Differential Distribution, Clustering, and Lateral Diffusion of Subtypes of the Inositol 1,4,5-Trisphosphate Receptor

Regulation of inositol 1,4,5-trisphosphate (IP(3)) receptors (IP(3)R) by IP(3) and Ca(2+) allows them to initiate and regeneratively propagate intracellular Ca(2+) signals. The distribution and mobility of IP(3)R determines the spatial organization of these Ca(2+) signals. Until now, there has been no systematic comparison of the distribution and mobility of the three mammalian IP(3)R subtypes in a uniform background. We used confocal microscopy and fluorescence recovery after photobleaching to define these properties for each IP(3)R subtype expressed heterologously in COS-7 cells. IP(3)R1 and IP(3)R3 were uniformly distributed within the membranes of the endoplasmic reticulum (ER), but the distribution of IP(3)R2 was punctate. The mobile fractions (M(f) = 84 ± 2 and 80 ± 2%) and diffusion coefficients (D = 0.018 ± 0.001 and 0.016 ± 0.002 μm(2)/s) of IP(3)R1 and IP(3)R3 were similar. Other ER membrane proteins (ryanodine receptor type 1 and sarco/endoplasmic reticulum Ca(2+)-ATPase type 1) and a luminal protein (enhanced GFP with a KDEL retrieval sequence) had similar mobile fractions, suggesting that IP(3)R1 and IP(3)R3 move freely within an ER that is largely, although not entirely, continuous. IP(3)R2 was less mobile, but IP(3)R2 mobility differed between perinuclear (M(f) = 47 ± 4% and D = 0.004 ± 0.001 μm(2)/s) and near-plasma membrane (M(f) = 64 ± 6% and D = 0.013 ± 0.004 μm(2)/s) regions, whereas IP(3)R3 behaved similarly in both regions. We conclude that IP(3)R1 and IP(3)R3 diffuse freely within a largely continuous ER, but IP(3)R2 is more heterogeneously distributed and less mobile, and its mobility differs between regions of the cell.