Heterogeneity of heart failure (HF) phenotypes indicates contributions from underlying common polymorphisms. We considered polymorphisms in the beta(1)-adrenergic receptor (beta(1)AR), a beta-blocker target, as candidate pharmacogenomic loci. Transfected cells, genotyped human nonfailing and failing ventricles, and a clinical trial were used to ascertain phenotype and mechanism. In nonfailing and failing isolated ventricles, beta(1)-Arg-389 had respective 2.8 +/- 0.3- and 4.3 +/- 2.1-fold greater agonist-promoted contractility vs. beta(1)-Gly-389, defining enhanced physiologic coupling under relevant conditions of endogenous expression and HF. The beta-blocker bucindolol was an inverse agonist in failing Arg, but not Gly, ventricles, without partial agonist activity at either receptor; carvedilol was a genotype-independent neutral antagonist. In transfected cells, bucindolol antagonized agonist-stimulated cAMP, with a greater absolute decrease observed for Arg-389 (435 +/- 80 vs. 115 +/- 23 fmol per well). Potential pathophysiologic correlates were assessed in a placebo-controlled trial of bucindolol in 1,040 HF patients. No outcome was associated with genotype in the placebo group, indicating little impact on the natural course of HF. However, the Arg-389 homozygotes treated with bucindolol had an age-, sex-, and race-adjusted 38% reduction in mortality (P = 0.03) and 34% reduction in mortality or hospitalization (P = 0.004) vs. placebo. In contrast, Gly-389 carriers had no clinical response to bucindolol compared with placebo. Those with Arg-389 and high baseline norepinephrine levels trended toward improved survival, but no advantage with this allele and exaggerated sympatholysis was identified. We conclude that beta(1)AR-389 variation alters signaling in multiple models and affects the beta-blocker therapeutic response in HF and, thus, might be used to individualize treatment of the syndrome.

Ca 2؉ /calmodulin (CaM)-dependent protein kinase II (CaMKII) is a major mediator of cellular Ca 2؉ signaling. Several inhibitors are commonly used to study CaMKII function, but these inhibitors all lack specificity. CaM-KIIN is a natural, specific CaMKII inhibitor protein. CN21 (derived from CaM-KIIN amino acids 43-63) showed full specificity and potency of CaMKII inhibition. CNs completely blocked Ca 2؉ -stimulated and autonomous substrate phosphorylation by CaMKII and autophosphorylation at T305. However, T286 autophosphorylation (the autophosphorylation generating autonomous activity) was only mildly affected. Two mechanisms can explain this unusual differential inhibitor effect. First, CNs inhibited activity by interacting with the CaMKII T-site (and thereby also interfered with NMDA-type glutamate receptor binding to the T-site). Because of this, the CaMKII region surrounding T286 competed with CNs for T-site interaction, whereas other substrates did not. Second, the intersubunit T286 autophosphorylation requires CaM binding both to the "kinase" and the "substrate" subunit. CNs dramatically decreased CaM dissociation, thus facilitating the ability of CaM to make T286 accessible for phosphorylation. Tat-fusion made CN21 cell penetrating, as demonstrated by a strong inhibition of filopodia motility in neurons and insulin secrection from isolated Langerhans' islets. These results reveal the inhibitory mechanism of CaM-KIIN and establish a powerful new tool for dissecting CaMKII function. INTRODUCTIONCa 2ϩ /calmodulin-dependent protein kinase II (CaMKII) is a multifunctional protein kinase best known for its critical role in learning and memory (for review, see Lisman and McIntyre, 2001;Soderling et al., 2001;Hudmon and Schulman, 2002;Lisman et al., 2002). CaMKII is highly expressed in the brain (Erondu and Kennedy, 1985), but at least one of its four isoforms (␣, ␤, ␥, and ␦) has been found in every cell type examined (Tobimatsu and Fujisawa, 1989;Bayer et al., 1999;Tombes et al., 2003). Numerous cellular functions of CaMKII have been described previously, both in and outside the nervous system. These include regulation of various ion channels (Worrell and Frizzell, 1991;Wang and Best, 1992;Roeper et al., 1997;Derkach et al., 1999;Dzhura et al., 2000), gene expression (Nghiem et al., 1994;Ramirez et al., 1997;Meffert et al., 2003), cell cycle/proliferation control (Baitinger et al., 1990;Patel et al., 1999;Matsumoto and Maller, 2002;Illario et al., 2003), apoptotic and excitotoxic cell death (Laabich and Cooper, 2000;Fladmark et al., 2002), cell morphology (Wu and Cline, 1998;Fink et al., 2003), and filopodia motility (Fink et al., 2003). CaMKII also has been implicated in regulation of insulin secretion (for review, see Easom, 1999); however, this conclusion is largely based on experiments using KN inhibitors, which also affect the Ca 2ϩ channels required for secretion (see below).CaMKII forms multimeric holoenzymes (Bennett et al., 1983;Kanaseki et al., 1991;Kolodziej et al., 2000;Morris and Torok, 2001;Hoel...

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MicroRNAs (miRNAs) are small, noncoding ∼22-nucleotide regulatory RNAs that are key regulators of gene expression programs. Their role in the context of the cardiovascular system has only recently begun to be explored; however, changes in the expression of miRNAs have been associated with cardiac development and with several pathophysiological states including myocardial hypertrophy and heart failure. We demonstrate that miRNA expression patterns are distinct in two types of heart failure: idiopathic dilated cardiomyopathy and ischemic cardiomyopathy. To pursue the observation that changes in expression levels of individual miRNAs are functionally relevant, microRNA mimics and inhibitors to miR-92, miR-100 and miR-133b were expressed in primary cultures of neonatal rat cardiac myocytes. These studies demonstrated that over expression of miR-100 is involved in the β-adrenergic receptor mediated repression of "adult" cardiac genes (i.e., α-myosin heavy chain, SERCA2a), and that over-expression of miR-133b prevents changes in gene expression patterns mediated by β-adrenergic receptor stimulation. In conclusion, some miRNA expression patterns appear to be unique to the etiology of cardiomyopathy and changes in the expression level of miR's 100 and 133b contribute to regulation of the fetal gene program. It is likely that this miR-directed reprogramming of key remodeling genes is involved in the establishment and progression of common human cardiomyopathies.

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