domains
- Ligand binding domains
How do transactivation domains function?
By binding to other components of the transcriptional apparatus in order to induce transcription by RNA polymerase
Some transcription factors work in pairs. How is this called and how does this work
This pair is called a dimer. These transcription factors require a dimerization domain which facilitates protein-protein interactions between the two molecules
How can the activity of TF be regulated?
- Synthesis in particular cell types only
- Covalent modification such as phosphorylation
- Ligand binding
- Cell localization (if dimeric)
- Exchange of partner proteins
Which families of TF make up the AP-1 complex that plays a role in carcinogenesis?
The For and Jun family
Give an example of where the AP-1 complex plays a role in carcinogenesis
As TPA is a tumor promoter and the AP-1 complex binds to the TPA response element, an association of AP-1 with carcinogenesis was implicated early after this property was characterized. The first members of AP-1 identified, c-Jun and c-Fos were able to transform normal cells in culture to cancer cells and are frequently overexpressen in tumor cells
True/false: Steroid hormones only pass through the cell membrane and bind to their particular intracellular receptor in the cytoplasm to activate the TF
False, some don’t need to bind to a receptor and can directly enter the nucleus. Most, however, do need an intracellular receptor.
The retinoid acid receptor (RAR) is a member of the steroid hormone receptor family and is important during differentiation. RA is derived from vitamin A. Does the RAR act as a transcriptional activator or repressor in the absence of RA?
Repressor RAR acts as a transcriptional repressor in absence of RA.
What type of cancers are characteristic of aberrant forms of RARs?
Leukemias
Fill in: Somatic mutations upstream of the … oncogene, which codes for a basic helix-loop-helix transcription factor, have been shown to create a super-enhancer that upregulates the expression of this oncogene in T-cell leukemia
TAL1
What are super-enhancers?
Super-enhancers are clusters of regulatory DNA elements that recruit transcriptional and chromatin-modifying proteins and may be transcribed into RNA. Many are associated with key genes that control cell state
In a study was shown that DNA response elements for the MYB transcription factor are created by acquired insertional mutation. What does MYB then recruit?
MYB recruits important transcriptional (e.g. GATA3, Pol II) and epigenetic (e.g. histone lysine acetylene) regulators for strong expression of the TAL1 oncogene.
Note: a similar phenotype is exhibited when a deletion occurs that causes the TAL1 gene coding sequence to come under the regulation of a ubiquitously expressed promoter of the STIL gene
What is chromatin immunoprecipitation (ChIP) sequencing?
Chromatin immunoprecipitation (ChIP)-sequencing is a technique that is used to identify protein-DNA interactions at precise DNA sequences. The typical protocol involves cross-linking proteins to DNA, sonicating DNA to produce small fragments, adding bead-bound antibodies to capture your protein of interest for immunoprecipitation, unlinking protein from DNA, purifying DNA, and sequencing.
In a study by Mansour et al in 2014, a MYB-specific antibody was used to produce genomic maps of MYB binding in cells from the Jor-kat T-cell leukemia cell line and MOLT-3 cells that harbored insertional mutations of interest. What did the analysis show?
Precise alignment of MYB binding with other transcriptional and epigenetic regulators such as GATA3 and Pol II. This data suggests the formation of a super-enhancer.
What is the length of DNA when fully extended?
Over a meter (which is why it needs to be packaged tightly)
DNA is tightly packed around a nucleosome. What pattern does a row of nucleosomes show?
“beands-on-a-string”
The histone core is an October of histones. Which 4 histones (that are duplicated) are present there?
H2A, H2B, H3 and H4
Each histone contains domains. Can you name all the domains?
A domain for:
- histone-histone and histone-DNA interactions
- NH2-terminal lysine-rich
- COOH-terminal “tail” domains (which can be post-translationally modified)
What does H1 do?
It is a linker histone that binds to DNA outside the core
What is a secondary level of organization of DNA? (nucleosomes are primary)
Formation of 30 nm fibers
What tertiary structure can the 30 nm fibers form?
Radial loops. For an overview, see this figure
True/false: Histone modifications and DNA methylation can both be acquired or inherited
True
What post-translational modifications can occur on histone proteins?
Acetylation, methylation, phosphorylation and ubiquination
Fill in: Acetylation acts as a … (1) signal for the recruitment or the repulsion of … (2) factors.
1: docking 2: chromatin-modifying
Fill in: Acetylation of histones neutralizes the … charge on lysine residues and relaxes chromatin folding
Positive
Which enzyme adds, and which enzyme removes acetyl groups
HATs: add acetyl groups HDACs: remove acetyl groups
Transcriptional activators often recruit HATs and other chromatin-remodeling enzymes to the promoter region. Can you give an example of this?
The retinoblastoma tumor suppressor protein exerts is effects, in part, by recruiting HDACs to specific gene promoters. Thus a signaling network seems to underlie chromatin modeling.
Where, in DNA methylation, is the methyl group added?
To position 5 of cytosine, at the 5’ end of a guanine nucleotide (CpG) (3-4% of all cytosines are methylated)
Mature miRNAs require several steps of processing. Can you explain these steps?
- After they are transcribed by RNA polymerase II from intergenepic regions or from regions that code for introns, the primary transcript is processed by ribonucleases Drosha and DGCR8 in the nucleus.
- This processing produces pre-miRNAs, hairpin-shaped intermediates of 70-100 nucleotides.
- Exportin-5 transports pre-miRNAs into the cytoplasm where they are further processed by ribonuclease Dicer into a double-stranded miRNA.
- The strands separate, and a mature single-stranded molecule joins an RNA-induced silencing complex (RISC).
What are the two type of inhibitors that are used against DNA methylation?
There are two 5’-modified analogs of deoxcytidine, 5-azacytidine (5-azaC) and 5-aza-2’-deoxycytidine to target DNA methytransverases.
(remember DNA methylation occurs at position 5 on cytosine)
(also these have both been approved by the FDA)
