Eukaryotic Transcription and RNA Processing

Eukaryotic Transcription and RNA Processing

Introduction

This section covers the mechanisms of transcription and RNA processing in eukaryotic cells, highlighting the roles of transcription factors, the assembly of the transcription initiation complex, and the steps involved in mRNA maturation. Comparisons to prokaryotic systems are included to emphasize key differences.

Transcription Factors in Eukaryotes and Prokaryotes

General and Specific Transcription Factors

• General Transcription Factors (GTFs): Required for the recruitment of RNA polymerase II (Pol II) to the core promoter. They are necessary but not sufficient for physiological specificity of gene expression.

• Specific Transcription Factors (Activators): Bind to enhancers and are required for normal expression levels in vivo. They allow genes to respond to specific conditions.

Transcription Factors in Prokaryotes

• Prokaryotes use both basal and activator transcription factors.

• Regulation under different conditions is achieved by sigma factor switching.

• Note: In prokaryotes, sigma factors are needed for transcription initiation, but not for elongation.

RNA Polymerase II Initiation in Eukaryotes

Assembly of the Preinitiation Complex (PIC)

• The PIC is composed of general transcription factors and RNA polymerase II assembled at the core promoter.

• Assembly is an ordered process:

  • TFIID (composed of TBP and TAFs) binds the TATA box first.

  • Followed by the sequential binding of TFIIB, TFIIA, TFIIF, RNA polymerase II, TFIIE, and TFIIH.

Comparison: Initiation in Bacteria

• In bacteria, the switch from closed to open complex is mediated by sigma factors.

• Promoter escape is a key step in both systems.

Transcription Initiation and Promoter Escape

Steps in Initiation

• Additional basal transcription factors (TFIIJ and TFIIH) join the complex.

• TFIIH has helicase activity to melt the DNA, allowing the formation of the open complex.

• Promoter clearance/escape marks the transition to elongation.

• Abortive initiation may occur, where short RNA fragments are synthesized and released.

• Phosphorylation of the Pol II C-terminal domain (CTD) by TFIIH is required for promoter escape.

• Most transcription factors are released during promoter escape, but TFIIH remains associated with the polymerase during elongation.

Functions of TFIIH

• Kinase activity: Phosphorylates the Pol II CTD to initiate transcription and promote promoter escape.

• Helicase activity: Unwinds DNA for transcription initiation and is also involved in DNA repair (both genomic and transcription-coupled repair).

• TFIIH is unique in that it is not released from the polymerase during elongation.

The Carboxy Terminal Domain (CTD) of RNA Polymerase II

Structure and Function

• The CTD is a repeated heptapeptide sequence (YSPTSPS) found in the largest subunit of Pol II.

• Human Pol II CTD contains 52 repeats of this sequence.

• Multiple phosphorylation events occur on serine and threonine residues within the repeats.

• Phosphorylation state of the CTD regulates the transition from initiation to elongation and coordinates RNA processing events.

Comparison of Eukaryotic and Prokaryotic Transcription

Feature	Eukaryotes	Prokaryotes
Promoter Elements	TATA box (A/T rich consensus)	-10 (Pribnow box), -35 regions
Initiation Factors	General transcription factors (GTFs)	Sigma factors
RNA Polymerase Binding	Requires GTFs	Direct binding with sigma factor
mRNA Processing	Extensive (capping, splicing, polyadenylation)	Minimal (no capping or splicing)
Transcription-Translation Coupling	Separated (nucleus vs. cytoplasm)	Coupled (both in cytoplasm)
Introns	Present (removed by splicing)	Absent (rare exceptions)

Summary

• Eukaryotic transcription is a highly regulated, multi-step process involving numerous protein factors and complex RNA processing events.

• Key differences from prokaryotic transcription include the use of general transcription factors, the presence of introns, and the spatial separation of transcription and translation.

• The CTD of RNA Pol II plays a central role in coordinating transcription with RNA processing.

Additional info: Some details, such as the exact order of factor assembly and the number of CTD repeats, are standard textbook knowledge and have been clarified for completeness.