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. 2009 Nov 17;106(49):20776–20781. doi: 10.1073/pnas.0906998106

Fig. 4.

Fig. 4.

Effect of HAX-1 on PLN-PLN and PLN-SERCA fluorescence resonance energy transfer (FRET). (A) Fluorescence microscopy showed that acceptor-selective photobleaching of YFP-PLN fluorescence resulted in an increase in CFP-SERCA fluorescence, indicating FRET. (B) Image quantification showed CFP-SERCA fluorescence (blue circles) increased exponentially as YFP-PLN (green triangles) was photobleached. The magnitude of CFP-SERCA fluorescence enhancement was greater in cells cotransfected with HAX-1 (black squares), suggesting increased FRET compared to control. (C) CFP-PLN fluorescence (blue circles) increased as YFP-PLN (green triangles) was bleached, indicating intrapentameric FRET. The magnitude of the CFP-PLN fluorescence enhancement was smaller in cells cotransfected with HAX-1 (black squares), suggesting reduced FRET. (D) Summary of FRET microscopy results. Coexpression of HAX-1 caused a 32% decrease in overall FRET between CFP-PLN and YFP-PLN, and a 35% increase in overall FRET between CFP-SERCA and YFP-PLN. Data are mean ± SEM, *, P < 0.01, compared with control. (E) A model for HAX-1 regulation of cardiac calcium handling. HAX-1 shifts the PLN pentamer/monomer equilibrium toward the active, monomeric species, and promotes formation of the PLN-SERCA regulatory complex.