Topic 5 - Gene Transcription

Question 1

What technique could be used to determine specific mRNA levels in cells?

Answer 1

mRNA can be quantitated by reverse transcription-PCR (RT-PCR)

Question 2

What effect does DNA methylation have on the interaction of DNA with transcription factors?

Answer 2

Since transcription factors recognise specific sequences of bases, the presence of a bulky methyl substituent on the cytosine would sterically hinder recognition of the base by the transcription factor.

Question 3

Define epigenetics

Answer 3

The study of epigenetic changes to DNA, acting at different levels, between single genes and entire genomes.

Question 4

Define genomic imprinting

Answer 4

Epigenetic marks on individual genes or alleles, which are retained in daughter cells following cell division, and may be passed between generations of a species.

Question 5

Define inducible genes

Answer 5

Genes that are normally not expressed by a cell, but which may be transcribed following appropriate activation of the cell.

Question 6

Define differentiation.

Answer 6

The changes that occur in a cell as it becomes specialised. Differentiation occurs as a result of differential gene expression.

Question 7

What are housekeeping genes?

Answer 7

Genes that are required by and expressed in (virtually) all cell types in an individual. For example, the genes for the enzymes that catalyse glycolysis.

Question 8

What is dedifferentiation?

Answer 8

The reversal of the process of differentiation, so that a cell becomes less specialised, and may acquire the potential to differentiate into other cell types.

Question 9

What are induced pluripotent stem cells (iPSCs)?

Answer 9

Pluripotent stem cells created from fully differentiated cells by the re-expression of a core set of stem cell transcription factors, such as the Yamanaka factors: Klf4, Oct4, Sox2 and c-Myc.

Question 10

What are promoters?

Answer 10

Regions of a gene, usually 5’ to the transcription initiation point, where transcription factors bind that promote transcription.

Question 11

What is a core promoter?

Answer 11

A region of a gene, usually within 100 nucleotides of the transcription start site, where RNA polymerase II complex is assembled.

Question 12

What are target motifs?

Answer 12

A DNA sequence that binds to a particular DNA-binding protein.

Question 13

What are enhancers?

Answer 13

DNA sequences that elevate transcription rate in a position- and orientation-independent manner. Enhancers are often active only in specific cell types, and at specific developmental stages.

Question 14

What are silencers?

Answer 14

DNA sequences that reduce transcription in a position- and orientation-independent manner. These are often active only in specific cell types, and at specific developmental stages.

Question 15

What is the TATA-binding protein (TBP)?

Answer 15

A DNA-binding protein that recognises the sequence TATA and is part of the TFII complex required for assembly of RNA polymerase II at the core promoter.

Question 16

What is CAAT enhancer binding protein?

Answer 16

A ‘leucine-zipper’ transcription factor, which binds to the consensus sequence CCAAT found at the core promoter in many eukaryotic genes.

Question 17

what functions do RNA polymerase I and RNA polymerase III have?

Answer 17

RNA polymerase I transcribes the main ribosomal RNA (rRNA).
RNA polymerase III transcribes tRNAs, the small rRNA unit and other small RNAs found in the nucleus and cytosol, which are involved in mRNA splicing and gene control.

Question 18

What is a consensus sequence?

Answer 18

A consensus sequence is a common order of nucleotides in DNA, or amino acids in proteins. Proteins interact with DNA, and with other proteins, via specified consensus sequences. In DNA, the consensus sequence is a generalised nucleotide sequence derived from comparisons of closely related DNA sequences from many places within, or between, genomes.

Question 19

Consider Sp1; what would be the consequences for an organism if it were genetically engineered to lack this transcription factor? Why is this so? (Note: the common term for an organism in such circumstances is a ‘knockout organism’.)

Answer 19

Sp1 is required for the activation of many hundreds of genes needed for both basic and specialised cell functions. Therefore, knockout of the Sp1 gene is not compatible with life.

Question 20

How might topoisomerases be involved in regulating the accessibility of the DNA?

Answer 20

If the stress is not relieved, DNA wound around nucleosomes becomes supercoiled. By relieving the stress, topoisomerase allows the DNA to become more relaxed so that loops of 30 nm chromatin fibres are more acc

Question 21

What are histone marks?

Answer 21

Individual histone modifications and their distribution on a gene.

Question 22

What is the histone code?

Answer 22

The relationship between modifications to the histone N-terminal tails and the effects of these modifications, including gene activation and silencing and chromatin compaction.

Question 23

What are CpG islands?

Answer 23

Regions of a eukaryotic gene which contain a high proportion of cytosine and guanosine, often near the core promoter.

Question 24

What are transcription factories?

Answer 24

Regions within the nucleus of a eukaryotic cell where transcription of a number of genes is taking place.

Question 25

What is chromatin remodelling?

Answer 25

Changes in the chromatin structure caused, for example, by addition or removal of nucleosomes.

Question 26

What are hypersensitive sites in DNA mapping?

Answer 26

Regions of the genome that are particularly sensitive to digestion with DNAse.

Question 27

What is chromatin immunoprecipitation (ChIP)?

Answer 27

A sensitive and common technique for analysing which proteins or specific protein modifications are associated with a region of DNA in vivo through the use of antibodies specific for chromatin-bound proteins, followed by isolation and analysis of the DNA strands to which they are bound.

Question 28

If you wanted to know what genes are being actively transcribed into mRNA in a cell, what antibody could you use for ChIP?

Answer 28

An antibody against RNA polymerase II or another component of the RNA polymerase II complex would be suitable.

Question 29

ChIP is used to determine whether a protein is associated with a specific region of the DNA. What technique can be used to determine whether a protein is associated with another protein?

Answer 29

Co-immunoprecipitation (CoIP).

Question 30

The first codon of every eukaryotic mRNA has the sequence AUG which encodes methionine. What will be the sequence in the corresponding anticodon of the methionine-tRNA that translates this codon?

Answer 30

CAT – it is the antisense sequence complementary to the sense sequence AUG

Question 31

What are isoforms?

Answer 31

Variant forms of an individual protein, produced either from different, related genes, or by differential splicing of the mRNA from a single gene.

Question 32

Name a family of genes that have related functions but are differentially expressed in different cells.

Answer 32

There are many examples, but the Sp family and CEBP family are functionally diverse and expressed selectively in particular cell types.

Question 33

How can similar genes from a family of proteins come to be expressed in differ

Answer 33

The promoters and other controlling elements have diversified.

Question 34

What is RNA splicing?

Answer 34

The removal of introns from a primary RNA transcript to produce mRNA.

Question 35

What is differential splicing?

Answer 35

The splicing of different combinations of exons in a primary transcript that can result in synthesis of different polypeptides or mRNAs with different stabilities. Also called alternative splicing.

Question 36

What is the spliceosome?

Answer 36

A complex of proteins and RNA molecules that catalyses mRNA splicing.

Question 37

What is the polyadenylation signal sequence?

Answer 37

A sequence of nucleotides in the mRNA transcript that recruits poly-A polymerase to initiate production of the 3’ poly-A tail on eukaryotic mRNA.

Question 38

How could the process of polyadenylation be affected by factors that control differential splicing?

Answer 38

Whether an internal polyadenylation site will be used depends on whether the segment of RNA containing that site survives splicing long enough to be recognised by polyadenylation factors.

Question 39

What are splicing silencers?

Answer 39

Regions in an RNA transcript that bind to splicing repressors (proteins or ribonucleoproteins) to reduce the likelihood of splicing at a particular intron–exon boundary.

Question 40

What are splicing enhancers?

Answer 40

Regions in an RNA transcript that bind to splicing activators (proteins or ribonucleoproteins) and increase the likelihood of splicing at a particular intron–exon boundary.

Question 41

What are exon junction complexes (EJCs)?

Answer 41

A complex of proteins that mark the position of splice sites in eukaryotic mRNA.

Question 42

What is nonsense-mediated decay (NMD)?

Answer 42

A translation-associated pathway that recognises the presence of aberrant or premature stop codons within an open reading frame (ORF) of a eukaryotic mRNA. It targets such mRNAs for degradation.

Question 43

What is an exosome?

Answer 43

In the context of gene expression, and exosome is a complex of proteins that degrades RNA molecules from the 3’ or 5’ end. Note that this term can also refer to membrane-bound vesicles that are released from cells and carry nucleic acids.