Transcription

 

I. Introduction: the central dogma of  molecular biology

A. gene expression is DNA->RNA->Protein
B. DNA->RNA is transcription
C. RNA->Protein is translation
D. the difference that compartmentalization makes: prokaryotic vs. eukaryotic gene expression.

II. Transcription is the process that provides the molecular definition of a gene.

A.  a gene is a sequence of DNA that is transcribed.
B.  in other words, a gene is the DNA ranging from the promoter to the terminator (inclusive).
C. later on, well encounter less ambiguous terminology - "open reading frame"

III. Promoters defined

A. evidence from partial diploid p mutants (classic example, lacP minus mutants)

1. the term "promoter" was coined based on the behavior of a peculiar type of lac- mutant.  Here's how it behaved:

a.  lacP- lacZ+  had Lac- phenotype (no problem)
b. lacP+ lacZ-    ditto (again, no problem)
c. merodiploid  lacP- lacZ+ / lacP+ lacZ+  was Lac+ (ok, so lacP- appears to be recessive, no problem still)
d. but, check this one out: lacP- LacZ+ / lacP+ lacZ- is Lac- !? why doesn't this complement? It is not behaving like a recessive mutation here.

2. answer: lacP is a "cis-acting" regulatory sequence.  This means, to get a wild-type phenotype, you must have the wild-type lacP ON THE SAME MOLECULE as the wild-type lacZ.  No other combinations will work.  Specifcally we call mutants of the lacP- type cis-dominant.
3.  Beckwith proved that the lacP region (termed promoter by Jacob and Monod), is the site where transcription begins.

B. evidence from "sequence gazing": the Pribnow box (or -10 consensus sequence) and the -35 sequence
C. DNase I footprinting
D. Odds and ends

1. RNA polymerase structure (core polymerase)
2. sigma factors

IV. The mechanism of transcription (see the Flash animation, courtesy of Media del Gekko )

A. Initiation
B. Elongation
C. Termination

V. The molecular events of transcription
        A. initiation
                1. Closed complex
                2. Open complex
        B. Elongation and sigma factor release
        C. Termination

VI.  Terminators defined
        A. rho dependent
        B. rho independent

VII. Eukaryotic transcription

A different RNA polymerases transcribe different classes of genes in eukaryotes
            1. RNA pol I does rRNA except 5S rRNA
            2. RNA pol II makes mRNA and snRNA
            3. RNA pol III does tRNA, 5S rRNA

B. typical core promoter (leaving out the regulatory elements for now)
             1. TATA box helps establish the transcription start site
             2. CAAT boxes and GC boxes recruit general transcription factors and RNA polymerase

C. Processing of eukaryotic RNAs

1. capping
2. polyadenylation
3. splicing