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. 2016 Jan 11;291(10):4846–4860. doi: 10.1074/jbc.M115.705301

FIGURE 2.

FIGURE 2.

IP3R concatenated dimers are stably expressed in DT40 3KO cells, form tetramers, and are functionally indistinguishable from IP3R monomers. A, lysates from DT40-3KO or DT40-3KO cells stably expressing IP3R constructs were resolved on 4% SDS-PAGE and probed with α-R2NT or α-R3. B, lysates from DT40-3KO cells stably expressing IP3R constructs were resolved on 3–12% native-PAGETM Novex gels. Immunoblots were processed using α-R2NT or α-R3. C, [3H]IP3 binding curves of R1, R2, and R3 monomers, compared with R1R1, R2R2, and R3R3 homodimers, generated using homologous competitive binding assays. D, [IP3]-response relationships of R1, R2, and R3 monomers, compared with R1R1, R2R,2 and R3R3 homodimers. High throughput permeabilized cell IICR assays were carried out using FlexStation 3. IP3-mediated Ca2+ release was induced through addition of varying [IP3] in the presence of 5 mm ATP and 200 nm free Ca2+. All values were normalized to the maximal release rate. Each point is the mean ± S.E. of eight wells from at least three experiments.