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Transcription: Synthesis of RNA – Biochemistry Study Notes

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Transcription: Synthesis of RNA

Overview of Transcription

Transcription is the process by which RNA is synthesized from a DNA template. This fundamental process is catalyzed by RNA polymerases, which generate single-stranded RNA molecules complementary to one strand of DNA. Transcription is essential for gene expression and regulation in both prokaryotic and eukaryotic cells.

  • RNA polymerase: Enzyme responsible for synthesizing RNA from DNA.

  • Template strand: The DNA strand used as a template for RNA synthesis.

  • Promoter: DNA sequence where RNA polymerase binds to initiate transcription.

  • Directionality: RNA is synthesized in the 5' to 3' direction.

  • Regulatory sequences: Elements such as enhancers and promoter-proximal elements influence transcription efficiency.

Action of RNA Polymerase

RNA polymerase catalyzes the polymerization of ribonucleotides into RNA using a DNA template. The enzyme recognizes specific promoter regions to initiate transcription and proceeds along the template strand, synthesizing RNA by adding ribonucleoside triphosphates (ATP, GTP, CTP, UTP).

  • Polymerization reaction:

  • Base pairing: RNA nucleotides pair with complementary DNA bases (A-U, G-C).

  • Pyrophosphate release: Each nucleotide addition releases pyrophosphate ().

  • High fidelity: Error rate of RNA polymerase is approximately 1 in 100,000 bases.

Types of RNA Polymerases

Both prokaryotes and eukaryotes possess multiple RNA polymerases, each responsible for synthesizing different types of RNA.

Polymerase

Product

RNA polymerase I

rRNA (except 5S rRNA)

RNA polymerase II

mRNA, snRNA, miRNA

RNA polymerase III

tRNA, 5S rRNA, other small RNAs

Additional info: In prokaryotes, a single RNA polymerase synthesizes all major RNA types.

Sequences of Genes

Genes consist of a coding strand and a template strand. The coding strand has the same sequence as the RNA (except T is replaced by U), while the template strand is used for RNA synthesis.

  • Sense strand: Coding strand, matches mRNA sequence.

  • Antisense strand: Template strand, used by RNA polymerase.

  • Transcription unit: Region of DNA transcribed into RNA.

Promoter Regions and Recognition

Promoters are specific DNA sequences that direct RNA polymerase to the correct start site for transcription. In eukaryotes, promoters contain elements such as the TATA box, CAAT box, and GC box. In prokaryotes, promoters have -10 and -35 regions recognized by sigma factors.

Promoter Element

Location

Function

TATA box

-25 to -30

Core promoter, binding site for transcription factors

CAAT box

-75

Upstream promoter element

GC box

-90

Upstream promoter element

-10 region (Pribnow box)

-10 (prokaryotes)

Promoter recognition

-35 region

-35 (prokaryotes)

Promoter recognition

Transcription in Prokaryotes

In bacteria, transcription is initiated by RNA polymerase binding to the promoter region. Genes are often organized in operons, allowing for polycistronic mRNA synthesis (one mRNA encoding multiple proteins).

  • Polycistronic mRNA: Single mRNA encoding several proteins.

  • Operon: Cluster of genes under control of a single promoter.

  • Termination: Occurs via rho-dependent or rho-independent mechanisms.

Transcription in Eukaryotes

Eukaryotic transcription is more complex, involving multiple RNA polymerases and extensive regulation. Transcription factors and co-activators are required for initiation, and the process occurs in the nucleus.

  • Transcription factors: Proteins that assist RNA polymerase binding and initiation.

  • Basal transcription factors: TFIIA, TFIIB, TFIID, TFIIE, TFIIF, TFIIH.

  • Co-activators: Mediate interactions between transcription factors and RNA polymerase.

  • RNA processing: Includes 5' capping, polyadenylation, and splicing.

RNA Processing in Eukaryotes

Newly synthesized eukaryotic mRNA (hnRNA) undergoes several modifications before becoming mature mRNA.

  • 5' Capping: Addition of a 7-methylguanosine cap to the 5' end.

  • Polyadenylation: Addition of a poly(A) tail at the 3' end, signaled by the AAUAAA sequence.

  • Splicing: Removal of introns and joining of exons by the spliceosome complex.

Modification

Function

5' Cap

Protects mRNA from degradation, assists in ribosome binding

Poly(A) tail

Stabilizes mRNA, aids in export from nucleus

Splicing

Removes non-coding introns, joins coding exons

Splicing Mechanism

Splicing is carried out by the spliceosome, a complex of snRNAs and proteins. It recognizes specific sequences at exon-intron boundaries (5' GU and 3' AG) and removes introns via two transesterification reactions, forming a lariat structure.

  • snRNAs: U1, U2, U4, U5, U6 are involved in spliceosome assembly and function.

  • Lariat formation: Intron is looped out and excised.

Synthesis of rRNA and tRNA

Ribosomal RNA (rRNA) and transfer RNA (tRNA) are transcribed and processed in the nucleolus. rRNA genes are organized in clusters, and tRNA genes contain introns that are removed during maturation.

  • rRNA: Forms the core of ribosomal subunits (18S, 5.8S, 28S in eukaryotes).

  • tRNA: Adopts a cloverleaf structure, contains modified bases (e.g., pseudouridine, ribothymidine).

Differences Between Eukaryotic and Prokaryotic DNA/RNA

Several key differences exist between eukaryotic and prokaryotic gene structure and transcription mechanisms.

Parameter

Eukaryotes

Prokaryotes

Genome

Diploid

Haploid

Introns

Present

Absent

Poly(A) tail

Present

Absent

mRNA

Monocistronic

Polycistronic

Transcription/Translation

Separated (nucleus/cytoplasm)

Coupled

Transcription and HIV

HIV uses reverse transcriptase to synthesize DNA from its RNA genome, which then integrates into the host genome. Transcription of viral genes produces new viral RNA and proteins.

  • Reverse transcriptase: Enzyme that synthesizes DNA from RNA.

  • Integration: Viral DNA integrates into host cell DNA.

  • Transcription: Host machinery transcribes viral genes.

Key Concepts

  • Transcription is the synthesis of RNA from a DNA template.

  • RNA polymerases require promoter sequences to initiate transcription.

  • Prokaryotic and eukaryotic transcription differ in complexity and regulation.

  • RNA processing in eukaryotes includes capping, polyadenylation, and splicing.

  • Gene expression is regulated at multiple levels, including transcription initiation and RNA processing.

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