Heart Failure Re-Distributes Phospholamban between Nuclear Membranes and Sarcoplasmic Reticulum in Cardiomyocytes

Abstract

We recently documented that phospholamban (PLB), a sarcoplasmic reticulum (SR) resident protein regulator of cardiac SR Ca-ATPase (SERCA2a), is localized to the nuclear envelope (NE) of cardiomyocytes (CMs). It is well-known that heart failure (HF) remodeling involves altered protein expression of PLB and SERCA2a; however, with the demonstrated compartmentalization of PLB accumulation, we asked whether HF alters trafficking between ER/SR subcompartments affecting steady-state intracellular distribution of PLB and SERCA. In this study, we examined heart tissue from non-failing and pacing-induced HF dogs with the monoclonal PLB (2D12 or 1F1) and SERCA (2A7-A1) antibodies. Confocal immunofluorescence microscopy was used to compare subcellular concentration of PLB and SERCA. At least 3 regions from NE and SR in 10 CMs were analyzed for each dog, n = 4 dogs). Both PLB and SERCA antibodies stained SR and NE in these CMs. Fluorescence intensity ratios between NE and SR for PLB was 2.05±0.82 (mean ± SD) in control dog CMs, consistent with results just reported. In HF CMs we found a highly decreased ratio of PLB between NE and SR to 1.26 ± 0.34 (n = 8 dogs). In contrast to re-distribution of PLB, intensity ratios for SERCA in NE and SR remained unchanged (1.01±0.14 in Control, and 0.98±0.26 in HF). Furthermore, while there was only minor levels of phosphoPLB in Control CMs, we found that phosphoPLB (S16) were very significantly higher in NE of failed CMs. The decreased PLB relative to SERCA in the NE of CMs suggests that retention and accumulation of PLB in the NE is decreased in HF, leading to a smaller, possibly hyper-phosphorylated steady-state of PLB concentration in the NE.