DNA-Launched Alphavirus Replicons Encoding a Fusion of Mycobacterial Antigens Acr and Ag85B Are Immunogenic and Protective in a Murine Model of TB Infection

There is an urgent need for effective prophylactic measures against Mycobacterium tuberculosis (Mtb) infection, particularly given the highly variable efficacy of Bacille Calmette-Guerin (BCG), the only licensed vaccine against tuberculosis (TB). Most studies indicate that cell-mediated immune responses involving both CD4+ and CD8+ T cells are necessary for effective immunity against Mtb. Genetic vaccination induces humoral and cellular immune responses, including CD4+ and CD8+ T-cell responses, against a variety of bacterial, viral, parasitic and tumor antigens, and this strategy may therefore hold promise for the development of more effective TB vaccines. Novel formulations and delivery strategies to improve the immunogenicity of DNA-based vaccines have recently been evaluated, and have shown varying degrees of success. In the present study, we evaluated DNA-launched Venezuelan equine encephalitis replicons (Vrep) encoding a novel fusion of the mycobacterial antigens α-crystallin (Acr) and antigen 85B (Ag85B), termed Vrep-Acr/Ag85B, for their immunogenicity and protective efficacy in a murine model of pulmonary TB. Vrep-Acr/Ag85B generated antigen-specific CD4+ and CD8+ T cell responses that persisted for at least 10 wk post-immunization. Interestingly, parenterally administered Vrep-Acr/Ag85B also induced T cell responses in the lung tissues, the primary site of infection, and inhibited bacterial growth in both the lungs and spleens following aerosol challenge with Mtb. DNA-launched Vrep may, therefore, represent an effective approach to the development of gene-based vaccines against TB, particularly as components of heterologous prime-boost strategies or as BCG boosters.     

In vitro development of angiosperm floral buds and organs

In the last twenty-five years, young inflorescences, floral buds and individual floral organs of a number of species have been cultured in vitro. There is considerable variability in the requirement of plant growth regulators and nutritional factors for flower development of different species. This variability is compounded by the fact that the hormonal and nutritional requirements are different at various stages of organ and floral development. Experimental studies on normal and mutant flowers in vitro have provided insights into some of the regulatory processes in floral organogenesis. The potential use of the in vitro technique in elucidating the various mechanisms in flower development is stressed.     

Microsporogenesis in a normal line and in the ogu cytoplasmic male-sterile line of Brassica napus

Summary In the ogu cytoplasmic male-sterile (CMS) line of Brassica napus, stamen morphology was influenced by temperature conditions. Under a high temperature regime (27° C/23° C; day/ night) CMS stamens had a near-normal morphology, but microsporogenesis proceeded to a maximum of the microspore stage. However, compared to the normal stamens, the occurrence of sporopollenin-like deposits in the tapetum and deposition of exine on the microspores was sparse. Also, the tapetal cells of the CMS line were often highly vacuolate and failed to degenerate at the same stage as the normal. Ultrastructural changes in the mitochondrial matrix and cristae plus dilation of the endoplasmic reticulum, which occurred during development in sporogenous tissues of the normal line, were often lacking or mistimed in the mutant. Due to extensive variation, even between adjacent locules, the cytological differences between the normal and CMS anthers cannot be ascribed as the cause of male sterility in the ogu CMS line of B. napus, rather they may be the consequence of it.     

Cell-free translation of mRNA encoding an arterial smooth muscle cell proteoglycan core protein

The size and immunological reactivity of the primary gene products of a small non-aggregating dermatan sulfate proteoglycan from bovine and monkey arterial smooth muscle cells were examined after cell-free translation of mRNA. Antisera against the dermatan sulfate proteoglycans from bovine articular cartilage, DSPG II [Rosenberg et al. J. Biol. Chem. 260, 6304 (1985)] and human skin fibroblasts [Glossl et al. J. Biol. Chem. 259, 14144 (1984)] were used to show that the unmodified smooth muscle precursor core protein was immunologically related to both the cartilage and fibroblast core proteins. The size of the precursor core proteins within each species was identical regardless of the tissue source. Comparison of the precursor core proteins synthesized by primate and bovine cells revealed that the bovine core proteins were approximately 1500 Da larger than the primate core proteins as determined by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. A similar size difference was observed when the mature core proteins of monkey smooth muscle cells and bovine articular chondrocytes were compared after removal of the glycosaminoglycan chains. These results indicate that arterial smooth muscle cells synthesize a dermatan sulfate proteoglycan whose core protein is similar to, if not the same as, the cartilage and fibroblast dermatan sulfate proteoglycan core proteins. These core proteins may be encoded by the same gene that has diverged in size during speciation.     

Temperature control of germination and its possible role in the survival of a non-dormant population of Avena fatua

Germination of freshly harvested seeds of a non-dormant (ND) line (Stonehouse 319) of wild oats ( Avena fatua L.) was inhibited by incubation of the seeds at relatively high temperatures of 25 and 30°C. The germination inhibition in these seeds appeared to be a case of thermo-inhibition which was the direct effect of hightemperature treatment (HIT), since it did not persist after transferring the seeds to an optimum germination temperature of 20°C. Even a prolonged HTT of 30°C for over 5 weeks did not prevent germination of about 80% of the seeds transferred to 20°C. However, in a significant proportion of the seeds, thermo-dormancy was induced by 10 days of HTT at 30°C if the seeds were then incubated at sub-optimal temperatures of 5 to 15°C. This thermo-dormancy would appear to be 'restrictive' in form, since its expression was restricted to very specific conditions. Relatively low inclubation temperaturs of 5 and 10°C markedly slowed germination whether HTT was applied or not. The results suggest that thermo-inhibition and thermo-dormancy, induced during seasonal temperature fluctuations, may provide a survival mechanism for seeds of such ND lines as Stonehouse 319.     

Temperature-induced modifications in the surface features of stamens of a tomato mutant: An SEM study

Summary Stamenless-2 (sl₂/sl₂) is a temperature-sensitive mutant of tomato (Lycopersicon esculentum) which exhibits altered stamen development under different temperatures (Sawhney 1983). By using scanning electron microscopy, this study was conducted to investigate the differentiation of surface features of mutant and normal stamens grown under different temperatures, with the view to further determine the role of temperature in gene expression in stamen development. Mutant stamens grown under intermediate temperatures (23 °C day/18 °C night) differed from the normal in hair production, the shape of epidermal cells and in the pattern of cuticular thickenings. Under low temperatures (18 °C day/15 °C night), all surface features of mutant stamens closely resembled the normal, whereas under high temperatures (28 °C day/23 °C night), the patterns and types of hairs, epidermal cells, stomata, and cuticular thickenings on mutant stamens were similar to that of a gynoecium. The staminal features of normal stamens were not affected by different temperatures. This study shows that the expression of the sl₂/sl₂ allele is influenced by temperature conditions to the extent that the pattern of cellular differentiation characteristic of either the stamens or the carpels can be induced in mutant stamens.     

Polyamines and Flower Development in the Male Sterile Stamenless-2 Mutant of Tomato (Lycopersicon esculentum Mill.) : I. Level of Polyamines and Their Biosynthesis in Normal and Mutant Flowers

The levels of free putrescine, spermidine, and spermine, and the activities of ornithine decarboxylase and s-adenosylmethionine decarboxylase were determined in the floral organs of the normal and a male sterile stamenless-2 (sl-2/sl-2) mutant of tomato (Lycopersicon esculentum Mill.). Under the intermediate temperature regime, all mutant floral organs possessed significantly higher levels of polyamines and enzyme activities than their normal counterparts. In the low temperature-reverted mutant stamens, the polyamine levels and the activity of PA biosynthetic enzymes were not significantly different from the normal. It is suggested that the abnormal stamen development in the sl-2/sl-2 mutant is, in part, related to elevated levels of endogenous PAs.     

Polyamines and Flower Development in the Male Sterile Stamenless-2 Mutant of Tomato (Lycopersicon esculentum Mill.) : II. Effects of Polyamines and Their Biosynthetic Inhibitors on the Development of Normal and Mutant Floral Buds Cultured in Vitro

The floral organs of the male sterile stamenless-2 (sl-2/sl-2) mutant of tomato (Lycopersicon esculentum Mill.) contain significantly higher level of polyamines than those of the normal (R Rastogi, VK Sawhney [1990] Plant Physiol 93: 439-445). The effects of putrescine, spermidine and spermine, and three different inhibitors of polyamine biosynthesis on the in vitro development of floral buds of the normal and sl-2/sl-2 mutant were studied. The polyamines were inhibitory to the in vitro growth and development of both the normal and mutant floral buds and they induced abnormal stamen development in normal flowers. The inhibitors of polyamine biosynthesis also inhibited the growth and development of floral organs of the two genotypes, but the normal flowers showed greater sensitivity than the mutant. The inhibitors also promoted the formation of normal-looking pollen in stamens of some mutant flowers. The effect of the inhibitors on polyamine levels was not determined. The polyamine-induced abnormal stamen development in the normal, and the inhibitor-induced production of normal-looking pollen in mutant flowers support the suggestion that the elevated polyamine levels contribute to abnormal stamen development in the sl-2/sl-2 mutant of tomato.     

Genetic research as the valid base of strategies for breeding rust resistant wheats

It is known that few wheat cultivars maintain their resistance to rust diseases for a long period of time, particularly when crop populations become genetically more uniform. A number of genetically diverse, so far unexploited, sources of rust resistance in the natural as well as mutagenized population of wheat cultivars were identified. Several of these genes were placed in agronomically superior well-adapted backgrounds so that they could be used as pre-breeding stocks for introducing genetic diversity for resistance in a crop population. Some of these stocks when employed as parents in several cross combinations in a breeding programme have generated a number of promising cultivars with diversity for resistance.Many presently grown wheats in India, near-isogenic lines each with Lr14b, Lr14ab, Lr30 and certain international cultivars were identified as possessing diverse sources of adult plant resistance (APR) to leaf rust. Prolonged leaf rust resistance in some of the Indian cultivars was attributed to the likely presence of Lr34 either alone or in combination with other APR components. Tests of allelism carried out in certain cultivars that continue to show adequate levels of field resistance confirm the presence of Lr34, which explains the role that this gene has played in imparting durability for resistance to leaf rust. Also, Lr34 in combination with other APR components increases the levels of resistance, which suggests that combination of certain APR components should be another important strategy for breeding cultivars conferring durable and adequate levels of resistance. A new adult plant leaf rust resistance source that seems to be associated with durability in Arjun has been postulated. Likewise, cultivars possessing Sr2 in combination with certain other specific genes have maintained resistance to stem rust.Further, non-specific resistances that were transferred across widely different genotypes into two of the popular Indian wheats provided easily usable materials to the national breeding programmes for imparting durable resistance to stripe rust.