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3
0
hydroxyl group of the upstream exon so it can attack the phosphodiester bond at
the 3
0
splice site, linking the exons together and releasing the lariat intron for
debranching and degradation (
Wachtel and Manley, 2009
).
The process of mRNA 3
0
end formation is highly conserved among eukaryotes.
Several groups of proteins are involved, and research into this process, along with
the related process of transcription termination, is rapidly progressing. There are
scores of proteins involved in 3
0
cleavage and transcription termination. Of these
proteins, two different complexes appear to play crucially important roles and
have been studied most thoroughly, usually in mammalian systems: CPSF (cleav-
age and polyadenylation specificity factor) and CstF (cleavage stimulation fac-
tor). In yeast, the protein constituents of these complexes associate somewhat
differently, and the complexes are CF1A and CPF (reviewed in
Mandel et al.,
2008
).
In vertebrates, The CPSF complex catalyzes cleavage of the RNA at a site
30 nt
downstream of the AAUAAA polyadenylation signal to which it binds (
Fitzgerald and
Shenk, 1981
). CPSF contains five proteins: CPSF-160, CPSF-100, CPSF-73, CPSF-
30, and Fip1 (
Kaufmann et al., 2004; Mandel et al.,2008
). CPSF-160 recognizes and
binds directly to the AAUAAA. It also may link the CPSF complex to the CstF
complex. CPSF-73 is the endonuclease that cleaves the RNA (
Mandel et al.,2006
).
The CstF complex contains three polypeptides: CstF-77, CstF-64, and CstF-50
(
Mandel et al., 2008
). CstF-77 acts as a scaffold around which the complex
assembles (
Legrand et al., 2007
). It also interacts with CPSF-160, effectively
linking the CPSF and CstF complexes. The CstF-64 binds to a less-well-defined
GU-rich region close to, but downstream of, the cleavage site on the RNA. The
CstF-50 subunit binds to the C-terminal domain of the largest subunit of RNA
polymerase, thereby physically linking the 3
0
end formation and transcriptional
machineries (
McCracken et al.,1997
).
Evidence from ChIP experiments indicates that the complexes travel along the
transcribed DNAwith RNA polymerase (
Glover-Cutter et al., 2008
). They scan the
nascent RNA for their recognition sequences, and, when both AAUAAA and a
downstream GU-rich sequence are found in the proper orientation, 3
0
end formation
can occur. Some variability is tolerated at the polyadenylation signal. A survey in
humans and mice showed that 70% of all polyadenylation signals contained the
canonical sequence AAUAAA. The only variant commonly seen was AUUAAA
(15%), indicating that there is little plasticity in CPSF
'
s ability to recognize this 3
0
processing signal in mammals (
Mandel et al., 2008
).
B. Operons
An operon is a cluster of genes transcribed together from a common upstream
promoter. In the operons of bacteria and archaea (
Brown et al., 1989; Jacob and
Monod, 1961
), genes encoding enzymes in the same metabolic pathway are
often located next to each other on the chromosome. A common upstream
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