Abstract
An inhibitory, 'dominant-negative,' form of the calcineurin catalytic (A) subunit was prepared, which lacks the calmodulin-binding domain, autoinhibitory domain and most of its catalytic core but possesses die regulatory (B) subunit binding domain. When tested for its ability to block calcineurin-dependent signaling in Jurkat cells, expression of this 'B-subunit knock-out' (BKO) construct suppressed reporter gene activity driven by NF-AT, the pivotal promoter element for interleukin (IL)-2 gene induction. Immunoprecipitation of epitope-labeled BKO demonstrated for the formation of a tight complex with endogenous B subunit in Jurkat cells, consistent with an inhibitory mechanism that involves the sequestration of the B subunit. Furthermore, the sharply reduced NF-AT activity produced by co-transfecting BKO could be 'rescued' by overexpression of transfected B subunit, suggesting that depletion of this subunit was responsible for the inhibition. These data suggest the potential utility of agents that disrupt calcineurin-mediated signal transduction pathways by blocking formation of the catalytically active dimer of calcineurin A and B subunits.
| Original language | English |
|---|---|
| Pages (from-to) | 466-472 |
| Number of pages | 7 |
| Journal | Biochemical and Biophysical Research Communications |
| Volume | 218 |
| Issue number | 2 |
| DOIs | |
| Publication status | Published - 1996 Jan 17 |
| Externally published | Yes |
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ASJC Scopus subject areas
- Biochemistry
- Biophysics
- Molecular Biology
Cite this
Inhibition of NF-AT signal transduction events by a dominant-negative form of calcineurin. / Muramatsu, Taro; Kincaid, Randall L.
In: Biochemical and Biophysical Research Communications, Vol. 218, No. 2, 17.01.1996, p. 466-472.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Inhibition of NF-AT signal transduction events by a dominant-negative form of calcineurin
AU - Muramatsu, Taro
AU - Kincaid, Randall L.
PY - 1996/1/17
Y1 - 1996/1/17
N2 - An inhibitory, 'dominant-negative,' form of the calcineurin catalytic (A) subunit was prepared, which lacks the calmodulin-binding domain, autoinhibitory domain and most of its catalytic core but possesses die regulatory (B) subunit binding domain. When tested for its ability to block calcineurin-dependent signaling in Jurkat cells, expression of this 'B-subunit knock-out' (BKO) construct suppressed reporter gene activity driven by NF-AT, the pivotal promoter element for interleukin (IL)-2 gene induction. Immunoprecipitation of epitope-labeled BKO demonstrated for the formation of a tight complex with endogenous B subunit in Jurkat cells, consistent with an inhibitory mechanism that involves the sequestration of the B subunit. Furthermore, the sharply reduced NF-AT activity produced by co-transfecting BKO could be 'rescued' by overexpression of transfected B subunit, suggesting that depletion of this subunit was responsible for the inhibition. These data suggest the potential utility of agents that disrupt calcineurin-mediated signal transduction pathways by blocking formation of the catalytically active dimer of calcineurin A and B subunits.
AB - An inhibitory, 'dominant-negative,' form of the calcineurin catalytic (A) subunit was prepared, which lacks the calmodulin-binding domain, autoinhibitory domain and most of its catalytic core but possesses die regulatory (B) subunit binding domain. When tested for its ability to block calcineurin-dependent signaling in Jurkat cells, expression of this 'B-subunit knock-out' (BKO) construct suppressed reporter gene activity driven by NF-AT, the pivotal promoter element for interleukin (IL)-2 gene induction. Immunoprecipitation of epitope-labeled BKO demonstrated for the formation of a tight complex with endogenous B subunit in Jurkat cells, consistent with an inhibitory mechanism that involves the sequestration of the B subunit. Furthermore, the sharply reduced NF-AT activity produced by co-transfecting BKO could be 'rescued' by overexpression of transfected B subunit, suggesting that depletion of this subunit was responsible for the inhibition. These data suggest the potential utility of agents that disrupt calcineurin-mediated signal transduction pathways by blocking formation of the catalytically active dimer of calcineurin A and B subunits.
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U2 - 10.1006/bbrc.1996.0083
DO - 10.1006/bbrc.1996.0083
M3 - Article
VL - 218
SP - 466
EP - 472
JO - Biochemical and Biophysical Research Communications
JF - Biochemical and Biophysical Research Communications
SN - 0006-291X
IS - 2
ER -