Human TRP-1 Has Tyrosine Hydroxylase but no DOPA Oxidase Activity

Human TRP-1 has been immunopurified from normal human melanocytes cultured from black neonatal subjects and used to investigate the catalytic function of TRP-1 for the two substrates, L-tyrosine and L-DOPA. Immunopurified TRP-1 did not demonstrate DOPA staining on SDS/PAGE nor DOPA oxidase (DO) activity with either routine or modified assays. The purified TRP-1 also demonstrated no tyrosine hydroxylase (TH) activity using the routine Pomerantz assay. However, there was apparent TH activity exhibited by immunopurified TRP-1 under conditions with low tyrosine concentration (≤0.8 μCi/ml of ³H-tyrosine), prolonged incubation time (i.e., overnight) and in the absence of the cofactor L-DOPA. Using these latter specific conditions, TH activity was also detected in cell lysates from a tyrosinase-negative albino melanocyte line which exhibited no TH activity with the routine Pomerantz assay. In addition, TH activity under low substrate assay conditions was not exhibited in a melanocyte line derived from a TRP-1 deficient, Brown albino individual. However, the absence of TH in this Brown albino cell line could be compensated for by the addition of L-DOPA to the assay. These results suggested that TRP-1 has some tyrosine hydroxylase but no DOPA oxidase activity. We propose that one function of TRP-1 is to modulate tyrosinase activity by making DOPA available as a cofactor to perpetuate the initial steps in melanogenesis.     

Description of a New Species of Microsporidia from Muscidifurax raptor (Hymenoptera: Pteromalidae), a Pupal Parasitoid of Muscoid Flies

ABSTRACT. A microsporidian parasite, Nosema muscidifuracis n. sp., has been found in Muscidifurax raptor , a parasitoid of muscoid flies. Stages of the parasite developed in direct contact with the host cell cytoplasm and were detected in midgut epithelium, Malpighian tubules, ovaries (including oocytes) and fat body of larvae and adults. Spores were also detected within eggs deposited on the host. Light and electron microscopy revealed a developmental cycle with diplokaryotic stages dividing by binary fission and disporous sporulation sequences producing diplokaryotic spores of three morphological classes, differing significantly only in length of the polar filament. Two of the classes were found in larvae, pupae and adults. One of these, with about five turns in the coiled polar filament, is presumed to be responsible for transmission from cell to cell within the host (autoinfection) and the other, with about 10 turns, responsible for transmission from host to host. A third class, with about 15 turns in the polar filament, was found in eggs of M. raptor . It is, presumably, either involved in initiation and spread of the infection at eclosion or is responsible for horizontal transmission to a new host individual when eggs are cannibalized.     

Mechanism of Isoxaben Tolerance in Agrostis palustris var. Penncross

Previous work has demonstrated that isoxaben tolerant mutantsof Arabidopsis thaliana var. Columbia are most likely alteredat the site of isoxaben binding. The salient question becomeswhether or not species selectivity to this herbicide might alsobe a result of differential target site binding. Grasses aregenerally more tolerant to isoxaben than dicots. In this communicationwe show that Agrostis palustris var. Penncross, a grass, is83-fold more tolerant in a soil incorporation test and 170-foldmore tolerant to inhibition of glucose incorporation into cellulosethan is Arabidopsis, a dicot. Cell wall fractionation of Agrostisshows a specific effect on cellulose biosynthesis. At most,5-fold of the 170-fold tolerance exhibited by Agrostis in termsof cellulose biosynthesis can be attributed to decreased isoxabenuptake under the test conditions. Furthermore, Agrostis is unableto metabolize isoxaben to any significant degree. Therefore,we suggest that the major portion of the tolerance in Agrostismight be due to differences in isoxaben binding. Key words: Isoxaben, cellulose, Arabidopsis, Agrostis, herbicide tolerance     

The influence of temperature on light enhanced glycoalkaloid synthesis in potato

The influence of temperature on total glycoalkaloid (TGA) synthesis in tubers exposed to light (250 jumol m“² s”² PAR, Photosynthetically Active Radiation) or dark environments for 96 h was examined in three potato cultivars. Following 96 h light or dark the tubers were stored without light at 5°C or 24°C and TGA concentrations monitored over the subsequent 30 and 90 days. Exposure to light and cultivar were found to be major factors influencing TGA concentrations; temperature had no significant effect. TGA content in illuminated tubers of cvs ‘Pentland Hawk’ and ‘Kerrs Pink’ were significantly higher (P < 0.01) compared with tubers placed in the dark. TGA concentrations in cv. ‘Desiree’ increased significantly only following exposure to light at low temperatures (P < 0.05). Removal of tubers from storage at 5°C and immediate illumination at 24°C altered the ratio of glycoalkaloids in cvs ‘Pentland Hawk’ and ‘Kerrs Pink’. Regardless of cultivar and storage temperature TGA concentrations were higher at the end of the storage period compared with initial TGA concentrations. During storage TGA concentrations fluctuated widely and gradual accumulation of glycoalkaloids with time was rarely demonstrated except in cv. ‘Desiree’. Tubers stored at 24°C accumulated higher TGA concentrations than those stored at 5°C in cv. ‘Kerrs Pink’ but not in cvs ‘Pentland Hawk’ and ‘Desiree’. Tubers of cv. ‘Kerrs Pink’ exposed to light prior to storage accumulated glycoalkaloids more rapidly than unexposed tubers during storage at 24°C and occasionally at 5°C. Light enhanced glycoalkaloids are not degraded over time.     

Histological,Biochemical, and Ultrastructural Studies on Hyperpigmented Human Skin Xenografts

The mechanisms for hyperpigmentation observed in human cutaneous xenografts placed on athymic nude mice was investigated. Histologic, biochemical, histochemical, and ultrastructural examinations were performed on human skin prior to grafting and at various times ranging from 2 weeks to 30 weeks post-grafting (PG). Hyperpigmentation was macroscopically visible on the graft as early as 4–6 weeks. The number of Dopa-positive melanocytes per unit area was increased at 2 weeks PG and remained elevated until 20 weeks PG. The surface area of the melanocytes, a measure of the activity of the cells, also increased significantly and remained above the pre-grafting size throughout the study. Western blot analysis using tyrosinase specific antibody (αTy-SP) revealed the presence of tyrosinase exclusively in the grafted skin from 2 weeks to 12 weeks PG tested. Histological and ultrastructural observations revealed the presence of numerous dendritic melanocytes, indeterminant clear cells suggestive of Langerhans cells, and dermal melanophages. The results of this study suggest that the observed hyperpigmentation in grafted tissue is caused by an increase in the number of Dopa-positive melanocytes and probably from enhanced melanin production. Extracts of proteins from the xenografts exhibited prominent differences in low and high molecular proteins between pre- and post-grafted skin. Among them, the exclusive appearance of a protein doublet with apparent mw ～14 kDa was found in grafted skin, and subsequent studies indicate it has potent effects on melanocyte function.