Time-resolved fluoroimmunoassay of potato virus M with monoclonal antibodies

Mouse monoclonal antibodies (MAbs) specific for potato virus M (PVM) were prepared and the properties of three of them were studied. MAb M4C1 is IgG2b, it binds with high affinity to PVM coat protein, to purified virus preparations and recognises PVM in infected potato leaves and tubers. MAb M6D5 is IgG2a and also reacts with PVM coat protein, purified PVM and with PVM in potato leaf and tuber extracts. In double-antibody sandwich ELISA (DAS ELISA) MAbs M4C1 and M6D5 reacted with all 17 PVM isolates tested. MAb M7 is IgG2b and recognises PVM only in indirect dot ELISA on nitrocellulose filters and viral coat protein on Western blots. MAbs against PVM were used as capture antibodies and europium-labelled MAbs as conjugates in time-resolved fluoroimmunoassay (EuTRFIA). The standard EuTRFIA curve of PVM detection is approximately linear over a range of PVM concentrations from 0.5 ng/ml to 1000 ng/ml. The lowest PVM concentration detectable in EuTRFIA was 0.5 ng/ml and correspondingly 6 ng/ml in DAS ELISA. The use of the europium chelate label allows PVM detection in potato leaf and tuber sap at dilutions greater than 10^--⁴ with very low background fluorescence. EuTRFIA with MAbs, with either one or two incubations is about 10–20 times more sensitive for PVM detection than is DAS ELISA. PVM and PVX, mixed with healthy potato tuber sap, were simultaneously tested in a single sample at concentrations lower than 10 ng/ml by double-label TRFIA using europium-labelled MAbs to PVM and samarium-labelled MAbs to PVX.     

Genome size in gymnosperms

The DNA 2C and per chromosome values of 57 species belonging to 22 genera of gymnosperms have been analysed. The overall range is 12-fold with a modal value of about 30.0 pg.Cycadales exhibit a 2-fold difference. AmongConiferales with a 4-fold variation, thePinaceae have higher mean DNA contents as well as a greater range and diversity than other families. Remarkable interspecific differences are found inCycas, Picea, Larix, Pinus, Callitris, Cupressus, andChamaecyparis. Despite this, there is a constancy of basikaryotypes within these genera.Gnetum shows a distinctly low DNA value.     

Correlations between 2C DNA values and habit inCassia (Leguminosae:Caesalpinioideae)

Ten species of the genusCassia show a range of 2C DNA amounts from 1.30 to 2.54 pg at the same ploidy level. Remarkably, a distinct 2-fold increase is depicted by an arboreal speciesC. excelsa while the rest comprising of herbs, trees and shrubs have a range from 1.30 to 1.47 pg. These form a natural grouping with respect to mean DNA amounts which differ by 0.05 pg in the herbs, trees and shrubs respectively.     

Molecular differentiation of Chenopodium album complex and some related species using ISSR profiles and ITS sequences

The present study was undertaken to understand the genetic differentiation and relationships in various components of C. album complex, C. giganteum and some related species using inter simple sequence repeats (ISSR) profiles and internal transcribed spacer (ITS) sequences. The relationships based on UPGMA dendrograms have shown the heterogenous nature of C. album complex. The 2x taxa while showing close relation among themselves are sharply segregated from 4x and 6x taxa belonging to C. album and C. giganteum. Among the three cytotypes from North Indian plains the 4x shows greater similarity to 6x than to 2x which is corroborated by the karyotypic studies. Furthermore, the 6x C. album and C. giganteum accessions of American and European origin are clearly segregated from those of Indian origin which may show their separate origin. Other related species show relationships according to their taxonomic position. The present study based on ISSR profiles and ITS sequences has therefore been very useful in explaining the relationships between various components of C. album complex and related species. However, more work needs to be done using different CpDNA loci to define correct species boundary of the taxa under C. album complex from Himalayas and North Indian Plains.     

The melanosomal protein PMEL17 as a target for antibody drug conjugate therapy in melanoma

Melanocytes uniquely express specialized genes required for pigment formation, some of which are maintained following their transformation to melanoma. Here we exploit this property to selectively target melanoma with an antibody drug conjugate (ADC) specific to PMEL17, the product of the SILV pigment-forming gene. We describe new PMEL17 antibodies that detect the endogenous protein. These antibodies help define the secretory fate of PMEL17 and demonstrate its utility as an ADC target. Although newly synthesized PMEL17 is ultimately routed to the melanosome, we find substantial amounts accessible to our antibodies at the cell surface that undergo internalization and routing to a LAMP1-enriched, lysosome-related organelle. Accordingly, an ADC reactive with PMEL17 exhibits target-dependent tumor cell killing in vitro and in vivo.     

Nuclear DNA amounts in 112 species of tropical hardwoods -- new estimates

The 4C DNA values of 112 species, belonging to 37 families have a range from 0.83 pg (Bixa orellana) to 15.54 pg (Thryallis angustifolia), showing a 18.72-fold variation. The genome size varies from 0.21 pg (Bixa orellana) to 3.32 (Thespesia populnea), with a 15.8-fold difference. The Bombacaceae has the minimum range (1.08-fold) of variation, while the maximum (5.0-fold) is shown by the Fabaceae. The Boraginaceae, Lauraceae, Malpighiaceae, and Malvaceae generally have higher 4C DNA values of > 10 pg, while the Bixaceae, Caricaceae, Oxalidaceae, and Santalaceae have lower values of < 2.0 pg. These data add further to our knowledge on variation in DNA amount in tropical hardwoods.     

Jasmonic acid-induced tolerance to root-knot nematodes in tomato plants through altered photosynthetic and antioxidative defense mechanisms

Plant parasitic nematodes cause severe damage to cultivated crops globally. Management of nematode population is a major concern as chemicals used as nematicides have negative impact on the environment. Natural plant products can be safely used for the control of nematodes. Among various plant metabolites, plant hormones play an essential role in developmental and physiological processes and also assist the plants to encounter stressful conditions. Keeping this in mind, the present study was designed to evaluate the effect of jasmonic acid (JA) on the growth, pigments, polyphenols, antioxidants, osmolytes, and organic acids under nematode infection in tomato seedlings. It was observed that nematode inoculation reduced the growth of seedlings. Treatment with JA improved root growth (32.79%), total chlorophylls (71.51%), xanthophylls (94.63%), anthocyanins (37.5%), and flavonoids content (21.11%) when compared to inoculated seedlings alone. The JA application enhanced the total antioxidant capacity (lipid- and water-soluble antioxidants) by 38.23 and 34.37%, respectively, in comparison to infected seedlings. Confocal studies revealed that there was higher accumulation of glutathione in hormone-treated seedlings under nematode infection. Treatment with JA increased total polyphenols content (74.56%) in comparison to nematode-infested seedlings. JA-treated seedlings also enhanced osmolyte and organic acid contents under nematode stress. Overall, treatment with JA improved growth, enhanced pigment levels, modulated antioxidant content, and enhanced osmolyte and organic acid content in nematode-infected seedlings.     

Procathepsin D secreted by HaCaT keratinocyte cells - A novel regulator of keratinocyte growth

Procathepsin D (pCD), the precursor form of lysosomal aspartic protease, is overexpressed and secreted by various carcinomas. The fact that secreted pCD plays an essential role in progression of cancer has been established. In this study, we describe substantial secretion of pCD by the human keratinocyte cell line HaCaT, under serum-free conditions. Moreover, exogenous addition of purified pCD enhanced the proliferation of HaCaT cells. The proliferative effect of pCD was inhibited by a monoclonal antibody against the activation peptide (AP) of pCD. Treatment of HaCaT cells with pCD or AP led to the secretion of a set of cytokines that might promote the growth of cells in a paracrine manner. The role of secreted pCD and its mechanism of action were studied in a scratch wound model and the presence of pCD and AP enhanced regeneration, while this effect was reversed by the addition of anti-AP antibody. Expression and secretion of pCD was upregulated in HaCaT cells exposed to various stress conditions. Taken together, our results strongly suggest that the secretion of pCD is not only linked to cancer cells but also plays a role in normal physiological conditions like wound healing and tissue remodeling.     

Structure-activity relationship studies of flavonoids as potent inhibitors of human platelet 12-hLO, reticulocyte 15-hLO-1, and prostate epithelial 15-hLO-2

Human lipoxygenase (hLO) isozymes have been implicated in a number of disease states and have attracted much attention with respect to their inhibition. One class of inhibitors, the flavonoids, have been shown to be potent lipoxygenase inhibitors but their study has been restricted to those compounds found in nature, which have limited structural variability. We have therefore carried out a comprehensive study to determine the structural requirements for flavonoid potency and selectivity against platelet 12-hLO, reticulocyte 15-hLO-1, and prostate epithelial 15-hLO-2. We conclude from this study that catechols are essential for high potency, that isoflavones and isoflavonones tend to select against 12-hLO, that isoflavons tend to select against 15-hLO-1, but few flavonoids target 15-hLO-2.