Effect of Temperature on the Formation of Microsclerotia of Verticillium dahliae

Microsclerotium formation by six isolates of Verticillium dahliae was studied at different temperatures both in vitro and in Arabidopsis thaliana . In vitro mycelial growth was optimal at 25°C, but microsclerotium formation was greatest at 20°C (two isolates) or 15–20°C (one isolate). Seedlings of A. thaliana were root-dipped in a conidial suspension, planted, and either placed at 5, 10, 15, or 25°C, or left at 20°C until the onset of senescence, after which some of the plants were placed at 5, 10, 15, or 25°C. The amount of microsclerotia per unit of shoot weight was assessed in relation to isolate and temperature. The optimal temperature for production of microsclerotia was 15–25°C. Two isolates each produced about 10 times more microsclerotia than each of the other four isolates. For these isolates, high R ²_adj.-values of 0.77 and 0.66 were obtained, with temperature and its square as highly significant (P   < 0.001) independent variables. R ²_adj.-values for the other isolates varied between 0.28 and 0.39. Moving plants to different temperatures at the onset of senescence led to microsclerotial densities that were intermediate between densities on plants that had grown at constantly 20°C and plants grown at other temperatures. This suggests that vascular colonization rate and rate of microsclerotium formation are similarly affected by temperature. The senescence rate of plants appeared unimportant except for plants grown at 25°C, which showed the highest amounts of microsclerotia per unit of plant weight in the most rapidly senescing plants.     

Latency of Botrytis cinerea Pers.: Fr. and Biochemical Studies During Growth and Ripening of Two Grape Berry Cultivars, Respectively Susceptible and Resistant to Grey Mould

Latency and development of Botrytis cinerea were assessed under field conditions and after artificial inoculation of two grape varieties, Gamay (susceptible) and Gamaret (resistant). When the percentage of latent Botrytis was the same for both varieties, severity of visible grey mould remained very low in Gamaret berries, while Gamay clusters were destroyed by the disease to a high percentage. Some biochemical parameters were measured in berries, such as constitutive and induced anti‐fungal compounds, polymeric proanthocyanidins and lipid peroxidation products as markers of senescence. Differences were observed in polymeric proanthocyanidins (PPRA) of Gamaret compared with those of Gamay. Concentration and mean degree of polymerization (mDP) of PPRA were always higher in the berries of the resistant variety. The inhibitory effect of Gamaret PPRA on enzyme activity remained until harvest whereas Gamay PPRA lost their inhibitory activity at the beginning of véraison. Based on these results, resistance to B. cinerea seems to be linked to the maintainance of the fungus in its latent form in berries, mainly due to the ability of Gamaret PPRA to inhibit macerating fungal enzyme activities.     

A free-radical hypothesis for the instability and evolution of genotype and phenotypein vitro

It has been known for several decades that cultured murine cells undergo a defined series of changes, i.e., anin vitro evolution, which includes crisis, spontaneous transformation (immortalization), aneuploidy, and spontaneous neoplastic transformation. These changes have been shown to be caused by thein vitro environment rather than an inherent instability of the murine phenotype or genotype. Serum amine oxidases were recently identified as a predominant cause of crisis. These enzymes generate hydrogen peroxide from polyamine substrates that enter the extracellular milieu. This finding implicates free-radical toxicity as the underlying cause ofin vitro evolution. We propose an oxyradical hypothesis to explain each of the stages ofin vitro evolution and discuss its significance for cytotechnology and long-term cultivation of mammalian cell types.ORR, CDER, FDA Mod-1, Room 2023, 8301 Muirkirk Road, Laurel MD 20708, USA     

Change in floral nectar components from fresh to senescent flowers ofCapparis spinosa (Capparidaceae), a nocturnally flowering Mediterranean shrub

We studied the nectar characteristics in relation to flower age of the summer flowering Mediterranean shrubCapparis spinosa in three localities in Southern Greece. Anthesis was nocturnal. Nectar volume, concentration, and sucrose/hexose ratio varied with site, year, and between individual plants; amino acid concentration varied only with site. The sucrose/hexose ratio decreased considerably with flower age, while the glucose/fructose ratio remained constant (ca. 1), implying that nectar sucrose broke down in the course of anthesis. Sugar breakdown increased with water content of nectar. Amino acid concentration was strongly age-dependent: It was low in fresh flowers, relatively high in middle-aged ones (except aspartic acid that was extremely increased), and very high in senescent ones. We attribute the amino acid changes to phenomena related to flower senescence in the dark.     

Evolution of aging: Testing the theory usingDrosophila

Evolutionary explanations of aging (or senescence) fall into two classes. First, organisms might have evolved the optimal life history, in which survival and fertility late in life are sacrificed for the sake of early reproduction or high pre-adult survival. Second, the life history might be depressed below this optimal compromise by the influx of deleterious mutations; since selection against late-acting mutations is weaker, deleterious mutations will impose a greater load on late life. We discuss ways in which these theories might be investigated and distinguished, with reference to experimental work withDrosophila. While genetic correlations between life history traits determine the immediate response to selection, they are hard to measure, and may not reflect the fundamental constraints on life history. Long term selection experiments are more likely to be informative. The third approach of using experimental manipulations suffers from some of the same problems as measures of genetic correlations; however, these two approaches may be fruitful when used together. The experimental results so far suggest that aging inDrosophila has evolved in part as a consequence of selection for an optimal life history, and in part as a result of accumulation of predominantly late-acting deleterious mutations. Quantification of these effects presents a major challenge for the future.     

SEASONAL PATTERNS OF MORPHOLOGICAL VARIABILITY IN SARGASSUM POLYCERATIUM (PHAEOPHYTA)1

Temporal variability in certain morphological and taxonomically important features was quantified for Sargassum polyceratium Mont. from a population in the Content Keys, Florida (U.S.A.). Patterns of blade development, senescence, and loss caused pronounced seasonal changes in blade length-width ratios. Blade length and width were maximal early in the growing season (August-November) and decreased as the annual stems matured. Early in the growing season, plants had broader blades with randomly distributed cryptostomata. Late in the growing season (February-April), plants had more linear blades with cryptostomata approximately arranged in two rows, one on each side of the midrib. The length-width ratio of blades increased acropetally along the stems and were directly correlated to the size of the cryptostomatal opening and inversely correlated with the number of cryptostomata. The branching pattern of the annual stems ranged from short spur branches to well-developed, lateral axillary branches. The frequency of bifurcated blades increased significantly late in the growing season. Vesicle shape and size and pedicel length were temporally stable. Alated pedicels and mucronate vesicles occurred in low frequencies. The variability of the morphological features used to delineate species within the genus Sargassum on the tropical eastern coasts of the Americas is poorly understood.     

Cell size in aging monolayer cultures

Summary Changes in the size of the area covered by individual cultured WI-38 cells as the cultures age have been studied by using a new microphotographic paper cutout technique. This method is nondestructive and nonintrusive and avoids a number of artifacts which can occur in the measurement of suspended cells. The measurements reveal that the decreased cell yield of late passage cultures-reflects not only the appearance of a subpopulation of larger cells but also the failure of the cells to utilize all the growth surface available to them. This work was supported in part by USPHS research grant AG-00378 and by a fellowship, AG-05019, from the National Institute on Aging.     

ZMAT3 hypomethylation contributes to early senescence of preadipocytes from healthy first‐degree relatives of type 2 diabetics

Senescence of adipose precursor cells (APC) impairs adipogenesis, contributes to the age‐related subcutaneous adipose tissue (SAT) dysfunction, and increases risk of type 2 diabetes (T2D). First‐degree relatives of T2D individuals (FDR) feature restricted adipogenesis, reflecting the detrimental effects of APC senescence earlier in life and rendering FDR more vulnerable to T2D. Epigenetics may contribute to these abnormalities but the underlying mechanisms remain unclear. In previous methylome comparison in APC from FDR and individuals with no diabetes familiarity (CTRL), ZMAT3 emerged as one of the top‐ranked senescence‐related genes featuring hypomethylation in FDR and associated with T2D risk. Here, we investigated whether and how DNA methylation changes at ZMAT3 promote early APC senescence. APC from FDR individuals revealed increases in multiple senescence markers compared to CTRL. Senescence in these cells was accompanied by ZMAT3 hypomethylation, which caused ZMAT3 upregulation. Demethylation at this gene in CTRL APC led to increased ZMAT3 expression and premature senescence, which were reverted by ZMAT3 siRNA. Furthermore, ZMAT3 overexpression in APC determined senescence and activation of the p53/p21 pathway, as observed in FDR APC. Adipogenesis was also inhibited in ZMAT3‐overexpressing APC. In FDR APC, rescue of ZMAT3 methylation through senolytic exposure simultaneously downregulated ZMAT3 expression and improved adipogenesis. Interestingly, in human SAT, aging and T2D were associated with significantly increased expression of both ZMAT3 and the P53 senescence marker. Thus, DNA hypomethylation causes ZMAT3 upregulation in FDR APC accompanied by acquisition of the senescence phenotype and impaired adipogenesis, which may contribute to FDR predisposition for T2D.