MBMB 451A Section One - Fall 2006

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Transcription - Part A 

I.    RNA polymerase     

	A. E. coli RNA polymerase
		1. core enzyme = bb'(a)2 has catalytic activity but cannot recognize start site of transcription
			~500,000 daltons  
			dimensions: 100 X 100 X 160 angstroms
			requires Mg2+ for activity
			b' binds 2 Zn atoms
		2.  holoenzyme = core enzyme + sigma factor (s)
			carries out four functions: (i) template binding, (ii) RNA chain initiation
			   		           (iii) chain elongation, (iv) chain termination

	B.  Eukaryotic RNA polymerases	(RNAP)
		1.  3 nuclear RNA polymerases table 31.2
			a.  RNAP I- transcribes  rRNA genes
			b.  RNAP II - transcribes  mRNA genes
			c.  RNAP III - transcribes tRNA, 5S rRNA, and other small RNA genes
			d.  have 10-17 different subunits, large multisubunit complexes are functionally similar to E. coli RNA 
			     polymerase
			e.  cannot bind to their respective promoters alone, but requires transcription factors
			    for promoter specific recruitment		
		2.  organelle specific RNA polymerases
			a. chloroplast
			b.  mitochrondria	
		3.  RNAP II
			a.  core subunits - have sequence similarity to the core subunits of
			    E. coli core RNA polymerase or subunits of other eukaryotic RNA polymerases
			b.  shared or common subunits	
			     same subunits found in RNAP III and II or in RNAP I and RNAP II
			c.  unique subunits - no similar homologs found anywhere else
			d.  CTD or C-terminal domain of the largest subunit
				i.  a heptapeptide repeat that is repeated 52 times in humans and 26 times in yeast
				ii.  can be highly phosphorylated	at Ser and Thr
				iii.  three subforms of RNAP II: IIo, IIa, and IIb
			e.  cycling of phosphorylated and dephosphorylated forms of RNAP II associated 
			    with different stages in transcription	Figure 21.16
			f.  CTD is also required to recruit proteins for capping of 5'-end of mRNA, as well as for 
				splicing and polyadenylation of the 3'end of mRNA Figure 21.17
		4.  structure of yeast RNA polymerase II at 2.8 A˚ resolution figure 31.20, 31.21 and 31.22

II.  Promoter structure

	A.  rRNA genes - transcribed by RNAP I
		1.  Core promoter element  (-45 to +20) figure 21.5 (Genes VIII)
			alone is sufficient for transcription
		2.  Upstream control element  (-180 to -107)
			required for efficient transcription
		3.  transcription factor UBF binds to both promoter elements
		4.  which then helps recruit a second transcription factor called SLI1 
			a.  contains 4 subunits
			b.  one of the subunits is TBP
			c.  referred to as specificity factor - species specific

	B.  mRNA genes transcribed by RNAP II
		1.  TATA box element figure 21.10 (Genes VIII)
			located between -30 and -20 bps
		2.  Initiator region or In: centered on the start site of transcription
		3.  DPE: downstream promoter element
		4.  Response elements (RE) figure 21.21 (Genes VIII)
			a.  upstream of the TATA box
			b.  many different kinds - help respond to signals
			c.  multiple RE present - synergy
		5.  enhancers figure 21.22 (Genes VIII)
			a.  can be located at great distances (>1000 bps) from start site of transcription	
				eother from the 5' or 3' end of gene
			b.  stimulates transcription (~100 times)
			c.  orientation independent
			d.  two models of how enhancers might work figure 21.24 (Genes VIII)
				i.  entry point of RNAP II
					by preventing nucleosomes from binding or an altered DNA
					conformation thate promotes RNAP II recognition
				ii.  transcription factors bound to enhancer will stimulate binding of RNAP II
					to promoter regions closer to the start site of transcription

	C.  genes transcribed by RNAP III figure 21.6 (Genes VIII)
		1.  tRNA genes figure 21.7 and 21.8 (Genes VIII)
			a.  box A: crucial for start site selection			
			b.  box B 	
			c.  spacing between the two is variable	
			d. TFIIIC is bound to both box A and B
			e.  recruits TFIIIB to DNA upstream of start site of transcription	
		2.  5S rRNA gene figure 21.9 (Genes VIII)
			a.  box A			
			b.  box C is bound by TFIIIA
			c.  TFIIIA helps recruit TFIIIC and in turn TFIIIB		
		3.  U6 snRNA gene
			a.  TATA box		
			b.  PSE: proximal stimulatory element	
				centered near bp -50
			c.  DSE:  distal stimulatory element

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Nucleic Acids  |  Methods-Nucleic Acids: Part A and Part B and Molecular Cloning |  DNA Supercoiling  |  Nucleosome  | Transcription: Part B  |  Higher Ordered Structure  |  Chromatin Transcription  |  Regulation of Transcription  |  Course Material

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Last updated on September 6, 2005.