UNIT 6 – Organisation of Gene (Q&A) | MZO-002 | MSCZOO | M.Sc. Zoology | IGNOU
SAQ 1
i) Fill in the blanks:
(a) The organisation of genes in prokaryotes is ....................... .
Answer: simple
(b) A series of genes involved in a particular metabolic pathway arranged together form a functional group called ...................... .
Answer: operon
(c) ....................... is the sequence of nucleotides that defines the direction of transcription in the gene.
Answer: the TATA box
(d) The short sequence of DNA usually present in the upstream region of the gene often within 200 bp of the transcription start site is .......................... .
Answer: promoter
(e) ........................ are the genes that have lost their ability for expression or code protein.
Answer: Pseudogenes
ii. Answer in one word:
a) Coding sequences present in a genome.
Answer: Exons
b) Sequences that were discovered by Phillip Sharp and Richard Roberts during studies of the replication of adenovirus in cultured human cells.
Answer: Introns
c) Nucleotide sequences that enhance the transcription of the associated gene when bound by specific transcription factors.
Answer: Enhancers
d) An area that contains a stretch of codons and has the potential to code for protein/peptide.
Answer: Open reading frame (ORF)
e) The sequence consists of a series of conserved combinations of nucleotides in eukaryotes.
Answer: Kozak sequence
SAQ 2
Fill in the blanks:
a) ....................... are the proteins that bind specific DNA sequences and regulate gene expression.
Answer: Transcription factors (TFs)
b) Small regulatory non-coding RNAs that influence gene regulation are called ........................ .
Answer: microRNAs (miRNAs)
c) The specific domain in transcription factors that bind to DNA are .......................... .
Answer: DNA binding domain (DBD)
d) During regulation of gene activity TFs bind to specific short DNA base pair patterns in upstream, intron, or downstream regions of target genes called ........................ .
Answer: motifs or cis-regulatory elements (CREs)
e) ........................ transcription factor is involved in recognizing the promoter sequence (TATA box).
Answer: TFIID
SAQ 3
State whether the statements are 'True' or 'False'
a) Enhancers can regulate the expression of genes that can be located relatively far away in the DNA molecule.
Answer: True
b) Small Nucleolar RNA (snoRNA) present in the nucleoli aid in the addition of methyl groups to the rRNA after assembly of the rRNA into the ribosomes.
Answer: False
c) tRNA ensures that the correct amino acid is attached to the growing polypeptide chain according to the sequence specified by the mRNA that is being translated.
Answer: True
d) Negative Regulatory Elements mediate the repression of transcription by an active mechanism.
Answer: False
e) iRNA and miRNA regulate gene expression by attaching to the mRNA of those genes (via base pairing) and subsequently causing the mRNAs to be degraded resulting in the termination of the expression of the genes.
Answer: True
SAQ 4
Write the full form of the abbreviations:
a) CpG
Answer: Cytosine-phosphatidyl-Guanine
b) DNMTs
Answer: DNA menthytransferases
c) HATs
Answer: Histone acetyltransferases
d) HDACs
Answer: Histon deacetylases
TERMINAL QUESTIONS
1. How is the organisation of genes different between prokaryotes and eukaryotes?
The organisation of genes in prokaryotes and eukaryotes is quite different due to their structural, functional and evolutionary differences. These differences are seen in the way genes are arranged on the DNA, how they are regulated and how they are transcribed and translated. Below is a detailed explanation of their gene organisation based on major points:
1. Arrangement of Genes
Prokaryotes:
Genes are often arranged in clusters called operons. An operon is a group of genes under the control of a single promoter and transcribed together as one mRNA. These genes usually have related functions. For example, the lac operon in E. coli includes genes required for lactose metabolism.
Eukaryotes:
Genes are usually arranged individually. Each gene has its own promoter, enhancer and regulatory elements. Eukaryotic genes are not usually grouped by function. They are transcribed separately into different mRNAs.
2. Coding and Non-coding Regions
Prokaryotes:
Their genes are mostly made of exons (coding sequences). There are very few introns (non-coding regions) in prokaryotic genes. So the transcription and translation processes are direct and fast.
Eukaryotes:
Most genes contain both exons (coding sequences) and introns (non-coding sequences). After transcription, the primary mRNA undergoes splicing to remove introns before translation. This adds complexity and regulation to gene expression.
3. Regulation of Genes
Prokaryotes:
Gene regulation is simpler and often at the level of transcription initiation. For example, the presence or absence of a substrate (like lactose) can switch genes on or off.
Eukaryotes:
Gene regulation is more complex. It happens at multiple levels, transcription, post-transcription, translation and post-translation. Regulatory sequences like enhancers and silencers may be located far from the gene they regulate. Proteins called transcription factors and epigenetic modifications (like methylation) also play important roles.
4. Transcription and Translation Location
Prokaryotes:
Transcription and translation occur in the same compartment (cytoplasm), often at the same time. This is called coupled transcription and translation.
Eukaryotes:
Transcription takes place in the nucleus and translation occurs in the cytoplasm. The mRNA must first be processed and exported from the nucleus.
5. Gene Copy Number and Complexity
Prokaryotes:
Generally have a single circular chromosome with fewer genes. The genome is small and compact with minimal non-coding DNA.
Eukaryotes:
Have multiple linear chromosomes. Their genome is large and contains a high amount of non-coding DNA, including repetitive sequences, pseudogenes and regulatory elements.
6. Histones and Chromatin Structure
Prokaryotes:
Do not have histones (with few exceptions in Archaea). Their DNA is not wrapped in chromatin but exists as a simple nucleoid.
Eukaryotes:
DNA is tightly packed with histone proteins to form chromatin. The organisation into euchromatin and heterochromatin controls gene accessibility and expression.
2. What are the regulatory sequences of a typical eukaryotic gene? Give examples.
In eukaryotic cells, gene expression is highly controlled by specific regulatory DNA sequences. These sequences do not code for proteins, but they decide when, where and how much a gene should be expressed. They mainly control the process of transcription, where RNA is made from DNA. A typical eukaryotic gene contains several important regulatory sequences, such as:
- Promoters
- Enhancers
- Silencers
- Insulators
- Response elements
1. Promoter
The promoter is the main regulatory region of a gene. It lies just before the transcription start site, which is the point where RNA starts getting made. This is the site where RNA polymerase and transcription factors attach to begin transcription. One well-known part of the promoter is the TATA box, found around 25 to 35 base pairs before the start site. It helps RNA polymerase find the correct place to begin. Another element, the Initiator (Inr) sequence, is present near the +1 position and supports proper transcription initiation.
Example: Housekeeping genes usually have strong promoters with clear TATA boxes to ensure continuous activity.
2. Enhancer
Enhancers are DNA elements that boost the level of transcription. They can be located far before or after the gene, or even inside introns. Enhancers bind to activator proteins, which help RNA polymerase work more efficiently. Sometimes, the DNA loops to bring the enhancer close to the promoter.
Example: The β-globin gene enhancer controls the gene's activity in red blood cells and is located thousands of base pairs away from the gene.
3. Silencer
Silencers are opposite to enhancers. They reduce or completely block transcription. They work by binding to repressor proteins, which prevent RNA polymerase from working properly. Silencers also act from a distance and can be present in different locations relative to the gene.
Example: The NRSE silencer prevents neuronal genes from being expressed in non-neuronal cells.
4. Insulator
Insulators serve as barriers. They stop enhancers from activating the wrong gene and also prevent the spread of silencing effects. This helps in keeping nearby genes regulated independently.
Example: The CTCF-binding site in the H19/Igf2 region is important for gene boundary control during genomic imprinting.
5. Response Elements
Response elements allow genes to respond to external or internal signals like hormones or stress. These elements ensure that genes are expressed only under the right conditions.
Examples:
- Estrogen Response Element (ERE) activates estrogen-responsive genes.
- Heat Shock Element (HSE) helps turn on genes during heat stress.
3. Describe the components of the promoter region of a eukaryotic gene.
In eukaryotic genes, the promoter region is a special stretch of DNA that lies just before the gene. Its main job is to control when and where transcription starts. It does this by providing binding sites for RNA polymerase II and other transcription factors.
The promoter region can be divided into two main parts:
- Core Promoter
- Proximal Promoter Elements
Each of these parts has different types of DNA sequences that help start and regulate transcription.
1. Core Promoter
This is the most essential part of the promoter and lies near the transcription start site (called the +1 position). It directly helps in assembling the transcription machinery. The core promoter usually includes the following elements:
- TATA Box:
- A short DNA sequence (TATAAA) usually located 25–35 base pairs upstream from +1 site. It helps in positioning RNA polymerase II. A special protein called TBP (TATA-binding protein) binds here to start the transcription complex.
- Initiator (Inr) Sequence:
- Found around the +1 site. It helps start transcription at the correct location. Some promoters have Inr even if they do not have a TATA box.
- BRE (TFIIB Recognition Element):
- Found either just upstream or downstream of the TATA box. It binds the TFIIB protein which supports recruitment of RNA polymerase II.
- DPE (Downstream Promoter Element):
- Found downstream of the transcription start site (usually at +28 to +32). Especially useful in promoters that lack a TATA box.
2. Proximal Promoter Elements
These are found a little further upstream of the core promoter. They help enhance the level of transcription by binding extra transcription factors.
- CAAT Box:
- Located around 75 base pairs upstream. It binds proteins like NF-Y which increase transcription strength.
- GC Box:
- A region rich in guanine and cytosine. It binds the Sp1 protein and supports basic transcription activity. These are often present in housekeeping genes.
4. What are transcription factors? Describe the different categories of transcription factors.
Transcription factors are special types of proteins that control the process of transcription in eukaryotic cells. Transcription is the first step of gene expression where the DNA is copied into RNA. These factors do not make RNA directly but help the enzyme RNA polymerase to start, stop or control the speed of this process. Transcription factors help the cell know when to turn a gene ON, when to turn it OFF and how much product the gene should make. They play a very important role in development, cell cycle, cell differentiation, stress response and hormone signaling.
Transcription factors are mainly divided into two broad categories:
- Based on their function
- Based on their structure and DNA-binding domain
1. Types Based on Their Function
There are the following two types:
a) General Transcription Factors
These transcription factors are needed for the transcription of almost all protein-coding genes. They help in forming the transcription initiation complex near the promoter region. They guide the RNA polymerase II to attach at the correct place and begin transcription. These factors are not gene-specific.
Example: TFIID is a general transcription factor. It has a part called TATA-binding protein (TBP) which helps in recognizing the TATA box of the promoter.
b) Specific Transcription Factors
These transcription factors control only certain genes. They work in special conditions such as during stress, hormone action, or development. They bind to enhancer or silencer sequences and either activate or block the transcription of target genes. They are gene-specific and time-specific.
Example: Estrogen receptor is a specific transcription factor that activates genes in the presence of estrogen hormone.
2. Types Based on Their Structure and DNA-Binding Domain
There are the following four major types:
a) Helix-Turn-Helix (HTH)
This type has two alpha helices connected by a short loop or turn. One of the helices fits into the major groove of DNA and helps in DNA binding.
Example: Hox proteins, which are important for body pattern development in animals, are of this type.
b) Zinc Finger
These proteins have finger-like loops that are held together by zinc ions. These loops bind to DNA in a specific way and are very common in eukaryotic transcription factors.
Example: TFIIIA in Xenopus (a frog) was the first discovered zinc finger protein.
c) Leucine Zipper
These proteins have a leucine amino acid at every 7th position which helps in forming a dimer. Two such proteins come together to hold the DNA tightly, like a zip and help in gene regulation.
Example: c-Fos and c-Jun proteins form the AP-1 complex which controls cell growth and division.
d) Basic Helix-Loop-Helix (bHLH)
This type has two helices connected by a loop. They usually form dimers and bind to specific DNA sequences called E-boxes. These are common in developmental genes.
Example: MyoD is a bHLH transcription factor involved in muscle development.
5. What are the differences between gene enhancers and gene silencers? How do enhancers and silencers regulate eukaryotic gene expression?
Enhancers and silencers are two important types of regulatory DNA sequences that play opposite roles in controlling gene expression in eukaryotic cells. They are non-coding DNA elements which do not produce proteins but control when, where and how much a gene is expressed. They work by interacting with transcription factors and RNA polymerase to either increase or decrease the level of transcription.
Differences Between Gene Enhancers and Gene Silencers
Gene enhancers and gene silencers are both regulatory DNA elements found in eukaryotic genomes. Their main role is to control the level of gene expression, but they work in opposite directions. The differences between them can be explained based on the following criteria:
1. Based on Function
Enhancers increase the transcription of a gene. They make the gene more active and allow it to produce more RNA.
On the other hand, silencers reduce or completely block transcription. They stop or decrease the activity of the gene.
2. Based on the Type of Proteins That Bind
Enhancers work by binding with special proteins called activators. These activators help RNA polymerase and other transcription machinery to work more effectively.
In contrast, silencers bind with repressor proteins. These repressors block the action of RNA polymerase and reduce gene activity.
3. Based on Effect on Chromatin
Enhancers often promote open chromatin structure (euchromatin), which is easy for transcription machinery to access.
Silencers promote closed chromatin (heterochromatin), which makes the DNA less accessible and harder for transcription to happen.
4. Based on Impact on Cell-Specific Gene Expression
Enhancers help activate genes in specific cell types or under specific conditions. For example, they may turn on a gene in liver cells but not in brain cells.
Silencers help stop a gene from being expressed in the wrong cells. For example, a neuronal gene may be silenced in muscle cells by a silencer.
How Enhancers and Silencers Regulate Eukaryotic Gene Expression
There are two ways by which enhancers and silencers regulate gene expression:
A) By Binding to Specific Proteins
Enhancers bind with activator transcription factors, while silencers bind with repressor transcription factors. These protein–DNA interactions either support or block the binding of RNA polymerase to the promoter region. This decides whether transcription will happen or not.
- In case of enhancers, the activator proteins help to recruit RNA polymerase II and other general transcription factors to the promoter.
- In case of silencers, the repressor proteins block the recruitment of RNA polymerase or prevent the formation of transcription initiation complex.
B) By DNA Looping Mechanism
Even if these elements are far from the gene, DNA can loop to bring them near the promoter. This looping is supported by proteins like mediator complex and architectural proteins. This way, enhancers can activate and silencers can suppress the transcription machinery even from a distance.
So, enhancers and silencers act like gene switches that help the cell decide which genes to turn ON and which to keep OFF, depending on the cell type, environmental signals or stage of development.
6. What are non-coding genes? Give examples.
Non-coding genes are segments of DNA that do not code for any protein but still play very important roles in the cell. These genes are transcribed into functional RNA molecules instead of mRNA. These RNAs do not get translated into proteins but perform regulatory, structural, or catalytic roles directly as RNA. Non-coding genes are a major part of the eukaryotic genome and help in many cellular processes such as gene regulation, RNA processing and maintaining genome stability.
Many people think that only protein-coding genes matter, but non-coding genes also play a crucial role in regulating cellular activities. They help control when and how genes are turned on or off, assist in processing other RNAs and participate in building cell machinery like ribosomes and spliceosomes.
There are several important types of non-coding genes, which can be grouped based on the RNA they produce and their functions. The main categories are:
1. rRNA Genes (Ribosomal RNA Genes)
These genes produce ribosomal RNAs, which are core components of ribosomes. Ribosomes are the molecular machines that make proteins. rRNAs form the structural and catalytic parts of ribosomes, helping in protein synthesis by linking amino acids in the right order. Without rRNAs, ribosomes cannot assemble properly or carry out their function of translating genetic information into proteins.
Example:
- 18S, 5.8S and 28S rRNA genes transcribed by RNA Polymerase I.
- 5S rRNA transcribed by RNA Polymerase III.
2. tRNA Genes (Transfer RNA Genes)
tRNA genes make transfer RNAs, which bring amino acids to the ribosome during protein production. Each tRNA matches a specific mRNA codon with the right amino acid, ensuring the protein is made correctly. tRNAs have a unique three-dimensional structure that allows them to fit perfectly in the ribosome's active site during translation.
Example:
- tRNAᴾʰᵉ (for phenylalanine)
- tRNAᴸᵉᵘLeu (for leucine)
3. snRNA Genes (Small Nuclear RNA Genes)
snRNAs are part of the spliceosome complex that removes non-coding sequences called introns from pre-mRNA. This process is essential for making mature mRNA that can be translated into protein. snRNAs recognize specific sites on the pre-mRNA and help cut and join the exons together in the right order.
Example:
- U1, U2, U4, U5, U6 snRNAs.
4. snoRNA Genes (Small Nucleolar RNA Genes)
snoRNAs are located in the nucleolus and guide chemical modifications on rRNA, such as methylation and pseudouridylation, which are important for rRNA stability and function. These modifications help ribosomes maintain their shape and carry out protein synthesis efficiently.
Example:
- U3 snoRNA is important for processing 18S rRNA.
5. miRNA Genes (MicroRNA Genes)
MicroRNAs regulate gene expression by binding to messenger RNAs (mRNAs), preventing their translation into proteins or causing their breakdown. This process allows cells to control the amount of specific proteins, which is important for development, cell growth and disease prevention.
Example:
- miR-21 regulates genes related to cell growth and cancer.
- miR-155 is important in immune response.
6. lncRNA Genes (Long Non-Coding RNA Genes)
Long non-coding RNAs are more than 200 nucleotides long and regulate gene activity in many ways, such as changing chromatin structure, controlling transcription and influencing RNA stability. They can act in the nucleus or cytoplasm and are involved in processes like X-chromosome inactivation and gene silencing.
Example:
- XIST lncRNA controls X-chromosome inactivation in females.
- HOTAIR lncRNA is involved in modifying chromatin to silence genes.
7. What are epigenetic modifications? Give examples.
Epigenetic modifications are heritable and reversible changes in gene expression that occur without altering the DNA sequence itself. These changes play a major role in how genes are turned on or off in different cells and at different times. Epigenetics helps explain how the same DNA sequence can produce different types of cells like skin cells, nerve cells and liver cells in the same organism. These modifications are very important during development, cellular differentiation, X-chromosome inactivation in females, genomic imprinting, aging and in many diseases such as cancer.
There are the following three main types of epigenetic modifications:
1. DNA Methylation
This is the most studied and well-understood form of epigenetic modification. In this process, a methyl group (–CH₃) is added to the cytosine base in DNA, mainly at CpG dinucleotides. These CpG regions are often found in clusters called CpG islands, which are located near gene promoters. DNA methylation is done by enzymes called DNA methyltransferases (DNMTs). When methylation happens in the promoter region of a gene, it blocks the binding of transcription factors and RNA polymerase, leading to the silencing of that gene.
Example: In cancer cells, tumor suppressor genes like p16 or BRCA1 are often found to be silenced due to abnormal hypermethylation. In female mammals, one of the two X chromosomes is inactivated by heavy methylation to balance gene dosage between males and females.
2. Histone Modifications
DNA is wrapped around proteins called histones to form a structure known as chromatin. The N-terminal tails of histones can undergo various chemical modifications such as acetylation, methylation, phosphorylation, ubiquitination and sumoylation. These modifications affect how tightly or loosely the DNA is wrapped, which in turn controls gene accessibility.
- Histone Acetylation: Addition of acetyl groups by Histone Acetyl Transferases (HATs) loosens chromatin and increases transcription.
- Histone Deacetylation: Removal of acetyl groups by Histone Deacetylases (HDACs) tightens chromatin and represses transcription.
- Histone Methylation: Addition of methyl groups can either activate or repress transcription depending on the location and type of histone.
Example: Acetylation at H3K27 (histone 3, lysine 27) is associated with gene activation, while methylation at the same site (H3K27me3) is associated with gene silencing.
3. Non-coding RNAs (ncRNAs)
These are RNA molecules that do not code for proteins but help in regulating gene expression at transcriptional and post-transcriptional levels. They include microRNAs (miRNAs), long non-coding RNAs (lncRNAs) and small interfering RNAs (siRNAs). ncRNAs can bind to mRNA to block translation or cause degradation. Some ncRNAs also help in recruiting histone-modifying complexes to specific gene regions.
Example: The long non-coding RNA Xist is essential for X-chromosome inactivation in females. It spreads across the inactive X chromosome and attracts proteins that silence gene expression on that chromosome.
8. How can DNA methylation regulate gene expression in eukaryotes?
DNA methylation is a key epigenetic mechanism used by eukaryotic cells to control gene expression without changing the actual DNA sequence. It mainly involves the addition of a methyl group (–CH₃) to the 5th carbon of cytosine, especially at CpG (Cytosine phosphate Guanine) dinucleotides (where cytosine is followed by guanine). This chemical change plays an important role in silencing genes and maintaining cell identity.
Gene regulation through DNA methylation happens by the following interconnected processes:
1. Blocking of transcription by promoter methylation
Most genes have CpG (Cytosine phosphate Guanine) islands in their promoter regions. When these CpG islands are unmethylated, transcription factors and RNA polymerase can bind easily and gene expression takes place. But if these CpG islands become methylated, they physically block the access of transcription factors to the promoter. As a result, the gene cannot be transcribed and gene expression is turned off.
2. Recruitment of methylation-specific binding proteins
Methylated DNA is recognized by special proteins known as methyl-CpG-binding proteins (like MeCP2). These proteins do not act alone. Once they bind to methylated regions, they recruit co-repressor proteins and enzymes that modify histones. This builds up a larger complex that causes long-term gene silencing.
3. Formation of repressive chromatin structure
When methyl-binding proteins are in place, they attract histone deacetylases (HDACs) and other chromatin remodeling enzymes. HDACs remove acetyl groups from histones, which causes the chromatin to shift from a loose (euchromatin) state to a tight (heterochromatin) form. In this compact state, transcription machinery cannot reach the DNA, so the gene remains inactive.
4. Stable and heritable gene silencing
DNA methylation is not just a temporary signal. After cell division, maintenance DNA methyltransferases (like DNMT1) copy the methylation pattern onto the new DNA strand. This ensures that the gene stays silent in daughter cells also. This feature is especially important during development, cell differentiation and X-chromosome inactivation.
5. Silencing of unwanted DNA elements
Besides regulating normal genes, DNA methylation also suppresses transposable elements and repetitive sequences in the genome. This protects the genome from harmful mutations. Abnormal methylation patterns are also seen in diseases like cancer, where tumor suppressor genes may get methylated and silenced.
Comments
Post a Comment