Amyloid deposits localized to the islets of Langerhans are typical of non-insulin-dependent human diabetes mellitus and of diabetes mellitus in adult cats. Amyloid deposits also commonly occur in insulin-producing pancreatic tumors. We have purified a major protein-insulinoma or islet amyloid polypeptide (IAPP)-from human and cat islet amyloid and from amyloid of a human insulinoma. IAPP from human insulinoma contained 37 amino acid residues and had a theoretical molecular mass of 3850 Da. The amino acid sequence is unique but has >40% identity with the human calcitonin gelie-related peptide. A partial amino acid sequence of cat islet IAPP corresponding to positions 1-27 of human insulinoma IAPP was identical to the human IAPP except for substitutions in three positions. An antiserum raised to a synthetic human insulinoma IAPP-(7-17) undecapeptide showed specific immunohistochemical reactivity with human and cat islet amyloid and with islet B cells.

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Cloning and sequencing of the complementary DNA for platelet-derived endothelial cell growth factor indicates that it is a novel factor distinct from previously characterized proteins. The factor, a protein with a relative molecular mass of about 45,000, stimulates endothelial cell growth and chemotaxis in vitro and angiogenesis in vivo.

Hormone-sensitive lipase (HSL) is the rate-limiting enzyme in lipolysis. Stimulation of rat adipocytes with isoproterenol results in phosphorylation of HSL and a 50-fold increase in the rate of lipolysis. In this study, we used site-directed mutagenesis and two-dimensional phosphopeptide mapping to show that phosphorylation sites other than the previously identified Ser-563 are phosphorylated in HSL in response to isoproterenol stimulation of 32 P-labeled rat adipocytes. Phosphorylation of HSL in adipocytes in response to isoproterenol and in vitro phosphorylation of HSL containing Ser 3 Ala mutations in residues 563 and 565 (S563A,S565A) with protein kinase A (PKA), followed by tryptic phosphopeptide mapping resulted in two tryptic phosphopeptides. These tryptic phosphopeptides co-migrated with the phosphopeptides released by the same treatment of F 654 HPRRSSQGVLHMPLYSSPIVK 675 phosphorylated with PKA. Analysis of the phosphorylation site mutants, S659A, S660A, and S659A,S660A disclosed that mutagenesis of both Ser-659 and Ser-660 was necessary to abolish the activation of HSL toward a triolein substrate after phosphorylation with PKA. Mutation of Ser-563 to alanine did not cause significant change of activation compared with wild-type HSL. Hence, our results demonstrate that in addition to the previously identified Ser-563, two other PKA phosphorylation sites, Ser-659 and Ser-660, are present in HSL and, furthermore, that Ser-659 and Ser-660 are the major activity controlling sites in vitro.Free fatty acids stored as triacylglycerols in the adipocytes comprise the quantitatively most important energy substrate in mammals. Hormone-sensitive lipase (HSL) 1 (EC 3.1.1.3) catalyzes the first and rate-limiting step in the hydrolysis of stored triacylglycerols and is thereby a key enzyme in the mobilization of free fatty acids from adipose tissue (1). The hormonal and neural control of lipolysis is exerted by regulation of HSL activity, mediated by reversible phosphorylation (2, 3). In response to catecholamines and other fast-acting lipolytic hormones, HSL is activated through the phosphorylation by protein kinase A (PKA). The major antilipolytic hormone insulin prevents cAMP-mediated phosphorylation and activation of HSL (3). The antilipolytic effect of insulin is brought about mainly by activation of phosphodiesterase 3B (4).HSL has in intact rat adipocytes been reported to be phosphorylated at two sites (3). These sites were later identified as Ser-563 (5) and Ser-565 (6). In hormonally quiescent cells, only Ser-565 is phosphorylated (3). Stimulation with noradrenaline increases the phosphorylation extent of Ser-563 to that of Ser-565 (3). Ser-563 is also phosphorylated in vitro by PKA (5). Ser-565 has been shown to be phosphorylated in vitro by the 5Ј-AMP-activated kinase (6). This kinase, which phosphorylates and regulates the activity of other key enzymes in lipid metabolism (7), acetyl-CoA carboxylase (fatty acid synthesis), and 3-hydroxy-3-methylglutaryl-CoA (cholesterol synthesis), has also been suggested ...

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