Inositol-1,3,4,5-tetrakisphosphate binding sites in control and ras-transformed NIH/3T3 fibroblasts

Inositol-1,3,4,5-tetrakisphosphate (Ins(1,3,4,5)P₄) binding properties were investigated in NIH/3T3 fibroblasts and its ras-transformant (DT cells), in which inositol tetrakisphosphates induce Ca²⁺ influx. [³H]Ins(1,3,4,5)P₄ bound to membranes of both types of cells with Kd values of 10.6 and 8.6 nM, respectively. The rank order of inositol polyphosphates for displacing [³H]Ins(1,3,4,5)P₄ in DT cells was Ins(1,3,4,5)P₄ > inositol-1,3,4,5,6-pentakisphosphate > inositol hexakisphosphate > inositol-1,4,5-trisphosphate. This order is similar to that reported in two Ras-GTPase-activating proteins, GAP1(IP4BP) and GAP1^m, which are also the Ins(1,3,4,S)P, binding proteins. Northern blot analysis revealed that NIH/3T3 and DT cells expressed mRNA species that were hybridizable with GAP1^m cDNA. These results suggest that parental and ras-transformed NIH/3T3 fibroblasts possess GAP1-like proteins, which may be responsible for triggering inositol tetrakisphosphate-dependent Ca² influx.