SUMMARY Adrenergic stimulation of the heart engages cAMP and phosphoinositide second messenger signaling cascades. Cardiac phosphoinositide 3-kinase p110γ participates in these processes by sustaining β-adrenergic receptor internalization through its catalytic function and by controlling phosphodiesterase 3B (PDE3B) activity via an unknown kinase-independent mechanism. We have discovered that p110γ anchors protein kinase A (PKA) through a site in its N-terminal region. Anchored PKA activates PDE3B to enhance cAMP degradation and phosphorylates p110γ to inhibit PIP3 production. This provides local feedback control of PIP3 and cAMP signaling events. In congestive heart failure, p110γ is upregulated and escapes PKA-mediated inhibition, contributing to a reduction in β-adrenergic receptor density. Pharmacological inhibition of p110γ normalizes β-adrenergic receptor density and improves contractility in failing hearts.

The diaphanous gene is the founding member of a family of Diaphanous-related formin proteins (DRFs). We identified diaphanous in a screen for genes that are necessary for the normal growth and stabilization of the Drosophila neuromuscular junction (NMJ). Here, we demonstrate that diaphanous mutations perturb synaptic growth at the NMJ. Diaphanous protein is present both presynaptically and postsynaptically. However, genetic rescue experiments in combination with additional genetic interaction experiments support the conclusion that dia is necessary presynaptically for normal NMJ growth. We then document defects in both the actin and microtubule cytoskeletons in dia mutant nerve terminals. In so doing, we define and characterize a population of dynamic pioneer microtubules within the NMJ that are distinct from the bundled core of microtubules identified by the MAP1b-like protein Futsch. Defects in both synaptic actin and dynamic pioneer microtubules are correlated with impaired synaptic growth in dia mutants. Finally, we present genetic evidence that Dia functions downstream of the presynaptic receptor tyrosine phosphatase Dlar and the Rho-type GEF (guanine nucleotide exchange factor) trio to control NMJ growth. Based on the established function of DRFs as Rho-GTPase-dependent regulators of the cell cytoskeleton, we propose a model in which Diaphanous links receptor tyrosine phosphatase signaling at the plasma membrane to growth-dependent modulation of the synaptic actin and microtubule cytoskeletons.

A cell's response to its environment is often determined by signaling through the actions of enzyme cascades. The ability to organize these enzymes into multiprotein complexes allows for a high degree of fidelity, efficiency and spatial precision in signaling responses.

Mixtures of poly(ethylene terephthalate) and poly(ethylene naphthalene-2,6-dicarboxylate) have been transesterified at temperatures between 553 and 573 K. The partially transesterified polymers were analyzed using 1 H NMR, and the rate constants for transesterification were obtained using a kinetic expression based on a second-order reversible reaction mechanism. The hydroxyl end groups have been shown to have a significant effect on the kinetics of the reaction by comparing rate constants obtained for the same polyester mixture but with hydroxyl end groups quantitatively esterified by trifluoroacetic acid. Carboxyl end groups do not have such a significant role in transesterification, and from the variation of rate constant at one temperature for a series of PET polymers with differing carboxyl to hydroxyl end groups, it appears that transesterification by direct ester-ester interchange is very small. The NMR data have also been used to calculate the randomness factor and number-average sequence length of the terephthalate sequences in the copolymer as transesterification progresses. The initially immiscible polyester mixture becomes miscible after a critical amount of transesterification takes place; no matter what reaction temperature is used, this homogenization takes place when the randomness factor has a value of 0.5. This does not correspond to completely random copolymer formation but suggests that the number-average sequence length of the terephthalate units in the copolyester is ca. 4-8.