Functionally redundant control of cardiac hypertrophic signaling by inositol 1,4,5-trisphosphate receptors

Abstract

Calcium plays an integral role to many cellular processes including contraction, energy metabolism, gene expression, and cell death. The inositol 1, 4, 5-trisphosphate receptor (IP3R) is a calcium channel expressed in cardiac tissue. There are three IP 3R isoforms encoded by separate genes. In the heart, the IP3R-2 isoform is reported to being most predominant with regards to expression levels and functional significance. The functional roles of IP3R-1 and IP3R-3 in the heart are essentially unexplored despite measureable expression levels. Here we show that all three IP3Rs isoforms are expressed in both neonatal and adult rat ventricular cardiomyocytes, and in human heart tissue. The three IP3R proteins are expressed throughout the cardiomyocyte sarcoplasmic reticulum. Using isoform specific siRNA, we found that expression of all three IP 3R isoforms are required for hypertrophic signaling downstream of endothelin-1 stimulation. Mechanistically, IP 3Rs specifically contribute to activation of the hypertrophic program by mediating the positive inotropic effects of endothelin-1 and leading to downstream activation of nuclear factor of activated T-cells. Our findings highlight previously unidentified functions for IP 3R isoforms in the heart with specific implications for hypertrophic signaling in animal models and in human disease.

DHHC5 Mediates β-Adrenergic Signaling in Cardiomyocytes by Targeting Gα Proteins

“…The rat myoblast H9c2 and HEK293 cell lines were obtained from American Type Culture Collection and maintained according to American Type Culture Collection guidelines. Neonatal rat ventricular cardiomyocytes (NRVMs) were prepared from 1- to 2-day-old Sprague-Dawley rat pups, as previously described ( 22 ). Animals were purchased from Texas Animal Specialties (Humble, TX) and processed on the same day.…”

“…NRVMs were plated at a density of 300 cells per mm 2 on fibronectin-coated glass coverslips. Cardiomyocytes were loaded with Fura-2 AM as described previously ( 22 ). Images were taken on a Nikon TiS inverted microscope (Tokyo, Japan) with a 40× oil immersion objective, and images were taken every second with a Photometrics Evolve electron-multiplying charged-coupled device camera (Tucson, AZ).…”

“…An oscillation was counted when the Fura-2 ratio rose 10% above the baseline ratio. Oscillation frequencies before and after ISO were calculated from five coverslips for each condition and mean ± standard error were plotted as described ( 22 ).…”

“…NRVMs were plated at a density of 250 cells per mm 2 on fibronectin-coated glass coverslips. Immunofluorescence staining was performed as described previously ( 22 ). Anti-DHHC5 was used at a concentration of 1:100.…”

Calcium Signaling in Cardiomyocyte Function

“…Inositol 1,4,5-trisphosphate receptor (InsP 3 R) intracellular Ca 2+ release channels are expressed in cardiomyocytes (Lipp et al 2000; Kockskämper et al 2008b;Garcia et al 2017). Although all…”