Nucleic Acids - 7.2 Transcription (HL (+ some SL))

Question 1

Transcription = basic understandings

Answer 1

• one of the two main stages in protein synthesis
• involves making a copy (mRNA) of a section of DNA (a section of DNA unwinds inside the nucleus)
• THYMINE is replaced with URACIL in the mRNA molecule
• the mRNA molecule moves (from the nucleus) to the ribosomes where protein synthesis takes place

Transcription is the process by which a DNA sequence (gene) is copied into a complementary RNA sequence by RNA polymerase

1. Free nucleotides exist in the cell as nucleoside triphosphates (NTPs), which line up opposite their complementary base partner
2. RNA polymerase covalently binds the NTPs together in a reaction that involves the release of the two additional phosphates
3. The 5’-phosphate is linked to the 3’-end of the growing mRNA strand, hence transcription occurs in a 5’ → 3’ direction

Many RNA polymerase enzymes can transcribe a DNA sequence sequentially, producing a large number of transcripts

In eukaryotes, post-transcriptional modification of the RNA sequence is necessary to form mature mRNA

Question 2

7.2 - transcription and gene expression = UNDERSTANDINGS

Answer 2

1. Transcription occurs in a 5’ to 3’ direction
2. Nucleosomes help to regulate transcription in eukaryotes

(DNA is wrapped around histone proteins forming nucleosomes - the ends of histone proteins can be modified. When histone proteins are methylated the transcription of genes can be promoted or inhibited)

3. Eukaryotic cells modify mRNA after transcription

(The presence of a nuclear membrane in eukaryotic cells means that transcription and translation occur in different locations. mRNA can be extensively modified (post-translation modification) before it leaves the nucleus (eg. introns can be removed from mRNA before translation occurs)

4. Splicing of mRNA increases the number of different proteins an organism can occur

(Some genes code for multiple proteins - these genes have a number of (and alternative introns) introns that cant be spliced (cut) out of the mRNA = different proteins being formed)

5. Gene expression is regulated by proteins that bind to specific base sequences in DNA

Question 3

Nucleosomes in transcription

Answer 3

Nucleosomes help to regulate transcription in eukaryotes

(DNA is wrapped around histone proteins forming nucleosomes - the ends of histone proteins can be modified. When histone proteins are methylated the transcription of genes can be promoted or inhibited)

Question 4

Eukaryotic and mRNA after transcription

Answer 4

Eukaryotic cells modify mRNA after transcription

(The presence of a nuclear membrane in eukaryotic cells means that transcription and translation occur in different locations. mRNA can be extensively modified (post-translation modification) before it leaves the nucleus (eg. introns can be removed from mRNA before translation occours

Question 5

___

Answer 5

One side of the DNA acts as a template on which the complementary molecule of mRNA is made - this process is catalysed by the enzyme RNA polymerase (transcriptase) and only occurs in the 5’ to 3’ direction. RNA polymerase adds the 5’ end of the free RNA nucleotide tot he 3’ end of the growing mRNA molecule

Question 6

IMPORTANT NOTE - exons and introns

Answer 6

only the EXONS (coding regions) of the DNA are KEPT –> INTRONS (non-coding are edited/cut out)

Question 7

why introns are cut out (taken from google)

Answer 7

they actually have to be removed in order for the mRNA to encode a protein with the right sequence. If the spliceosome fails to remove an intron, an mRNA with extra “junk” in it will be made, and a wrong protein will get produced during translation.

Question 8

Promoters

Answer 8

= a sequence of DNA that is located near a gene
= binding site of RNA polymerase (the enzyme that catalyses the formation of the covalent bond between nucleotides during the synthesis of RNA)

(the promoter is not transcribed by is an example of non-coding DNA having a function

Question 9

(basic) outline the process of transcription

Answer 9

1. A section of DNA is unwound (inside the nucleus) and unzips - codes for a polypeptide
2. copy of the section (= transcription section) is made and called mRNA (one side of the DNA acts as the template)

= Processed by the catalyst RNA polymerase (transcriptase) and only occurs in 5’ to 3’ direction

Question 10

Controlling gene expression in prokaryotes - THE OPERON THEORY

Answer 10

worked out by F. jacobs and J. Monod on E.coli bacteria (a prokaryote) = THE LAC OPERON MODEL
(they put forward a hypothesis to explain why E.coli produced an enzyme galactosidase which breaks down lactose only when lactose is present)

Question 11

LAC OPERON MODEL / THE OPERON THEORY - steps (4)

Answer 11

1. One section of DNA that codes for the enzyme is called the STRUCTURAL GENE
2. situated close to this is another section of DNA called the (OPERATOR GENE) (this activates the structural gene when the enzyme is needed
3. the operator gene is controlled by the (REGULATOR GENE) situated further down the DNA chain - produces a REPRESSOR SUBSTANCE which turns the operator gene off ( = no enzyme is made = GENE REPRESSION)
4. When the enzyme is needed, the repressor substance is inhibited = operator gene can now turn on the structural gene and the enzyme is made = GENE INDUCTION

(it has been found that lactose itself can inactivate the repressor by combining with it)

Question 12

why is it important for cells to be able to control the production of certain enzymes?

Answer 12

control what is being produced = don’t make things we don’t use (+ wasting energy, space, resources)

Question 13

role of the promoter in the Lac Operon

Answer 13

= the promoter is used by the RNA polymerase to express the genes which synthesised lactase (BINDING POINT FOR RNA POLYMERASE)

Question 14

(when lactose is added to the E.coli environment discuss) what happens to start transcription of the lactose-utilization genes and why this is a benefit for cell such as E.coli

Answer 14

Lactose binds with the repressor - causing it to release from the operator and thus allowing the RNA polymerase to bind to the promoter and transcription can occur - it is a benefit as it stops the process from making unnecessary mRNA (process of DNA having mRNA taken from it)

Question 15

Epigenetics - basic breakdown

Answer 15

(scientifically not well understood)

understandings:
the environment of a cell and of an organism has an impact on gene expression

Epigenetics = is the study of environmental factors can turn genes on and off and affect how cells read genes
(differences in ‘traits’ that occur from the environment - there are ways that DNA can be changed after it has been replicated)

Question 16

The two types of epigenetics

Answer 16

1. DNA methylation
2. Histone modification

Question 17

DNA methylation - Epigenetics (mechanisms)

Answer 17

It is a process which methyl groups (-CH3) are added to the bases C and A in DNA - methylation modifies the function of the DNA (typically acting to suppress gene transcription - eg. preventing a cell from reverting to a stem cell or converting into a different cell type)

Question 18

Histone modification - Epigenetics (mechanisms)

Answer 18

DNA is wrapped around histones so alterations in the histone can affect the activity of DNA - Methylation of histones can promote or inhibit transcription

Modification of Histone Tails

Typically the histone tails have a positive charge and hence associate tightly with the negatively charged DNA

Adding an acetyl group to the tail (acetylation) neutralises the charge, making DNA less tightly coiled and increasing transcription
Adding a methyl group to the tail (methylation) maintains the positive charge, making DNA more coiled and reducing transcription

Question 19

Epigenetics research areas

Answer 19

• cancer
• developmental abnormalities (due to exposure to chemicals)
• trans-generational stress
• cardiovascular disease
• diabetes

Question 20

transcription vs translation

Answer 20

transcription:
- make RNA copies of individual genes
- produce mRNA, tRNA, rRNA, non-coding RNA
- 5’ cap adds to 3’ poly
- in the nucleus
- uses genes as templates to produce several functional form of RNA

translation:
- synthesis proteins that are used for cellular functions
- protein synthesis from mRNA template
- 2nd step of gene expressions
- rRNA = assembly plans, tRNA = translator for proteins
- produce proteins
- in the cytoplasm

Question 21

why is epigenetics important in cell specialisation

Answer 21

essential to controlling the heritable cellular memory of gene expression during development - research eg. cancer, developmental abnormalities due to exposure to chemicals….

Question 22

Skill: analysis of changes in the DNA methylation patterns

Answer 22

“during development the pattern of DNA methylation in the genome changes as a result of a dynamic process involving both de novo DNA methylation and demthylation - as a consequence, differentiated cells develop a stable and unique DNA methylation pattern that regulates tissue-specific gene transcription”

DNA methylation varies throughout a lifetime and can be affected by the enviornmal, age and lifestyle choices analysis of methylation patterns in a healthy cell and a cancerous cell can help identify disease marker genes. If we can identify cancer related genes then early diagnosis of the disease is more likely

Question 23

Outline two examples of trans generational epigenetics

Answer 23

Environmental impact:
Feast vs Famine
- in sweden males who had lived through good harvests had sons and grandsons with a higher rate of diabetes and heart disease while males who lived through poor harvests had sons and grandsons who lived on average 32 years longer

(DNA methylation controls transcription so the phenotypic (overvable characteristics = methylation patterns are different in healthy and diseased cells) expression of disease)

Question 24

Promoter - section of a gene

Answer 24

The non-coding sequence responsible for the initiation of transcription
The core promoter is typically located immediately upstream of the gene’s coding sequence
The promoter functions as a binding site for RNA polymerase (the enzyme responsible for transcription)
The binding of RNA polymerase to the promoter is mediated and controlled by an array of transcription factors in eukaryotes
These transcription factors bind to either proximal control elements (near the promoter) or distal control elements (at a distance)

Question 25

gene =

Answer 25

A gene is a sequence of DNA which is transcribed into RNA and contains three main parts

Question 26

Coding Sequence - section of a gene

Answer 26

After RNA polymerase has bound to the promoter, it causes the DNA strands to unwind and separate
The region of DNA that is transcribed by RNA polymerase is called the coding sequence

Question 27

Terminator - section of a gene

Answer 27

RNA polymerase will continue to transcribe the DNA until it reaches a terminator sequence
The mechanism for transcriptional termination differs between prokaryotes and eukaryotes

Question 28

Antisense vs Sense

Answer 28

A gene (DNA) consists of two polynucleotide strands, but only one is transcribed into RNA

The antisense strand is the strand that is transcribed into RNA
Its sequence is complementary to the RNA sequence and will be the “DNA version” of the tRNA anticodon sequence
The antisense strand is also referred to as the template strand

The sense strand is the strand that is not transcribed into RNA
Its sequence will be the “DNA version” of the RNA sequence (i.e. identical except for T instead of U)
The sense strand is also referred to as the coding strand (because it is a DNA copy of the RNA sequence)

Either of the 2 polynucleotide strands may contain a gene, and hence the determination of sense and antisense is gene specific

Question 29

Overview of Transcription

Answer 29

The process of transcription can be divided into three main steps: initiation, elongation and termination

Question 30

initiation - transcription

Answer 30

RNA polymerase binds to the promoter and causes the unwinding and separating of the DNA strands

Question 31

Elongation - transcription

Answer 31

occurs as the RNA polymerase moves along the coding sequence, synthesising RNA in a 5’ → 3’ direction

Question 32

termination - transcription

Answer 32

When RNA polymerase reaches the terminator, both the enzyme and nascent RNA strand detach and the DNA rewinds

Question 33

Messenger RNA

Answer 33

Understanding:
• Eukaryotic cells modify mRNA after transcription

In eukaryotes, there are three post-transcriptional events that must occur in order to form mature messenger RNA
1. Capping
2. Polyadenylation
3. Splicing

+ Post-Transcriptional Modifications

Question 34

post-transcriptional events to mature messenger RNA:
1. Capping

Answer 34

Capping involves the addition of a methyl group to the 5’-end of the transcribed RNA

The methylated cap provides protection against degradation by exonucleases

It also allows the transcript to be recognised by the cell’s translational machinery (e.g. nuclear export proteins and ribosome)

Question 35

post-transcriptional events to mature messenger RNA:
2. Polyadenylation

Answer 35

Polyadenylation describes the addition of a long chain of adenine nucleotides (a poly-A tail) to the 3’-end of the transcript

The poly-A tail improves the stability of the RNA transcript and facilitates its export from the nucleus

Question 36

post-transcriptional events to mature messenger RNA:
3. Splicing

Answer 36

Within eukaryotic genes are non-coding sequences called introns, which must be removed prior to forming mature mRNA

The coding regions are called exons and these are fused together when introns are removed to form a continuous sequence

Introns are intruding sequences whereas exons are expressing sequences

The process by which introns are removed is called splicing

Question 37

Gene expression (straight from bioninja)

Answer 37

Transcriptional activity is regulated by two groups of proteins that mediate binding of RNA polymerase to the promoter

Transcription factors form a complex with RNA polymerase at the promoter

RNA polymerase cannot initiate transcription without these factors and hence their levels regulate gene expression

Regulatory proteins bind to DNA sequences outside of the promoter and interact with the transcription factors

Activator proteins bind to enhancer sites and increase the rate of transcription (by mediating complex formation)
Repressor proteins bind to silencer sequences and decrease the rate of transcription (by preventing complex formation)

The presence of certain transcription factors or regulatory proteins may be tissue-specific

Additionally, chemical signals (e.g. hormones) can moderate protein levels and hence mediate a change in gene expression

Control Elements

The DNA sequences that regulatory proteins bind to are called control elements

Some control elements are located close to the promoter (proximal elements) while others are more distant (distal elements)
Regulatory proteins typically bind to distal control elements, whereas transcription factors usually bind to proximal elements
Most genes have multiple control elements and hence gene expression is a tightly controlled and coordinated process

Understanding:
• The environment of a cell and of an organism has an impact on gene expression

Changes in the external or internal environment can result in changes to gene expression patterns

Chemical signals within the cell can trigger changes in levels of regulatory proteins or transcription factors in response to stimuli
This allows gene expression to change in response to alterations in intracellular and extracellular conditions

There are a number of examples of organisms changing their gene expression patterns in response to environmental changes:

Hydrangeas change colour depending on the pH of the soil (acidic soil = blue flower ; alkaline soil = pink flower)
The Himalayan rabbit produces a different fur pigment depending on the temperature (>35ºC = white fur ; <30ºC = black fur)
Humans produce different amounts of melanin (skin pigment) depending on light exposure
Certain species of fish, reptile and amphibian can even change gender in response to social cues (e.g. mate availability)

Question 38

Types of Chromatin - Epigenetics

Answer 38

When DNA is supercoiled and not accessible for transcription, it exists as condensed heterochromatin

When the DNA is loosely packed and therefore accessible to the transcription machinery, it exists as euchromatin

Different cell types will have varying segments of DNA packaged as heterochromatin and euchromatin
Some segments of DNA may be permanently supercoiled, while other segments may change over the life cycle of the cell