Analysis of the transcriptional activity of endogenous NFAT5 in primary cells using transgenic NFAT-luciferase reporter mice

Table 1 Impaired activation of endogenous 9xNFAT-Luc transgenic reporter by hypertonicity in NFAT5-deficient lymphocytes.

		9xNFAT-Luc reporter activity (RLU)
		Experiment # 1		Experiment # 2		Experiment # 2
		NFAT5 +/+	NFAT5 -/-	NFAT5 +/+	NFAT5 -/-	NFAT5 +/+	NFAT5 -/-
8-hour stimulation
300 mOsm/kg (isotonic)	- FK506	19	25	45	19	16	16
	+ FK506	11	13	18	16	7	8
430 mOsm/kg (hypertonic)	- FK506	1026	117	825	191	387	27
	+ FK506	720	83	1316	13	579	24
480 mOsm/kg (hypertonic)	- FK506	12616	540	5677	42	10901	273
	+ FK506	19543	435	2894	51	7403	102
PMA+ ionomycin (isotonic)	- FK506	2445	10087	2080	2494	1726	6364
	+ FK506	24	19	34	31	37	158
24-hour stimulation
300 mOsm/kg (isotonic)	- FK506	21	21	21	20	19	33
	+ FK506	10	15	14	15	12	10
430 mOsm/kg (hypertonic)	- FK506	1351	188	297	239	348	102
	+ FK506	550	122	254	129	388	73
480 mOsm/kg (hypertonic)	- FK506	6930	359	1742	224	7267	245
	+ FK506	1507	232	1111	129	3977	129
PMA+ ionomycin (isotonic)	- FK506	1100	2516	791	825	418	1775
	+ FK506	15	17	16	8	30	30

Luciferase activity was measured in proliferating transgenic 9xNFAT-Luc T cells derived from littermate NFAT5^+/+ and NFAT5^-/- mice after stimulation with hypertonic medium or PMA plus ionomycin, in the absence or presence of 100 nM FK506. Luciferase activity is represented as relative light units per second (RLU) after normalization with endogenous lactate dehydrogenase.

ISSN: 1471-2199