Chapter 5

Question 1

What are the four things changes in gene expression are involved in?

Answer 1

1. Response of cells to their environment
2. Regulation of cell cycle
3. Distinct activities and coordination of different cell types in tissue and organ function
4. Cell differentiation and development

Question 2

What are the differences and similarities between DNA polymerase and RNA polymerase?

Answer 2

RNA P. Are responsible for RNA synthesis
DNA P. Are responsible for DNA synthesis
Both made up of multiple polypeptide chains
DNA P. Needs a primer to start making DNA where RNA P. Does not need primer

Question 3

What are the three types of RNA polymerases in eukaryotic cells?

Answer 3

Messenger RNAs- templates for protein synthesis (RNA polymerase II), product of large number of different protein coding genes, correct gene is transcribed in the correct cell type at the correct time in the correct amount
Ribosomal RNAs- component of ribosomes (DNA polymerase I only does ribosomal RNAs and nothing else, and DNA polymerase III)
Transfer RNAs- adaptor molecules that align amino acids along the mRNA template
(DNA polymerase III)

All share several features common with eachother

Question 4

What does cis acting DNA mean?

Answer 4

Cis means within the same molecule
Cis acting DNA sequence means working on the same molecule being talked about
These are also called promoters and enhancer elements that regulate expression of genes

Study diagram of the cis acting regulatory elements of eukaryotic genes transcribed by RNA polymerase on slide 14

Question 5

What is the basal or core promoter?

What is the upstream promoter and enhancers?

Answer 5

Basal or core promoter- genes transcribed by RNA polymerase II contain core promoter elements and an initiator (TATA and INR on slide 14 graph)
Upstream promoter and enhancers- transcriptional regulatory sequences that can be located at a significant distance from the promoter (can be up or downstream)
Promoters and enhancers are structurally the same
Examples of core/upstream promoters on slide 17 and enhancers and promoters on slide 18-19

Question 6

How does your body identify eukaryotic gene regulatory sequences using reporter genes?

Answer 6

Reporter gene quickly assesses and reports back to you on the activity of a promoter
Slide 16 picture

Question 7

What is the difference between the position of promoters and enhancers?

Answer 7

Enhancers are position independent (can be anywhere)
The sequence is the important part
Promoters are position dependent and sequence dependent

Heat shock protein example on slides 20-23

Question 8

What are the two categories of transcription factors?

Answer 8

General transcription factors are involved in transcription from all polymerase II promoters (basic transcription machinery) form transcription initiation complex (known as preinitiation complex or basal transcription complex)
Gene specific transcription factors- not part of general machinery, bind to promoter or enhancer elements of different genes
Modulate activity of the basal transcription complex (modify rate of transcription initiation or rate of assembly of BTC)

Question 9

What is the transcription initiation complex (or preinitiation complex or basal transcription complex)?

Answer 9

Complex formed by general transcription factors and RNA polymerase II when they assemble inna step wise manner around basal promoter region
This initiates transcription but does not affect the rate of it
Necessary but not sufficient for transcription
Like a engine to a car, you need it but it can’t move the car on its own
Slide 26

Question 10

What specifies the amount of transcriptional initiation by RNA polymerase II?

Answer 10

The combo of transcriptional factors interacting at a specific genes promoter/enhancer sequences at any one point in time
They constitute the molecular dimmer switch

Question 11

What is DNA-affinity chromatography?

Answer 11

A method used to isolate transcription factors based on their binding to specific DNA sequences in order to study them further
Example on slide 34

Question 12

What is chromatin immunoprecipitation?

Answer 12

It identifies DNA binding sites of a transcription factor within the genome of a cell
Requires an antibody that specifically recognizes the transcription factor of interest
Pictures on slides 35 and 36

Question 13

What are the two things transcriptional factors can function as?

Answer 13

They can function as transcriptional activators and transcriptional repressors
Upregulate or downregulate gene expression
The factors contain separate DNA binding and activation/repression domains
Many transcription factors contain many distinct types of sequence specific, DNA-binding domains

Question 14

What are the two examples of gene specific transcription factors?

Answer 14

Steroid hormone receptors- regulate gene transcription in response to hormones like estrogen and testosterone
Homeodomain protiens- regulation of gene expression during embryonic development

Question 15

Once a DNA binding protein binds specifically, what happens next?

Answer 15

2 general mechanism of action:
1. Interact with mediators and general transcription factors
Mediator is a large protein that binds to preinitiation complex and links general transcription factors to gene specific transcription factors
2. Interact with coactivators, which modify chromatin structure

Slides 40 and 41

Question 16

What is DNA looping?

Answer 16

When DNA loops around (slide 42) which allows transcription factors bound to a distant enhancer to interact with protiens in the RNA polymerase/Mediator complex at the core promoter
It brings a transcription factor at a distal sequence close to RNA polymerase II

Question 17

What are topologically associating domains (TODs) and insulators?

Answer 17

TODs are a small structural region in DNA that contains all the sequences that can regulate a specific gene
Insulators are boundaries of TODs that are the binding site for a protein known as CTCF and others
Slide 43

Question 18

What releases the RNA polymerase from the basal complex to initiate transcription?

Answer 18

The phosphorylation and helicase activities of TFIIH

Helicase unwinds DNA activity, takes double strand and unwinds/relaxes it

Question 19

What are the functions of eukaryotic transcriptional repressors?

Answer 19

Gene expression in eukaryotes is regulated by repressors and by transcriptional activators
Function by blocking binding of transcriptional activator to DNA sequence, repressing mediator, and interacting with co repressors
(Slow down or shut down chromatin activity)
Slide 46 picture

Question 20

What are the 2 mechanisms in which chromatin can be modified for transcriptional modification?

Answer 20

1. Chromatin remodelling- moving, removal or addition of histones opens up or relaxes chromatin confirmation which allows for transcription
2. Histone code- histones tail acetylation and deacetylation or other histone modifications
   Acetylation results in transcriptionally permissive chromatin, deacetylation gives rise to non permissive state
   Slide 51-53 picture

Question 21

What is DNA methylation?

Answer 21

Plays an important role in genomic imprinting where only one of the two parental alleles of some genes is expressed during embryonic development
Slide 56 picture

Question 22

What is pre-mRNA?

Answer 22

Primary transcript that is processed to form messenger RNA in eukaryotic cells
Primary transcripts can undergo extensive modifications before they are transported from the nucleus to the cytoplasm
Some stages of processing shown on slide 60

Question 23

What caps the 5’ free end of RNA at the start of the transcribed RNA?
(Step 1 in processing eukaryotic messenger)

Answer 23

7-methylguanosine cap is added during modification before transcription has ceased

Question 24

What is added to the 3’ ends of eukaryotic mRNAs in the processing of mRNA in Eukaryotes? (Step 2)

Answer 24

Poly-A tail is a tract of about 200 adenine nucleotides added to the 3’ ends of eukaryotic mRNAs
Polyadenylation is the process of adding a poly-A tail to pre-mRNA
Polyadenylation signal is AAUAAA, this is the site where it will occur

Question 25

What are the two steps of pre-mRNA?

Answer 25

Step 1: cleavage at 5’ splice site
Formation of lariat like intermediate
Step 2: cleavage at 3’ splice site
Ligation of exons
Exon 1 & 2 separate from RNA
slide 67

Question 26

What are the mechanisms used for splicing?

Spliceosomes, snRNAs, snRNPs

Answer 26

Spliceosomes are large complexes composed of proteins and RNAs
Small nuclear RNAs (snRNAs) form the RNA component of the Spliceosome
Small nuclear ribonucleoprotein particles (snRNPs) are complexes of snRNAs with proteins that play central roles in the splicing process
Multiple snRNP complex forms the spliceosome
Slide 69-71

Question 27

What is alternative splicing?

Answer 27

The generation of different mRNAs by varying the pattern of pre-mRNA splicing
Creates multiple different but related proteins from the same gene
Slide 73 hypothetical example

Question 28

What are SR splicing factors?

Answer 28

They assist in directing where a spliceosome will form by binding to exon sequences
SR proteins will differ in two different cell types
They help determine the final choice of exons that will appear in any one cell type for any one alternatively spliced mRNA

Question 29

What is the Dscam gene?

Answer 29

Contains four sets of alternative exons, with a single exon from each set being incorporated into the spliced mRNA
Dscam= cell surface adhesion molecule important in allowing neurons to find their target cells
Allows cell to read road markers as it finds its target muscle cell
Slide 76 example

Question 30

What are 5.8S, 18S and 28S rRNA?

Answer 30

Makes up majority of cellular RNA (over 80% of cellular RNA)
The products of one unique gene that is present in multiple repeated copies within the genome
Transcribed by RNA polymerase I (boring job only does one thing but does a lot of it)
Transcription of these on slide 79

Question 31

What is the rRNA “Christmas tree”?

Answer 31

A micro graph showing three tandem rRNA genes being transcribed by RNA polymerase I
Can be 100 RNA polymerase I molecules transcribing the same gene at once
Picture on slide 80