Role of Melanin in Release of Extracellular Enzymes and Selection of Aggressive Isolates of Bipolaris sorokiniana in Barley

Eighteen barley isolates of Bipolaris sorokiniana belonging to wild and clonal type of black, mixed and white subpopulations were quantitatively assayed for their melanin content and aggressiveness with respect to production of some of the extracellular enzymes such as cellulase, pectinase, amylase and protease. Cellulase and pectinase constituted major portion of the enzymes recovered from the black, mixed and white isolates. Enzyme production and aggressiveness were relatively higher in melanin devoid or low melanin isolates. The melanin deficient isolates were also differentiated from black and mixed isolates on the basis of variation in internal transcribed spacer region of the ribosomal DNA. Higher enzyme productions positively correlated with area under disease progress curve (AUDPC) and lesion development. Melanin content was negatively correlated with extracellular enzymes and aggressiveness of the isolates. Based on melanin content, lesion size, AUDPC and extracellular enzymes, the isolates were grouped in two major clusters (I and II) with further division of cluster II into two sub-clusters (II-A and II-B). The results appears to indicate a possible role of melanin in release of extracellular enzymes and hence in evolution and selection of aggressive isolates of B. sorokiniana in barley.     

Interactions of the receptor for insulin-like growth factor II with mannose-6-phosphate and antibodies to the mannose-6-phosphate receptor

Recently, the sequence of the human receptor for insulin-like growth factor II (IGF-II) was found to be 80% identical [Morgan et al., (1987) Nature 329, 301-307] to the sequence of a partial clone of the bovine cation-independent mannose-6-phosphate receptor [Lobel et al., (1987) Proc. Natl. Acad. Sci. USA 84, 2233-2237]. In the present study, the purified receptor for insulin-like growth factor II (IGF-II) was found to react with two different polyclonal antibodies to the purified mannose-6-phosphate receptor. Moreover, mannose-6-phosphate was found to stimulate the binding of labeled IGF-II to the IGF-II receptor by two-fold. This effect had the same specificity and affinity as the reported binding of mannose-6-phosphate to its receptor; mannose-1-phosphate and mannose had no effect on the binding of labeled IGF-II to its receptor, and the half-maximally effective concentration of mannose-6-phosphate was 0.3 mM. Also, mannose-6-phosphate did not affect labeled IGF-II binding to the insulin receptor. These results support the hypothesis that a single protein of Mr-250,000 binds both IGF-II and mannose-6-phosphate. Furthermore, they indicate that mannose-6-phosphate can modulate the interaction of IGF-II to its receptor.     

Expression and characterization of a functional human insulin-like growth factor I receptor

Stable transfectants of Chinese hamster ovary (CHO) cells were developed that expressed the protein encoded by a human insulin-like growth factor I (IGF-I) receptor cDNA. The transfected cells expressed approximately 25,000 high affinity receptors for IGF-I (apparent Kd of 1.5 X 10(-9) M), whereas the parental CHO cells expressed only 5,000 receptors per cell (apparent Kd of 1.3 X 10(-9) M). A monoclonal antibody specific for the human IGF-I receptor inhibited IGF-I binding to the expressed receptor and immunoprecipitated polypeptides of apparent Mr values approximately 135,000 and 95,000 from metabolically labeled lysates of the transfected cells but not control cells. The expressed receptor was also capable of binding IGF-II with high affinity (Kd approximately 3 nM) and weakly recognized insulin (with about 1% the potency of IGF-I). The human IGF-I receptor expressed in these cells was capable of IGF-I-stimulated autophosphorylation and phosphorylation of endogenous substrates in the intact cell. This receptor also mediated IGF-I-stimulated glucose uptake, glycogen synthesis, and DNA synthesis. The extent of these responses was comparable to the stimulation by insulin of the same biological responses in CHO cells expressing the human insulin receptor. These results indicate that the isolated cDNA encodes a functional IGF-I receptor and that there are no inherent differences in the abilities of the insulin and IGF-I receptors to mediate rapid and long term biological responses when expressed in the same cell type. The high affinity of this receptor for IGF-II also suggests that it may be important in mediating biological responses to IGF-II as well as IGF-I.     

Formation of wheat protein bodies: Involvement of the Golgi apparatus in gliadin transport

Developing wheat (Triticum aestivum L.) endosperm was examined using ultrathin sections prepared from tissues harvested at 5, 9, 16 and 25 d after flowering. Protein bodies were evident by 9 d and displayed a variety of membranous structures and inclusions. The Golgi apparatus was a prominent organelle at all stages, and by 9 d was associated with small electron-dense inclusions. By immunocytochemical techniques, gliadin (wheat prolamine) was localized within these vesicles and in homogeneous regions of protein bodies, but not in the lumen of the rough endoplasmic reticulum. The protein bodies appear to enlarge by fusion of smaller protein bodies resulting in larger, irregular-shaped organelles. The affinity of the Golgi-derived vesicles for gliadin-specific probes during the period of maximal storage-protein synthesis and deposition indicates that this organelle includes the bulk, if not all, of the gliadin produced. The involvement of the Golgi apparatus in the packaging of gliadins into protein bodies indicates a pathway which differs from the mode of prolamine deposition in other cereals such as maize, rice and sorghum, and resembles the mechanism employed for the storage of rice glutelin and legume globulins.Abbreviations ER endoplasmic reticulum - IgG immunoglobulin G - DAF days after flowering     

Enhanced degradation of gamma-irradiated lignocelluloses by a new xylanolytic Flavobacterium sp. isolated from soil

An extracellular chitosanase produced by Rhodotorula gracilis CFR-1 that catalyses a limited degradation of chitosan with no detectable generation of glucosamine or reducing groups was identified. Ultracentrifugation, polyacrylamide gel electrophoresis and gel permeation studies suggest that chitosan of average molecular mass 36000 Da was reduced by the enzymic catalysis to nearly one-fourth this size without further hydrolysis of the products. The enzyme, produced constitutively by this yeast, was partially purified and some of its properties were studied.     

Metastasis-associated alterations in phospholipids and fatty acids of human prostatic adenocarcinoma cell lines.

Metastatic variants of human prostatic adenocarcinoma cell lines (DU-145, LNCaP, and ND-1) were studied by using soft agar colony forming efficiency, nude mice tumorigenicity, in vitro invasion assay, and type IV collagenase assay. The DU-145 and ND-1 cell line showed higher metastatic potential than LNCaP. Lipids from DU-145, ND-1, and LNCaP cells were extracted and analyzed by thin-layer chromatography and gas-liquid chromatography. The major lipids were phosphatidylcholine, phosphatidylethanolamine, sphingomyelin, fatty acids, and cholesterol. The sphingomyelin level was significantly higher in highly metastatic cells (DU-145 and ND-1) compared with the lower metastatic variant (LNCaP). The increase in the synthetic pathway and decrease in degradation pathway of sphingomyelin in microsomal fractions was sufficient to account for the measured increase in sphingomyelin in DU-145 cells compared with LNCaP cells. The major fatty acids of these lipids were palmitic (16:0), stearic (18:0), oelic (18:1), and arachidonic acid (20:4). The arachidonic acid level was significantly decreased in DU-145 and ND-1 compared with LNCaP cells. Electron microscopic studies showed no significant changes in the morphology of DU-145, ND-1, and LNCaP cells. The results of these investigations demonstrate for the first time that sphingomyelin and arachidonic acid contents are different in high and low metastatic variants of human prostatic adenocarcinoma cell lines.     

Insulin-degrading enzyme is capable of degrading receptor-bound insulin

In the investigation of the intracellular sites of insulin degradation, it might be important whether receptor-bound insulin could be a substrate for insulin-degrading enzyme (IDE). Insulin receptor and IDE were purified from rat liver using a wheat germ agglutinin column and monoclonal anti-IDE antibody affinity column, respectively. [125I]insulin-receptor complex was incubated with various amounts of IDE at 0 degree C in the presence of disuccinimidyl suberate and analyzed by reduced 7.5% SDS-PAGE and autoradiography. With increasing amounts of IDE, the radioactivity of 135 kd band (insulin receptor alpha-subunit) decreased, whereas that of 110 kd band (IDE) appeared then gradually increased, suggesting that IDE could bind to receptor-bound insulin. During incubation of insulin-receptor complex with IDE at 37 degrees C, about half of the [125I]insulin was dissociated from the complex. However, the time course of [125I]insulin degradation in this incubation was essentially identical to that of free [125I]insulin degradation. Cross-linked, non-dissociable receptor-bound [125I]insulin was also degraded by IDE. Rebinding studies to IM-9 cells showed that the receptor binding activity of dissociated [125I]insulin from insulin-receptor complex incubated with IDE was significantly (p less than 0.001) decreased as compared with that without the enzyme. These results, therefore, show that IDE could recognize and degrade receptor-bound insulin, and suggest that IDE may be involved in insulin metabolism during receptor-mediated endocytosis through the degradation of receptor-bound insulin in early neutral vesicles before their internal pH is acidified.     

Histology of the pituitary gland of the grey mullet, Mugil cephalus L.

The present communication deals with a histological study of the pituitary gland of the teleost fish Mugil cephalus , found in the estuarine waters of Cochin area. Six different cell types were identified in the pituitary gland on the basis of their grouping, distribution and staining properties. The prolactin and the TSH cells (thyroid stimulating hormone producing cells or thyrotrops) were identified in the rostral pars distalis and the ACTH cells (adrenocorticotropic hormone producing cells or corticotrops) in the interphase between the neurohypophysis and the rostral pars distalis. The STH cells (somatotropic hormone producing cells or somatotrops) and the gonadotropic cells were distinguished in the proximal pars distalis and the MSH cells (melanin stimulating hormone producing cells or melanotrops) in the pars intermedia.