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The clathrin-associated protein complex 2 (AP-2 complex) is a group of proteins associated with clathrincoated vesicles and believed to interact with cytoplasmic domains of receptors found in the plasma membrane. AP-2 was purified as an assembly of several polypeptide chains (a, (, AP50, and AP17), of which only the a and 18 chains (100-115 kDa) show significant heterogeneity. We have obtained cDNA clones for two distinct rat brain (3 chains. We have also studied the domain organization of bovine brain AP-2 complexes by selective proteolysis. Results of these studies show that the a and 13 chains have a similar two-domain organization. Their amino-terminal domains are relatively invariant whereas their carboxyl-terminal domains are variable in both sequence and length. We propose that the variable domains select receptors for inclusion in coated vesicles.Clathrin-coated pits and -coated vesicles are associated at the plasma membrane with the early stages of receptor-mediated endocytosis (for review, see ref. 1) and in the Golgi region with organization of the nonconstitutive pathway of exocytosis (for review, see ref.2). The major structural proteins of the coat are the heavy and light chains of clathrin (3-5) and the clathrin assembly proteins (6-8), referred to herein as associated proteins (APs). Together, these species represent "90% of the protein content of the coat. The APs are complexes made up of several kinds of polypeptide chains, large (100-115 kDa), medium (45-50 kDa), and small (17-20 kDa). Two major families of AP complexes, designated AP-1 and AP-2 (9), have been found in close association with the trans-Golgi and plasma membrane, respectively (7,8 (8,11,12). A similar picture also appears to characterize complexes of the AP-1 family, which contain y and P3' large chains associated with medium and small chains, known, respectively, as AP47 and AP19 (8). Thus, the AP complexes display a heterogeneity that suggests a role in interaction with membrane proteins destined for endocytosis or export. Their location, toward the membrane side of the hollow clathrin lattice (13,14), is also consistent with such a function.We report here the molecular cloning and primary structure of two distinct rat brain AP-2 P3 chains.** We also describe the proteolytic dissection of bovine brain AP-2 complexes. We combine these results to establish a bipartite structure for the a and P chains in the AP complex. We show that the core of an AP-2 complex contains, in addition to AP50 and AP17, the amino-terminal domains of the a and /3chains. The carboxyl-terminal domains of the two /8 chains we have cloned differ more in sequence than do their amino-terminal domains. Moreover, the proteolytic dissection experiments show that in the a chains, it is the carboxylterminal domains that are responsible for significant size heterogeneity. On the basis of these results, we present a model for the organization of the AP complexes and for the functions of each of their two major parts. METHODS Clathrin and AP-2 Complexes. Cl...

Sangre de Grado extract used by Peruvian natives as a cicatrizant agent, was collected from trees of the species Croton lechleri growing in the Peruvian jungle. The Sangre de Grado was found to contain one alkaloid identified as taspine and which was shown to be the active cicatrizant principle by an in vivo test in mice. This alkaloid exhibited a dose-related cicatrizant effect and an ED50 of 0.375 mg/kg. Experiments with taspine hydrochloride in order to study its mechanism of action in cell culture systems showed that the alkaloid was non-toxic to human foreskin fibroblasts at concentrations below 150 ng/ml and that it had no effect on cell proliferation. On the other hand, taspine hydrochloride was found to increase the migration of human foreskin fibroblasts. This effect on the migration of fibroblasts is probably the mechanism by which Sangre de Grado and taspine hydrochloride accelerate the wound healing process. Using the two-stage mouse skin carcinogenesis system, we have been able to show that neither Sangre de Grado nor taspine hydrochloride had carcinogenic or tumour promoter activity after 17 months of treatment.

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Bioassay-guided fractionation of the EtOH extract of the bark of Uncaria guianensis (Aubl.) Gmel (Rubiaceae) using a yeast-based assay for DNA-damaging agents has furnished the two weakly but selectively active indole alkaloids uncarine C (1) and uncarine E (2) as the major bioactive constituents in this assay.