SUMMARY Thrombospondin (Thbs) proteins are induced in sites of tissue damage or active remodeling. The endoplasmic reticulum (ER) stress response is also prominently induced with disease where it regulates protein production and resolution of misfolded proteins. Here we describe a novel function for Thbs’ as ER resident effectors of an adaptive ER stress response. Thbs4 cardiac-specific transgenic mice were protected from myocardial injury while Thbs4−/− mice were sensitized to cardiac maladaptation. Thbs induction produced a unique profile of adaptive ER stress response factors and expansion of the ER and downstream vesicles. The type-3 repeat domain in Thbs’ bind the ER luminal domain of activating transcription factor 6α (Atf6α) to promote its nuclear shuttling. Thbs4−/−mice failed to show activation of Atf6α and other ER stress response factors with injury, and Thbs4-mediated protection was lost when Atf6α was deleted. Hence, Thbs’ can function inside the cell during disease/remodeling to augment ER function and protect through a mechanism involving regulation of Atf6α.

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To improve contractile function, the myocardium undergoes hypertrophic growth without myocyte proliferation in response to both pathologic and physiologic stimulation. Various membrane-bound receptors and intermediate signal transduction pathways regulate the induction of cardiac hypertrophy, but the cardioprotective regulatory pathways or effectors that antagonize cardiac hypertrophy remain poorly understood. Here we identify the small GTPase Cdc42 as a signaling intermediate that restrained the cardiac growth response to physiologic and pathologic stimuli. Cdc42 was specifically activated in the heart after pressure overload and in cultured cardiomyocytes by multiple agonists. Mice with a heart-specific deletion of Cdc42 developed greater cardiac hypertrophy at 2 and 8 weeks of stimulation and transitioned more quickly into heart failure than did wild-type controls. These mice also displayed greater cardiac hypertrophy in response to neuroendocrine agonist infusion for 2 weeks and, more remarkably, enhanced exercise-induced hypertrophy and sudden death. These pathologies were associated with an inability to activate JNK following stimulation through a MEKK1/ MKK4/MKK7 pathway, resulting in greater cardiac nuclear factor of activated T cells (NFAT) activity. Restoration of cardiac JNK signaling with an Mkk7 heart-specific transgene reversed the enhanced growth effect.These results identify what we believe to be a novel antihypertrophic and protective cardiac signaling pathway, whereby Cdc42-dependent JNK activation antagonizes calcineurin-NFAT activity to reduce hypertrophy and prevent transition to heart failure.

Members of the ADAR (adenosine deaminase that acts on RNA) enzyme family catalyze the hydrolytic deamination of adenosine to inosine within double-stranded RNAs, a poorly understood process that is critical to mammalian development. We have performed fluorescence resonance energy transfer experiments in mammalian cells transfected with fluorophore-bearing ADAR1 and ADAR2 fusion proteins to investigate the relationship between these proteins. These studies conclusively demonstrate the homodimerization of ADAR1 and ADAR2 and also show that ADAR1 and ADAR2 form heterodimers in human cells. RNase treatment of cells expressing these fusion proteins changes their localization but does not affect dimerization. Taken together these results suggest that homo-and heterodimerization are important for the activity of ADAR family members in vivo and that these associations are RNA independent. Double-stranded RNAs (dsRNAs)2 in eukaryotes are subject to a variety of processing reactions, including cleavage by the RNase III family members Drosha and Dicer in the micro RNA and small interfering RNA gene-silencing pathways and editing by members of the ADAR (adenosone deaminase that acts on RNA) enzyme family (1, 2). This latter reaction involves the hydrolytic deamination of adenosine (A) to inosine (I) within the context of dsRNA. Editing events of this type have been demonstrated in both cellular and viral transcripts and have been shown to function in altering the coding properties of the edited RNAs. For example, the life cycle of the Hepatitis ␦ virus is regulated by an editing event in the anti-genome in which a UAG stop codon is converted to a UIG tryptophan codon (3). An A to I modification is involved in the functional regulation of a growing number of cellular factors. These include the tissue-specific editing of the serotonin 5-HT2C receptor, which results in a reduction in response to serotonin agonists (4). Transcripts for subunits of the neural-specific AMPA class of glutamate-gated (GluR) ion channels undergo A to I modification at two positions, the Q/R and R/G editing sites, that affect the properties of the resulting channels (5, 6). In addition to the editing of these and other neuronal transcripts to effect codon changes, one deaminase family member, ADAR2, has been shown to autoregulate its own expression by the creation of a 3Ј-splice site (CAA to CAI) (7). Despite the identification of these editing substrates, the global role of A to I modification in higher eukaryotes remains unclear. Measurement of inosine levels in RNA isolated from rat tissue suggests a greater level of editing than indicated by known RNA substrates (8). A cloning protocol that depended upon an inosine-specific cleavage of RNA detected a large number of editing sites in non-coding regions of RNAs from Caenorhabditis elegans and humans that included sites in 5Ј-and 3Ј-untranslated regions and introns (9). Recent bioinformatic studies have suggested the presence of more than 12,000 editing sites corresponding to non-coding regions of t...