Questions: Transcription Initiation and Gene Regulation

Question 1

A gene has a fully functional TATA box and pre-initiation complex but all its enhancer sequences have been deleted. What would you expect?

• A. Transcription is completely abolished because enhancers are required for any transcription
• B. Low-level basal transcription occurs but the high-level cell-type-specific expression is lost
• C. Transcription is unaffected because enhancers only matter during development
• D. Transcription increases because repressors can no longer bind the enhancer to silence the gene

Answer: (B)

The TATA box and pre-initiation complex support *basal* transcription — a low, constitutive level of RNA synthesis. The thousand-fold differences in expression between cell types require enhancer-bound activators signaling through the Mediator complex to strongly stimulate RNA Pol II. Without enhancers, the gene is transcribed at a low background level but loses its capacity for regulated, high-level, cell-type-specific expression. This distinction between basal and activated transcription is the key insight: the promoter is the floor, the enhancer is the accelerator.

Question 2

What is the primary function of the Mediator complex in eukaryotic transcription?

• A. It is a general transcription factor that binds the TATA box and initiates PIC assembly
• B. It acts as a bridge between enhancer-bound activators and RNA Pol II at the promoter
• C. It is the helicase that unwinds DNA to create the transcription bubble
• D. It methylates histones to silence genes not needed in that cell type

Answer: (B)

Mediator is a large multi-subunit complex that does not bind DNA directly but serves as a molecular bridge: enhancer-bound transcriptional activators (which can be tens to hundreds of kilobases from the promoter) recruit Mediator, which in turn contacts RNA Pol II and the pre-initiation complex to stimulate transcription. Option A describes TFIID/TBP. Option C describes TFIIH's helicase subunit. Option D describes PRC2/EZH2 (Polycomb repressive complex). The Mediator's bridging function is what allows distant regulatory elements to control gene expression.

Question 3

Histone methylation typically represses transcription by compacting chromatin and blocking RNA polymerase access.

• T. True
• F. False

Answer: False

This is a common oversimplification. Histone methylation can be either activating or repressive depending on which residue is modified and to what degree. H3K4me3 (trimethylation of histone H3 at lysine 4) is found at active promoters and is associated with open, transcription-permissive chromatin. H3K27me3 and H3K9me3, by contrast, are repressive marks deposited by Polycomb and heterochromatin complexes. The same chemical modification on different residues has opposite effects — this context-specificity is central to the 'histone code' concept.

Question 4

Chromatin remodeling is a prerequisite for RNA Pol II to access and transcribe most eukaryotic genes because nucleosomes physically occlude the promoter and coding sequence.

• T. True
• F. False

Answer: True

Eukaryotic DNA is wound around histone octamers to form nucleosomes, which compact the genome but also block transcription factor binding and RNA polymerase progression. Before a gene can be robustly transcribed, nucleosomes at the promoter must be removed or repositioned by ATP-dependent chromatin remodeling complexes (like SWI/SNF) and histone-modifying enzymes. This is why activators typically recruit both Mediator (to stimulate Pol II) and histone acetyltransferases (to open chromatin). Chromatin state is the first gatekeeper of gene expression — an inaccessible promoter cannot be transcribed regardless of transcription factor availability.

Question 5

Explain why the same general transcription machinery (TFIID, RNA Pol II, Mediator) can produce thousands of different gene expression patterns across different cell types.

Model answer: The general transcription factors and RNA Pol II are constitutively expressed and support basal transcription at all promoters in principle. Cell-type-specific expression patterns arise from the combinatorial action of regulatory transcription factors that differ between cell types. Different cell types express different sets of activators and repressors that bind different enhancers; the particular combination of factors bound at a gene's enhancers determines how strongly that gene is stimulated. Additionally, each cell type has a distinct chromatin state (different genes are accessible or packed into repressive chromatin), further restricting which genes can be transcribed.

The multi-layered control system — transcription factor availability, enhancer binding, Mediator recruitment, and chromatin accessibility — acts like a combinatorial logic gate. Even small differences in which activators are present or which enhancers are accessible produce large differences in output. This is how a liver cell and a neuron, with identical genomes and identical general transcription machinery, express completely different proteomes.