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RELEVANT LEARNING OUTCOME:
(c) Describe the structure and function of non-coding DNA in eukaryotes (i.e. portions that do not encode protein or RNA, including introns, centromeres, telomeres, promoters, enhancers and silencers) (knowledge of transposons, satellite DNA, pseudo-genes and duplication of segments is not required).
(d) Describe the process of DNA replication and how the end replication problem arises.
❓Junk DNA❓
❓Junk DNA❓
Eukaryotic genome can be largely classified into 2 types of DNA: coding and non-coding.
This classification gives the false impression that coding is 'useful', while non-coding is 'not useful', or a commonly used word is 'Junk DNA', but is this really true?
Watch the video and consider the following:
Do these non-coding DNA serve a function, and is it right to describe them as 'Junk DNA'?
💡Structure and Function of non-coding DNA in Eukaryotes
💡Structure and Function of non-coding DNA in Eukaryotes
Non-coding DNA
Definition: sequence of DNA which does not code for protein or RNA (tRNA, rRNA).
In the syllabus, we will cover the following examples:
A. Centromere
B. Telomere
C. Promoter
D. Enhancers and Silencers
E. Introns
A. Centromere
A. Centromere
Watch the video from 3:40 to 4:17 to learn more about the centromere. Consider the following:
What is the role of centromeres between sister chromatids?
Why might the term 'centromere' be misleading?
Structure
Centromere are highly constricted (condensed) region of the chromosome
It consists of highly repetitive nucleotide sequences
note that diagram representation may make centromere look not part of the chromosome, but note that it is part of the DNA!
Function
Centromere is the site of binding for the kinetochore protein. (see Figure)
this is important for the binding of spindle fibers (to kinetochore) during cell cycle, ensuring that the chromosomes can be properly aligned along the metaphase plate.
Centromere also facilitate the joining of sister chromatids.
After DNA replication, identical sister chromatids are formed, and joined via the centromeres (facilitated by proteins)
During anaphase, spindle fibers shorten to pull apart sister chromatids.
Sister chromatids are then separated at the centromere region.
The last 2 functions are more for human applications (not a natural function required by the cells).
Different chromosomes have different size and also different centromere position, this allows for easy identification of different chromosomes (see Figure)
Having centromere provides a reference point on the chromosomes, which allows for mapping of genes on the chromosomes (each gene have a unique 'postal code', based on its position on the chromosome).
sister chromatids are joined together at their centromere region.
This is facilitated by proteins that binds to the centromere of each sister chromatid
centromere is not always at the centre of chromosomes, different chromosomes have different unique positions for the centromere.
B. Telomere
B. Telomere
Watch the video to gain an understanding of telomere. Consider the following questions while watching the video:
Where are telomeres found along the DNA?
What is the structure of telomeres in terms of nucleotide sequence and association with other biomolecules.
What happens to the telomeres after each round of cell division, and what is the significance of this, and how is it related to cancer?
Telomeres are found at the ends of the DNA molecule.
Telomeres consists of a repetitive nucleotide sequence, for examples, in humans the sequence 'TTAGGG' is repeated. Telomeres also associate with proteins called telomere proteins.
Telomeres shorten after each round of cell division. When telomeres become too short, the cell enters senescence, and stops dividing.
This prevents unlimited proliferation, and thereby prevents the cell from becoming cancerous.
Definition: A telomere is a region of repetitive short nucleotide sequences at each end of a eukaryotic linear chromosome.
Structure
consists of short repetitive sequences (Image A)
In humans, the sequence 'TTAGGG' is repeated
highly condensed (Image B):
this means that it is packed tightly (heterochromatin)
consists of a 3' overhang (Image C):
'3' overhang' refers to that the 3' ends of the telomere is longer than the 5' end.
This comes about due to the end-replication problem. (see below)
Function
Prevent the ends of chromosomes from being degraded by exonucleases.
Prevent ends of chromosomes from fusing.
Delay the degradation of genes.
Serve as a biological clock to determine the lifespan of a cell and the organism.
[Application] Provides a counting mechanism for the number of cell division a cell has undergone.
End-Replication Problem
Watch the following video to better understand the end-replication problem.
Consider the following:
What is the reason for the end-replication problem?
What is the impact of this problem?
What is the role of telomere in this problem?
DNA Polymerase III requires pre-existing 3' -OH ends for synthesis of DNA daughter strand.
However, when the last primer at the 5' end of the lagging strand is removed, there are no pre-existing 3' -OH ends.
This causes the DNA daughter strand to be shorter at the 5' -end, or commonly called a '3' - overhang', where the 3' end is longer than the 5' end (recall that DNA is anti-parallel). (refer to figure)
Telomeres being at the ends of chromosomes are the sequences that are lost due to end replication problem. They act as a buffer to protect the 'more important' coding regions (genes).
Telomerase
As the name suggests, telomerase is an enzyme that acts on telomere. Specifically to lengthen telomeres.
Recall from the video on telomeres, there are some cells like stem cells that needs to retain the ability to divide indefinitely. To do so, the cell needs to prevent telomere from shortening to the hayflick limit.
This is achieved by the activity of telomerase. Watch the video to gain an understanding of how this enzyme works, and note the following:
What are the components of telomerase?
Why is telomerase classified as a reverse transcriptase?
What are the steps that are repeated (in a cycle) in which telomerase lengthens telomeres?
Telomerase consists of:
Protein: which consists of the catalytic activity
Telomerase RNA: The RNA strand serves 2 functions:
Bind to existing telomeres at 3' overhang via complementary base paring
act as a template for the synthesis of DNA via complementary base pairing
Transcription is the synthesis of RNA from DNA. Telomerase is using RNA as a template to synthesise DNA, thus the process is termed as reverse transcription, and enzymes that catalyses this process are reverse transcriptase.
The synthesis of telomere by telomerase can be viewed as a cycle consisting of 3 repeated steps (refer to Figure):
(a) Binding: Telomerase binds to telomere 3' overhang using telomerase RNA via complementary base pairing.
(b) Reverse transcription: Telomerase RNA acts as a template for telomerase to catalyse the synthesis of DNA via complementary base pairing
(c) Translocation: Telomerase detaches and moves down the newly lengthened 3' overhang.
Telomerase then binds to the newly legnthened 3' overhang via complementary base pairing, repeating the cycle.
(d) The lengthening of the 3' overhang, which acts the template for the synthesis of the daughter strand:
more RNA primers can now be placed
DNA pol III can then lengthen the daughter strand
results in both strand being lengthened
Do note that the end replication problem will still exist! There will still be a 3' overhang at the end, however, overall the telomere (both strands) have been lengthened.
The following 3 examples (promoter, enhancer, and silencer) are classified as control elements.
Control: These non-coding DNA do not code for any protein / RNA, but instead, they control the rate of transcription.
Element: indicates that these are DNA in nature.
C. Promoter
C. Promoter
Watch the video to gain an understanding of promoters and enhancers. Take note of the following:
Where are promoters and enhancers located?
What is the unique sequence found in promoters?
What are the function of promoters and enhancers?
Structure
In eukaryotes, promoters are found at the start of each gene.
Promoter region usually contain a AT-rich region (adenine-thymine), called the TATA box. E.g., TATAAA (see Figure)
Function
Promoter region is recognised and bound by general transcription factors and RNA polymerase.
This allows for the formation of the transcription initiation complex, which is required for transcription to start.
D. Enhancers and Silencers
D. Enhancers and Silencers
Structure
Enhancers and silencers are considered distal control elements.
Distal refers to that they are located far from the gene that it regulates, it can be upstream (before), downstream (after) or even within an intron of the gene that it controls.
There are no generic sequence of enhancers and silencers like that of promoter.
Different enhancers and silencers of different genes can have different nucleotide sequences.
Function
Enhancers: Are bound by activators, which leads to an increase rate of transcription.
Silencers: Are bound by repressors, which leads ot a decrease rate of transcription.
E. Introns
E. Introns
Introns was covered under post transcription modification in DNA Genomics II.
Watch the video to gain a deeper understanding of introns. Consider the following:
What is the purpose of having introns if it is just going to be spliced out after transcription?
Structure
Introns are interspersed between exons (coding regions), which are transcribed and are part of pre-mRNA.
Introns also do not have a specific sequence like promoters.
However, there are unique sequences are the ends of introns, which allows spliceosomes to recognise introns.
Function
The presence of introns causes the need of splicing.
Having to perform splicing allows for alternative RNA splicing to occur.
Alternative RNA splicing allows for one gene to form different mature mRNA, consisting of different combinations of exons, leading to the synthesis of different forms of the protein (see Figure).
🖊️Check Your Understanding: Lecture Notes Checkpoint
🖊️Check Your Understanding: Lecture Notes Checkpoint
Which eukaryotic non-coding DNA may have repeating sequences?
1. Centromeres
2. Introns
3. Telomeres
A. 1 only
B. 3 only
C. 2 and 3 only
D. 1, 2 and 3
Answer: (D) - Centromeres and Telomeres have specific repeating sequnces. Introns can also contain short repetitive sequences.
2. The diagram shows the classification of several portions of eukaryotic DNA that do not encode RNA or protein.
Which combination correctly identifies the type of eukaryotic DNA?
Answer: (B) - Out of the 5 types of non-coding DNA (centromere, intron, telomere, enhancer and 5' UTR),
2 are transcribed into pre-mRNA: introns and 5' UTR (untranslated region)
introns contain sequences that regulates splicing --> Y
5' UTR faciliates binding of small ribosomal subunits during translation --> Z
3 are never transcribed: centromere, telomere and enhancer
sister chromatids (each a DNA double helix) are joined via centromeres --> V
enhancers binds to activators to upregulate transcription --> W
telomeres binds to telomere capping proteins that protects the ends of DNA from degradation --> X
🖊️Check Your Understanding
🖊️Check Your Understanding
Attempt qns 1-16 of "Section 1: Organisation of Eukaryotic Genome" in this SLS lesson.
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