Prokaryotic and Eukaryotic RNA Polymerase

Introduction

Ribonucleic Acid (RNA) Polymerase (RNAP) enzyme is a multi-subunit enzyme that applies its activity in the catalyzation of the transcription process of RNA synthesized from a DNA template.

RNA polymerase is a key enzyme involved in the process of transcription, which is a fundamental step in gene expression. Transcription is the process by which genetic information encoded in DNA is copied into RNA molecules, such as messenger RNA (mRNA), ribosomal RNA (rRNA), and transfer RNA (tRNA). RNA polymerase plays a central role in this process by synthesizing RNA molecules based on the information contained in DNA.

• And therefore, RNA polymerase enzyme is responsible for the copying of DNA sequences into RNA sequences during transcription.
• The function of RNA polymerase is to control the process of transcription, through which copying of information stored in DNA into a new molecule of messenger RNA (mRNA.)
• During transcription, the RNA polymer is contemporary to the template DNA that is synthesized in the direction of 5′ to 3′.
• The enzyme RNA polymerase interacts with proteins to enable it to function in catalyzation of the synthesis of RNA.
• The collaborator proteins assist in enabling the specific binding of RNA polymerase, assist in the unwinding of the double chemical structure of DNA, moderate the enzymatic activities of RNA polymerase and to control the speed of transcription.
• The RNA polymerase enzyme has an interrupted mechanism whereby it continuously synthesizes RNA polymers of over four thousand bases per minute but they pause or stop occasionally to maintain fidelity.
• RNA polymerase is an enzyme that is responsible for copying a DNA sequence into an RNA sequence, during the process of transcription. As a complex molecule composed of protein subunits, RNA polymerase controls the process of transcription, during which the information stored in a molecule of DNA is copied into a new molecule of messenger RNA.
• RNA polymerases have been found in all species, but the number and composition of these proteins vary across taxa.
• For instance, bacteria contain a single type of RNA polymerase, while eukaryotes (multicellular organisms and yeasts) contain three distinct types.
• In spite of these differences, there are striking similarities among transcriptional mechanisms.
• For example, all species require a mechanism by which transcription can be regulated in order to achieve spatial and temporal changes in gene expression.

Types of RNA polymerase

Prokaryotic (Bacteria, viruses, archaea) organisms have a single type of RNA polymerase that synthesizes all the subtypes of RNA, while eukaryotes (multicellular organisms) have 5 different types of RNA polymerases which perform different functions in the synthesis of different RNA molecules.

Prokaryotic RNA polymerase

• The prokaryotes have a single type of RNA polymerase (RNAP) which synthesizes all the classes of RNA, i.e mRNA, tRNA, rRNA, sRNA.
• The RNA Polymerase molecule is made up of 2 domains and 5 subunits:

1. Core and holoenzyme
2. Subunits (β, β’, α (αI and αII), ω,)

• The promoter is the sequence of DNA that is required for accurate and specific initiation of transcription, and also, it is the sequence of DNA to which RNA polymerase binds accurately to initiate transcription.
• The ‘a’ subunit is made up of two distinct domains. The N-terminal domain (a-NTD) and the C-terminal.
• The N-terminal is involved in dimerization forming a2 and further assembly of the RNA polymerase.
• The C-terminal domain functions such as binding to the Upstream Promoter (UP) DNA sequence at promoters for rRNA and tRNA genes and in communication with several transcriptional activators.
• Each of the subunit structure is as follows:

Prokaryotic RNA Polymerase Subunits

Eukaryotic RNA polymerase

• There are 5 known types of RNA polymerases each responsible for the synthesis of specific subtypes of RNA. These include:
• RNA polymerase I that synthesizes a pre-rRNA 45S (35S in yeast), which matures and forms the major RNA sections of the ribosome.
• RNA polymerase II synthesizes precursors of mRNAs and most snRNA and microRNAs.
• RNA polymerase III synthesizes tRNAs, rRNA 5S, and other small RNAs found in the nucleus and cytosol.
• RNA polymerase IV and V found in plants are not well understood, however, they make siRNA. The plant chloroplast encodes the ssRNAPs and uses bacteria-like RNA Polymerase.
• Each of the nuclear RNA polymerases is a large protein molecule with about 8 to 14 subunits and the molecular weight is approximately 500,000 for each.
• They commonly have 3 subunits, a, b and b’. The largest subunits being b and b’.
• These subunits are used as catalytic promoters and for assembly of proteins.
• Each of these polymerases has a different function:

RNA polymerase I

• This enzyme is located in the nucleolus of the cell.
• It is a specialized nuclear substructure where the ribosomal RNA (rRNA) is synthesized by transcription and assembled into ribosomes.
• The rRNA are component elements of the ribosomes and are important in the process of translation.
• Therefore, RNA polymerase I synthesize almost all rRNAs except 5S rRNA.
• In yeast, the enzyme has a mass of 600kDa and 13 subunits.

RNA polymerase II

• This enzyme is located in the nucleus.
• Most organisms that possess RNA polymerase II have a 12-subunit RNAP II (with a mass of about 550 kDa)
• It is structurally made up of holoenzyme and mediators, with General Transcriptional factors (GTFs).
• They contain transcription factors and transcriptional regulators.
• It functions by synthesizing all proteins that code for the nuclear pre-mRNAs in eukaryotic cells (mRNAs in prokaryotic cells).
• It is responsible for transcribing most of the eukaryotic genes and especially found in human genes.

RNA polymerase III

• It is located in the nucleus.
• The RNA polymerase III has 14 or more distinct subunits with a mass of approximately 700 kDa.
• Its function is to transcribe transfer RNA (tRNA), ribosomal RNA (rRNA), and other small RNAs.
• Some of its target points are important for the normal functioning of the cell

RNA polymerases IV and V

• They are exclusively found in plants, and they perform combined action in the formation of small interfering RNA and heterochromatin in the cell nucleus.
• In Plants, the RNA polymerase is found in the chloroplast (plastids) and mitochondria, encoded by the mitochondrial DNA.
• These enzymes are much more related to bacterial RNA polymerase than to the nuclear RNA polymerase.
• Their function is to catalyze specific transcription of organelle genes.

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Functions of RNA Polymerase

• Generally,  the RNA molecule is a messenger molecule that is used to export information that is coded in DNA out of the cell nucleus, to synthesize proteins in the cell cytoplasm.
• RNA polymerase is used in the production of molecules that play a wide range of roles, of which one of its functions is to regulate the number and type of RNA transcript that is formed in response to the requirements of the cell.
• The RNA polymerase enzyme interacts with different molecular proteins, transcription factors, and signaling molecules on the carboxyl-terminal, which regulates its mechanisms, which play a major role in gene expression and gene specialization in multicellular (eukaryotic) organisms.
• The RNA enzyme also ensures irregularities and errors during the conversion of DNA to RNA (transcription). Such as ensuring that the right nucleotide is added to the newly synthesized RNA strand, inserting the right amino acid-base which is complementary to the template of the DNA strand.
• When the right nucleotides have been inserted, the RNA polymerase can then catalyze and elongate the RNA strand, at the same time, proofread the new strand and remove incorrect bases.
• RNA polymerase is also involved in the post-transcription modification of RNAs, converting them into functional molecules that facilitate the transportation of molecules from the nucleus to their site of action.
• Besides its role in the synthesis of proteins, RNA performs other functions such as
• Protein coding
• Regulation of gene expression
• Act as enzymes
• Formation of gametes by the non-coding RNA (ncRNA)
• Production of regulatory molecules.

Advantages of RNA Polymerase

RNA polymerases have several advantages and unique features that make them essential enzymes in the process of transcription and gene expression. Here are some of the advantages of RNA polymerases:

• Specificity: RNA polymerases are highly specific enzymes. Each type of RNA polymerase (e.g., RNA polymerase I, II, and III in eukaryotes) is responsible for transcribing a specific class of RNA molecules, ensuring accurate synthesis of different types of RNA, such as mRNA, rRNA, and tRNA.
• Versatility: RNA polymerases are versatile enzymes capable of synthesizing RNA from DNA templates with high fidelity. They can initiate transcription at specific promoter sequences and elongate RNA molecules, producing single-stranded RNA transcripts.
• Template-Directed Synthesis: RNA polymerases read the DNA template strand and synthesize RNA molecules in a template-directed manner, ensuring that the RNA sequence is complementary to the DNA template. This fidelity is crucial for the accurate transfer of genetic information from DNA to RNA.
• Regulation: RNA polymerases are subject to tight regulation, allowing cells to control gene expression. Various transcription factors, activators, and repressors influence the activity of RNA polymerases, enabling precise control of when and where genes are transcribed.
• Dynamic Processivity: RNA polymerases can remain associated with the DNA template for extended periods during transcription, allowing for the synthesis of long RNA transcripts. This dynamic processivity is important for the efficient transcription of genes that span thousands of nucleotides.
• Termination Mechanisms: RNA polymerases use different termination mechanisms to release the RNA transcript at the appropriate location, depending on the type of polymerase and organism. These mechanisms ensure that RNA synthesis ends accurately.
• RNA Processing: RNA polymerases produce primary transcripts that often require additional processing steps, such as capping, splicing, and polyadenylation, before they become mature, functional RNAs. This post-transcriptional processing allows for the production of diverse and functional RNA molecules.
• Promoter Recognition: RNA polymerases recognize and bind to specific promoter sequences in the DNA, ensuring that transcription initiates at the correct location within a gene. Promoter recognition is a key step in the regulation of gene expression.
• Role in Gene Expression: RNA polymerases are central to the process of gene expression. They initiate the synthesis of RNA molecules from genes, which serves as a template for protein synthesis (mRNA), a structural component of ribosomes (rRNA), or adaptor molecules for protein synthesis (tRNA).
• Critical for Cell Viability: RNA polymerases are essential for the viability of all living organisms. Their ability to transcribe genes is fundamental to the central dogma of molecular biology, allowing genetic information to flow from DNA to RNA to proteins.

Disadvantages of RNA Polymerase

RNA polymerases are essential enzymes in the process of transcription and gene expression, but they do have some limitations and potential disadvantages. Here are a few disadvantages or challenges associated with RNA polymerases:

• Lack of Proofreading: RNA polymerases do not possess the proofreading mechanisms found in DNA polymerases. As a result, they have a higher error rate during transcription. This can lead to the incorporation of incorrect nucleotides into the growing RNA strand, potentially resulting in mutated or nonfunctional RNA molecules.
• Sensitivity to Damaging Agents: RNA polymerases are sensitive to various DNA-damaging agents, such as UV radiation and chemical mutagens. Exposure to these agents can disrupt the transcription process, leading to errors in RNA synthesis or even cell death.
• Susceptibility to Termination Signals: RNA polymerases are susceptible to signals that trigger transcription termination. Premature or incorrect termination can result in incomplete or truncated RNA transcripts, affecting the functionality of the resulting RNA molecules.
• No 3'-5' Exonuclease Activity: Unlike DNA polymerases, RNA polymerases lack 3'-5' exonuclease activity. This means they cannot remove and correct incorrectly added nucleotides during RNA synthesis. Errors in the RNA transcript may persist unless corrected during post-transcriptional processing.
• Limited Processivity: RNA polymerases have limited processivity, which means they may dissociate from the DNA template prematurely during transcription. This can result in incomplete transcripts and may require the polymerase to reinitiate transcription multiple times for long genes.
• Susceptibility to Inhibition: RNA polymerases can be inhibited by various factors, including specific drugs and toxins. These inhibitors can interfere with transcription and disrupt normal gene expression.
• Requirement for Accessory Proteins: RNA polymerases often require the assistance of accessory proteins, such as transcription factors, to initiate transcription. The reliance on these additional factors can complicate the regulatory mechanisms governing gene expression.
• Directionality: RNA polymerases synthesize RNA in the 5'-to-3' direction, which is the opposite direction of the coding strand of DNA. This directionality can create challenges during transcription, as the polymerase must synthesize the RNA strand antiparallel to the template DNA strand.
• Limited Types of RNA: Each type of RNA polymerase is specialized for the synthesis of specific types of RNA (e.g., RNA polymerase II for mRNA). This specialization means that multiple RNA polymerases are required in a cell, which can complicate the regulatory machinery controlling gene expression.