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et al.
, 2004; Johnson
et al.
, 2003
). Internal deletions of the let-7 promoter in
a GFP reporter assay identified this 116nt region, which contains a 9nt
inverted repeat and is conserved in
C. briggsae
, as necessary for expression of
GFP in the hypodermal seam cells (
Johnson
et al.
, 2003
). Compared to wild
type,
let-7
transgenes lacking the TRE exhibit reduced rescue activity. The
proteins that regulate transcription of let-7 through the TRE, as well as the
elements responsible for the oscillating transcription pattern, and expression
in other tissues are yet to be identified.
So far, two transcription factors have been found to regulate the
expression of
let-7
in
C. elegans
. The
hbl-1
gene encodes a zinc-finger
transcription factor that shares homology with the
Drosophila
Hunchback
gene (
Abrahante
et al.
, 2003; Fay
et al.
, 1999; Lin
et al.
, 2003
). HBL-1 is
predicted to bind to an A-rich sequence 18nt downstream of the TRE
and repress transcription of
let-7
in the hypodermal seam cells (
Fig. 1.7
;
Roush and Slack, 2009
). The nuclear hormone receptor DAF-12 regulates
the transcription of
let-7
in a hormone dependent manner (
Bethke
et al.
,
2009; Hammell
et al.
, 2009a
). In the absence of ligand, DAF-12 represses
the expression of
let-7
and several of its sister miRNAs. When bound to
ligand, DAF-12 activates expression of some
let-7
family members. DAF-
12 response elements have been identified in the promoters of
mir-241
and
mir-84,
but direct interaction with the
let-7
promoter remains to be
demonstrated (
Bethke
et al.
, 2009
). Interestingly, both
hbl-1
and
daf-12
are targets of regulation by
let-7
family miRNAs (
Abbott
et al.
, 2005;
Abrahante
et al.
, 2003; Großhans
et al.
, 2005; Lin
et al.
, 2003
). Through
multiple 3
0
UTR complementary sites, the let-7 sisters, mir-48, mir-84,
and mir-241, initiate repression of
hbl-1
expression during the transition
from the second to the third larval stage (
Abbott
et al.
, 2005
). These
miRNAs are expressed by the second larval stage, one stage earlier than
let-7, thus, providing a mechanism to reduce HBL-1 levels and allow for
transcription of let-7 in the seam cells (
Abbott
et al.
, 2005; Roush and
Slack, 2009
). Likewise, the let-7 sisters also target
daf-12
for downregula-
tion at the L3 stage, which may promote transcription of let-7 in some
tissues (
Bethke
et al.
, 2009; Hammell
et al.
, 2009a
). Expression of mature
let-7 in L3 augments repression of
hbl-1
and
daf-12
, adding to the feedback
loop of transcriptional and miRNA-mediated control (
Abbott
et al.
, 2005;
Abrahante
et al.
, 2003; Bethke
et al.
, 2009; Großhans
et al.
, 2005;
Hammell
et al.
, 2009a; Lin
et al.
, 2003
).
In
C. elegans
,
70% of all mRNAs are
trans
-spliced to one of two 22nt
splice leader (SL) sequences, SL1 or SL2 (
Blumenthal, 2005
). The
trans-
splicing
reaction is carried out by a spliceosome that functions similarly to the
cis
-
splicing complex, which removes introns, except that the 5
0
end of the SL
RNA acts as the 5
0
exon and is ligated to a 3
0
splice site downstream of the 5
0
cap in the acceptor mRNA (
Blumenthal, 2005
). The role of
trans
-splicing is
not fully understood, but is thought to aid in nuclear export and translation.
