THE ROTAVIRUS ENTEROTOXIN NSP4 MOBILIZES INTRACELLULAR CALCIUM IN HUMAN INTESTINAL-CELLS BY STIMULATING PHOSPHOLIPASE C-MEDIATED INOSITOL 1,4,5-TRISPHOSPHATE PRODUCTION
Yj. Dong et al., THE ROTAVIRUS ENTEROTOXIN NSP4 MOBILIZES INTRACELLULAR CALCIUM IN HUMAN INTESTINAL-CELLS BY STIMULATING PHOSPHOLIPASE C-MEDIATED INOSITOL 1,4,5-TRISPHOSPHATE PRODUCTION, Proceedings of the National Academy of Sciences of the United Statesof America, 94(8), 1997, pp. 3960-3965
Rotavirus infection is the leading cause of severe diarrhea in infants
and young children worldwide. The rotavirus nonstructural protein NSP
4 acts as a viral enterotoxin to induce diarrhea and causes Ca2+-depen
dent transepithelial Cl- secretion in young mice. The cellular basis o
f this phenomenon was investigated in an in vitro cell line model for
the human intestine. Intracellular calcium concentration ([Ca2+](i)) w
as monitored in fura-2-loaded HT-29 cells using microscope-based fluor
escence imaging. NSP4 (1 nM to 5 mu M) induced both Ca2+ release from
intracellular stores and plasmalemma Ca2+ influx. During NSP4-induced
[Ca2+](i) mobilization, [Na+](i) homeostasis was not disrupted, demons
trating that NSP4 selectively regulated extracellular Ca2+ entry into
these cells. The ED50 of the NSP4 effect on peak [Ca2+](i) mobilizatio
n was 4.6 +/- 0.8 nM. Pretreatment of cells with either 2.3 x 10(-3) u
nits/ml trypsin or 4.4 x 10(-2) units/ml chymotrypsin for 1-10 min abo
lished the NSP4-induced [Ca2+](i) mobilization. Superfusing cells with
U-73122, an inhibitor of phospholipase C, ablated the NSP4 response.
NSP4 induced a rapid onset and transient stimulation of inositol 1,4,5
-trisphosphate (IP3) production in an IP3-specific radioreceptor assay
. Taken together, these results suggest that NSP4 mobilizes [Ca2+](i)
in human intestinal cells through receptor-mediated phospholipase C ac
tivation and IP3 production.