Using degenerate polymerase chain reaction, we isolated a cDNA encoding a novel 493-amino acid protein from human and mouse adult heart cDNAs and have designated it angiopoietin-related protein-2 (ARP2). The NH 2 -terminal and COOH-terminal portions of ARP2 contain the characteristic coiled-coil domain and fibrinogen-like domain that are conserved in angiopoietins. ARP2 has two consensus glycosylation sites and a highly hydrophobic region at the NH 2 terminus that is typical of a secretory signal sequence. Recombinant ARP2 expressed in COS cells is secreted and glycosylated. In human adult tissues, ARP2 mRNA is most abundant in heart, small intestine, spleen, and stomach. In rat embryos, ARP2 mRNA is most abundant in the blood vessels and skeletal muscles. Endothelial and vascular smooth muscle cells also contain ARP2 mRNA. Recombinant ARP2 protein induces sprouting in vascular endothelial cells but does not bind to the Tie1 or Tie2 receptor. These results suggest that ARP2 may exert a function on endothelial cells through autocrine or paracrine action.

Using degenerate PCR we isolated a cDNA encoding a novel 406- and 410-amino acid protein from human and mouse embryonic cDNAs and have designated it 'hepatic fibrinogen/angiopoietin-related protein' (HFARP). The N-terminal and C-terminal portions of HFARP contain the characteristic coiled-coil domains and fibrinogen-like domains that are conserved in angiopoietins. In human and mouse tissues, HFARP mRNA is specifically expressed in the liver. HFARP mRNA and protein are mainly present in the hepatocytes. HFARP has a highly hydrophobic region at the N-terminus that is typical of a secretory signal sequence and one consensus glycosylation site. Recombinant HFARP expressed in COS-7 cells is secreted and glycosylated. HFARP protein is present not only in the hepatocytes, but also in the circulating blood. Recombinant HFARP acts as an apoptosis survival factor for vascular endothelial cells, but does not bind to Tie1 or Tie2 (endothelial-cell tyrosine kinase receptors). These results suggest that HFARP may exert a protective function on endothelial cells through an endocrine action.

Abstract-Tropomodulin is a tropomyosin-binding protein that terminates "pointed-end" actin filament polymerization. To test the hypothesis that regulation of tropomodulin:actin filament stoichiometry is critical for maintenance of actin filament length, tropomodulin levels were altered in cells by infection with recombinant adenoviral expression vectors, which produce either sense or antisense tropomodulin mRNA. Neonatal rat cardiomyocytes were infected, and sarcomeric actin filament organization was examined. Confocal microscopy indicated that overexpression of tropomodulin protein shortened actin filaments and caused myofibril degeneration. In contrast, decreased tropomodulin content resulted in the formation of abnormally long actin filament bundles. Despite changes in myofibril structure caused by altered tropomodulin expression, total protein turnover of the cardiomyocytes was unaffected. Biochemical analyses of infected cardiomyocytes indicated that changes in actin distribution, rather than altered actin content, accounted for myofibril reorganization. Ultrastructural analysis showed thin-filament disarray and revealed the presence of leptomeres after tropomodulin overexpression. Tropomodulin-mediated effects constitute a novel mechanism to control actin filaments, and our findings demonstrate that regulated tropomodulin expression is necessary to maintain stabilized actin filament structures in cardiac muscle cells. (Circ Res. 1998;82:94-105.)A ccurate control of thin-filament length in sarcomeres is critical for proper function of the contractile apparatus. Uniformity among actin filaments depends on consistent length specification and effective termination of polymerization. Tropomodulin has been implicated as the regulatory agent for inhibiting slowly growing (pointed) end elongation of actin filaments. Located at or near the ends of thin filaments in skeletal muscle at a calculated stoichiometry of 1.2 to 1.6 tropomodulin molecules per actin filament, tropomodulin is situated in the appropriate place at the right concentration to regulate thin filaments.1 Circumstantial evidence suggests that tropomodulin:actin filament stoichiometry is critical for maintaining actin filament structure. First, the stoichiometry of tropomodulins per actin filament is comparable between skeletal muscle and the erythrocyte cytoskeleton, despite vastly different actin organization in the two cell types.1 Second, tropomodulin releases tropomyosin from association with actin filaments in vitro when added in molar excess.2 Third, tropomyosin-coated actin filaments are stabilized by interaction with tropomodulin when both are present in equivalent stoichiometric levels. 3 The importance of sarcomeric protein stoichiometry for muscle function and the regulation of sarcomeric organization is suggested from studies that found mutations of either ␣-tropomyosin or cardiac troponin T as the basis for the characteristic myofibrillar disorganization observed in familial hypertrophic cardiomyopathy. 4 Tropomodulin is a component of t...