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INNATE IMMUNITY 3
pNiFty - PRR Signaling Reporter Plasmids
PRR activation triggers a complex signaling cascade that leads to the activation of different transcription factors, each playing an important role in the
subsequent immune response. To monitor the induction of PRR signaling in response to ligand stimulation in a simple and efficient manner, InvivoGen
has designed pNiFty, a family of reporter plasmids expressing a reporter gene under the control of a minimal promoter inducible by these different
transcription factors, either individually or in combination. Most pNiFty plasmids are selectable with Zeocin in both E. coli and mammalian cells, and
can be used to generate stable clones
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PLASMID NAME
Description
TRANSCRIPTION FACTOR
BINDING SITES
pNiFty plasmids are composed of three key elements: a proximal
promoter, repeated transcription factor binding sites (TFBS) and a
reporter gene. The proximal promoters are shorter than 500 bp and
contain transcription factor binding sites. Upon stimulation in 293 cells,
their expression level remains undetectable. With the addition of repeated
TFBS, the proximal promoters become inducible by the appropriate
stimulus and drive the expression of the reporter gene.
Minimal promoters
- ELAM promoter: the proximal promoter of the endothelial cellleukocyte
adhesion molecule (ELAM-1; E-selectin) gene contains three
NF-kB sites and is truly NF-kB specific, as it lacks an AP1/CREB site found
in the full-length promoter 1, 2 .
- IFN-b promoter: the mouse IFN-b minimal promoter comprises several
positive regulatory domains that bind different cooperating transcription
factors such as NF-kB, IRF3 and IRF7 3 .
- ISG-56K promoter: the minimal promoter of the human interferonstimulated
gene ISG-56K contains two interferon-stimulated regulatory
element (ISRE) sites and is fully inducible by type I IFNs and interferon
regulatory factors (IRFs) 4, 5 .
Transcription factor binding sites (TFBS)
- AP-1 binding site: Activator protein 1 (AP-1) is a transcription factor
activated by most PRRs. AP-1 is a heterodimeric complex composed of
members of Fos, Jun and, ATF protein families. AP-1 binds to the TPA
responsive element (TRE: ; TGAG/CTCA) 6 . AP-1 activation in TLR signaling
is mostly mediated by MAP kinases such as c-Jun N-terminal kinase (JNK),
p38 and extracellular signal regulated kinase (ERK).
MINIMAL
PROMOTER
www.invivogen.com/innate-immunity-pnifty
SELECTION REPORTER
CATALOG
CODE
pNiFty-SEAP NF-kB (x5) ELAM Ampicillin SEAP pnifty-seap
pNiFty-Luc NF-kB (x5) ELAM Ampicillin Luciferase pnifty-luc
pNiFty2-SEAP NF-kB (x5) ELAM Zeocin SEAP pnifty2-seap
pNiFty2-Luc NF-kB (x5) ELAM Zeocin Luciferase pnifty2-luc
pNiFty2-56K None ISG56 Zeocin SEAP pnf2-56sp
pNiFty3-SEAP NEW None IFN-b Zeocin SEAP pnf3-sp1
pNiFty3-N-SEAP NEW NF-kB (x5) IFN-b Zeocin SEAP pnf3-sp2
pNiFty3-A-SEAP NEW AP-1 (x5) IFN-b Zeocin SEAP pnf3-sp3
pNiFty3-I-SEAP NEW ISRE (x5) IFN-b Zeocin SEAP pnf3-sp4
pNiFty3-T-SEAP NEW NFAT (x5) IFN-b Zeocin SEAP pnf3-sp5
pNiFty3-AN-SEAP NEW AP-1 (x5) NF-kB (x5) IFN-b Zeocin SEAP pnf3-sp6
pNiFty3-IAN-SEAP NEW ISRE (x5) AP-1 (x5) NF-kB (x5) IFN-b Zeocin SEAP pnf3-sp7
pNiFty3-TAN-SEAP NEW NFAT (x5) AP-1 (x5) NF-kB (x5) IFN-b Zeocin SEAP pnf3-sp8
SEAP
or
Luciferase
SEAP
or
Luciferase