Abnormal Methylation of Several Tumor Suppressor Genes in Sporadic Breast Cancer

Multiplex methylation-sensitive PCR was employed in studying the methylation of CpG islands in the R1, p16/CDKN2, p15/CDKN2, p14/ARF, DH1, MGMT, HIC1, and N33 promoter regions in breast cancer (105 tumors). Methylation was often observed for the two major suppressor genes involved in controlling the cell cycle through the Cdk–Rb–E2F signaling pathway, R1 (18/105, 17%) and p16 (59/105, 56%); both genes were methylated in 13 tumors. Methylation involved p15 in two (2%) tumors; CDH1, in 83 (79%) tumors; MGMT, in eight (8%) tumors, and N33, in nine (9%) tumors. The p14 promoter was not methylated in the tumors examined.     

Enhanced stability of alpha B-crystallin in the presence of small heat shock protein Hsp27

Lens alpha-crystallin, alpha A- and alpha B-crystallin, and Hsp27 are members of the small heat shock protein family. Both alpha A- and alpha B-crystallin are expressed in the lens and serve as structural proteins and as chaperones, but alpha B-crystallin is also expressed in nonlenticular organs where Hsp27, rather than alpha A-crystallin, is expressed along with alpha B-crystallin. It is not known what additional function Hsp27 has besides as a heat shock protein, but it may serve, as alpha A-crystallin does in the lens, to stabilize alpha B-crystallin. In this study, we investigate aspects on conformation and thermal stability for the mixture of Hsp27 and alpha B-crystallin. Size exclusion chromatography, circular dichroism (CD), and light scattering measurements indicated that Hsp27 prevented alpha B-crystallin from heat-induced structural changes and high molecular weight (HMW) aggregation. The results indicate that Hsp27 indeed promotes stability of alpha B-crystallin.     

Effects of metallic silver island films on resonance energy transfer between N,N'-(dipropyl)-tetramethyl- indocarbocyanine (Cy3)- and N,N'-(dipropyl)-tetramethyl- indodicarbocyanine (Cy5)-labeled DNA

Resonance energy transfer (RET) is typically limited to distances below 60 A, which can be too short for some biomedical assays. We examined a new method for increasing the RET distances by placing donor- and acceptor-labeled DNA oligomers between two slides coated with metallic silver particles. A N,N'-(dipropyl)-tetramethylindocarbocyanine donor and a N,N'-(dipropyl)-tetramethylindodicarbocyanine acceptor were covalently bound to opposite 5' ends of complementary 23 base pair DNA oligomers. The transfer efficiency was 25% in the absence of silver particles or if only one slide was silvered, and it increased to an average value near 64% between two silvered slides. The average value of the Forster distance increased from 58 to 77 A. The energy transfer data were analyzed with a model assuming two populations of donor-acceptor pairs: unaffected and affected by silver island films. In an affected fraction of about 28%, the apparent energy transfer efficiency is near 87% and the Forster distance increases to 119 A. These results suggest the use of metallic silver particles to increase the distances over which RET occurs in biomedical and biotechnology assays.     

Heat and water transfer in a rotating drum containing solid substrate particles

In previous work we reported on the simulation of mixing behavior of a slowly rotating drum for solid-state fermentation (SSF) using a discrete particle model. In this investigation the discrete particle model is extended with heat and moisture transfer. Heat transfer is implemented in the model via interparticle contacts and the interparticle heat transfer coefficient is determined experimentally. The model is shown to accurately predict heat transfer and resulting temperature gradients in a mixed wheat grain bed. In addition to heat transfer, the addition and subsequent distribution of water in the substrate bed is also studied. The water is added to the bed via spray nozzles to overcome desiccation of the bed during evaporative cooling. The development of moisture profiles in the bed during spraying and mixing are studied experimentally with a water-soluble fluorescent tracer. Two processes that affect the water distribution are considered in the model: the intraparticle absorption process, and the interparticle transfer of free water. It is found that optimum distribution can be achieved when the free water present at the surface of the grains is quickly distributed in the bed, for example, by fast mixing. Alternatively, a short spraying period, followed by a period of mixing without water addition, can be applied. The discrete particle model developed is used successfully to examine the influence of process operation on the moisture distribution (e.g., fill level and rotation rate). It is concluded that the extended discrete particle model can be used as a powerful predictive tool to derive operating strategies and criteria for design and scale-up for mixed SSF and other processes with granular media.     

Radiation, diversity, and host-plant interactions among island and continental legume-feeding psyllids

Island archipelagos and insect-plant associations have both independently provided many useful systems for evolutionary study. The arytainine psyllid (Sternorrhyncha: Hemiptera) radiation on broom (Fabaceae: Genisteae) in the Canary Island archipelago provides a discrete system for examining the speciation of highly host-specific phytophagous insects in an island context. Phylogenetic reconstructions based on three datasets (adult and nymph morphological characters, and two mitochondrial DNA regions: part of the small subunit rRNA, and part of cytochrome oxidase I, cytochrome oxidase II and the intervening tRNA leucine) are generally consistent. The combined molecular tree provides a well-supported estimate of psyllid relationships and shows that there have been several colonizations of the Macaronesian islands but that only one has resulted in a significant radiation. Psyllid diversification has apparently been constrained by the presence of suitable host groups within the genistoid legumes, and the diversity, distribution, and abundance of those groups. The phylogeny, by indicating pairs of sister species, allows putative mechanisms of speciation to be assessed. The most common conditions associated with psyllid speciation are geographical allopatry with a host switch to closely related hosts (six examples), or geographical allopatry on the same host (four examples). Where allopatric speciation involves a host switch, these have all been to related hosts. There is some evidence that switches between unrelated host plants may be more likely in sympatry. Only one sister pair (Aryrtainilla cytisi and A. telonicola) and the putative host races of Arytinnis modica are sympatric but on unrelated hosts, which may be a necessary condition for sympatric speciation in these insects. Where several psyllids share the same host, resources appear to be partitioned by ecological specialization and differing psyllid phenology.     

Empirical evidence for an optimal body size in snakes

Abstract The concept of optimal size has been invoked to explain patterns in body size of terrestrial mammals. However, the generality of this phenomenon has not been tested with similarly complete data from other taxonomic groups. In this study we describe three statistical patterns of body size in snakes, all of which indicate an optimal length of 1.0 m. First, a distribution of largest body lengths of 618 snake species had a single mode at 1.0 m. Second, we found a positive relationship between the size of the largest member of an island snake assemblage and island area and a negative relationship between the size of the smallest member of an island snake assemblage and island area. Best-fit lines through these data cross at a point corresponding to 1.0 m in body length, the presumed optimal size for a one-species island. Third, mainland snake species smaller than 1.0 m become larger on islands whereas those larger than 1.0 m become smaller on islands. The observation that all three analyses converge on a common body size is concordant with patterns observed in mammals and partial analyses of four other disparate animal clades. Because snakes differ so strikingly from mammals (ectotherms, gape-limited predators, elongate body shape) the concordant patterns of these two groups provide strong evidence for the evolution of an optimal body size within independent monophyletic groups. However, snakes differ from other taxonomic groups that have been studied in exhibiting a body size distribution that is not obviously skewed in either direction. We suggest that idiosyncratic features of the natural history of ectotherms allow relatively unconstrained distributions of body size whereas physiological limitations of endotherms constrain distributions of body size to a right skew.     

Phosphorylation-dependent structural alterations in the small hsp30 chaperone are associated with cellular recovery

Small heat shock proteins (hsps) act as molecular chaperones by preventing the thermal aggregation and unfolding of cellular protein; however, the manner by which cells regulate chaperone activity remains unclear. In the present study, we examined the role of phosphorylation on the chaperone function of the Xenopus small hsp30. Both heat stress and sodium arsenite treatment in A6 cells resulted in a rapid activation of p38alpha and MAPKAPK-2. Surprisingly, the association of MAPKAPK-2 with hsp30 and its subsequent phosphorylation were more prevalent during recovery after heat stress. Treatment of A6 cells with SB203580, an inhibitor of the p38 MAP kinase pathway, resulted in a loss of hsp30 phosphorylation. Phosphorylation resulted in the formation of smaller multimeric hsp30 complexes and resulted in a significant loss of secondary structure. Consequently the phosphorylation-induced structural changes severely compromised the ability of hsp30 to prevent the heat-induced aggregation of citrate synthase and luciferase in vitro. We confirmed that the loss of chaperone activity was coincident with an attenuated binding of phosphorylated hsp30 with target proteins. Our data suggest that phosphorylation may be necessary to regulate the post-heat stress molecular chaperone activity of hsp30.     

Investigation of the mechanisms affecting Cu and Fe bioavailability from legumes

Chemical composition and content in polyphenols, phytic acid, and dietary fiber of whole cooked common bean (Phaseolus vulgaris L.) and faba bean (Vicia faba L.) and of soluble and insoluble fractions separated from them were determined. Simultaneous determination of Cu, Fe, and protein bioavailability in the small intestine of rat was carried out in single-dose, short-term (1 h) experiments. After cooking, about 80% of seed components (on a weight basis) of either legume was recovered in the precipitate (insoluble fraction) after extraction with water. Protein, lipid, starch, dietary fiber, and polyphenols underwent the most severe insolubilization, together with more than 70% of total Cu and Fe. Cu, Fe, and protein bioavailability showed a similar trend (i.e., the lower the protein, the lower the Cu and Fe availability). Availability of proteins, Cu, and Fe in the insoluble fractions were the lowest, but Cu bioavailability was higher than that of Fe in all fractions. The results provide evidence that the heat-induced insolubilization process adversely affects not only protein but also Cu and Fe bioavailability from legumes and that polyphenols are likely to be a major inhibitor on absorption.     

Breaking taboos in the tropics: incest promotes colonization by wood-boring beetles

• 1 Inbreeding and parthenogenesis are especially frequent in colonizing species of plants and animals, and inbreeding in wood‐boring species in the weevil families Scolytinae and Platypodidae is especially common on small islands. In order to study the relationship between colonization success, island attributes and mating system in these beetles, we analysed the relative proportions of inbreeders and outbreeders for 45 Pacific and Old World tropical islands plus two adjacent mainland sites, and scored islands for size, distance from nearest source population, and maximum altitude.
• 2 The numbers of wood‐borer species decreased with decreasing island size, as expected; the degree of isolation and maximum island altitude had negligible effects on total species numbers.
• 3 Numbers of outbreeding species decreased more rapidly with island size than did those of inbreeders. Comparing species with similar ecology (e.g. ambrosia beetles) showed that this difference was best explained by differential success in colonization, rather than by differences in resource utilization or sampling biases. This conclusion was further supported by analyses of data from small islands, which suggested that outbreeding species have a higher degree of endemism and that inbreeding species are generally more widespread.
• 4 Recently established small populations necessarily go through a period of severe inbreeding, which should affect inbreeding species much less than outbreeding ones. In addition, non‐genetic ecological and behavioural (‘Allee’) effects are also expected to reduce the success of outbreeding colonists much more than that of inbreeders: compared with inbreeders, outbreeders are expected to have slower growth rates, have greater difficulties with mate‐location and be vulnerable to random extinction over a longer period.