Nonconventional trafficking of Ras associated with Ras signal organization

Ras signaling to its downstream effectors appears to include combinations of extracellular-signal-regulated Ras activation at the plasma membrane (PM) and endomembranes, dynamic lateral segregation in the PM, and translocation of Ras from the PM to intracellular compartments. These processes are governed by the C-terminal polybasic farnesyl domain in K-Ras 4B and by the cysteine-palmitoylated C-terminal farnesyl domains in H-Ras and N-Ras. K-Ras 4B has no palmitoylated cysteines. Depalmitoylation/repalmitoylation of H-/N-Ras proteins promotes their cellular redistribution and signaling by mechanisms as yet unknown, possibly involving chaperones. Palmitoylation of H-/N-Ras also promotes their association with 'rasosomes', randomly diffusing nanoparticles that apparently provide a means by which multiple copies of activated Ras and its signal can spread rapidly. Ubiquitination of H-Ras evidently targets it to the endosomes. The polybasic farnesyl domain of K-Ras 4B was shown to act as a target for Ca++/calmodulin, which sequesters the active protein from the PM, thereby facilitating its trafficking to Golgi apparatus and early endosomes. Protein kinase C-dependent phosphorylation of S181 in K-Ras 4B was shown to provide a regulated farnesyl-electrostatic switch on K-Ras 4B, which promotes its translocation to the mitochondria. All these translocation events are characterized by nonconventional trafficking of the farnesyl-modified Ras proteins and seem to govern the selectivity and probably also the robustness of the Ras signal. In this review, we discuss the various modifications and interactions of the farnesylated C-terminus, the trafficking of Ras proteins in the PM and between the PM and the endomembranes, and the relevance of the subcellular localization of Ras for Ras function.     

Natural history,physiology and energetic strategies of Asellia tridens (Chiroptera)

We used radio-telemetry, observations and physiological measurements to study the basic biology and energetic strategies of Asellia tridens in northern Israel from 2009 to 2010. Between late May and early November, the bats occupied abandoned man-made structures in this area. Parturition occurred between late June and mid-July, and juveniles were independent by late August. A. tridens foraged near the roost in a vegetation-rich, cluttered background environment, catching insects flying close to vegetation. Its diet was diverse, with Coleoptera, Heteroptera, Diptera and Lepidoptera being the main diet components. During summer, males and females differed in their foraging patterns and energetic strategies: Lactating females departed for more frequent foraging bouts than males, and maintained euthermy throughout the day, while males became torpid on a daily basis.     

Mutation Rules and the Evolution of Sparseness and Modularity in Biological Systems

Biological systems exhibit two structural features on many levels of organization: sparseness, in which only a small fraction of possible interactions between components actually occur; and modularity – the near decomposability of the system into modules with distinct functionality. Recent work suggests that modularity can evolve in a variety of circumstances, including goals that vary in time such that they share the same subgoals (modularly varying goals), or when connections are costly. Here, we studied the origin of modularity and sparseness focusing on the nature of the mutation process, rather than on connection cost or variations in the goal. We use simulations of evolution with different mutation rules. We found that commonly used sum-rule mutations, in which interactions are mutated by adding random numbers, do not lead to modularity or sparseness except for in special situations. In contrast, product-rule mutations in which interactions are mutated by multiplying by random numbers – a better model for the effects of biological mutations – led to sparseness naturally. When the goals of evolution are modular, in the sense that specific groups of inputs affect specific groups of outputs, product-rule mutations also lead to modular structure; sum-rule mutations do not. Product-rule mutations generate sparseness and modularity because they tend to reduce interactions, and to keep small interaction terms small.     

Loss of net negative surface charge during MLC stimulation of human T lymphocytes: correlation to "stable" E-rosette formation and natural attachment to normal and malignant target cells.

Activation of human blood T lymphocytes in mixed lymphocyte culture (MLC) causes a reduction in the net negative surface charge, as indicated by the reduction in the electrophoretic mobility. Concomitantly, the activated cells acquire new properties, including the ability to form “stable” E rosettes, and attach to normal and malignant cells of the same species (natural attachment (NA)). These properties were found to be expressed by lymphocytes within the low electrophoretic fractions (cells with low negative charge) of the MLC populations. The formation of stable E rosettes and natural attachment capacities of human thymocytes were also found to correlate with the amount of surface negative charge. The slowly migrating (less negative charged) cortical thymocytes, reported earlier as being able to form stable E rosettes, were found also to exhibit NA activity. Medullary thymocytes carrying a high net negative surface charge lack these characteristics. We consider it likely that the reduction of negative charge during activation of peripheral T cells, facilitates cell-to-cell contacts, and thus prepares the (activated) cells to perform cooperative interactions with other cell types, and express the lytic activity of T cells.     

Representation of nonepistatic selection models and analysis of multilocus Hardy-Weinberg equilibrium configurations

Summary The paper develops conditions for the existence and the stability of central equilibria emanating from selection recombination interaction with generalized nonepistatic selection forms operating in multilocus multiallele systems. The selection structure admits a natural representation as simple sums of Kronecker products based on a common set of marginal selection components. A flexible parametrization of the recombination process is introduced leading to a canonical derivation of the transformation equations connecting gamete frequency states over successive generations. Conditions for the existence and stability of multilocus Hardy-Weinberg (H.W.) type equilibria are elaborated for the classical nonepistatic models (multiplicative and additive viability effects across loci) as well as for generalized nonepistatic selection expressions. It is established that the range of recombination distributions maintaining a stable H.W. polymorphic equilibrium is confined to loose linkage in the pure multiplicative case, but is not restricted in the additive model. In the bisexual case we ascertain for the generalized nonepistatic model the stability conditions of a common H.W. polymorphism.This paper was supported in part by NIH Grant GM 10452-14 and NSF Grant MCS 75-23608.     

Generation of a fluorescently labeled endogenous protein library in living human cells

We present a protocol to tag proteins expressed from their endogenous chromosomal locations in individual mammalian cells using central dogma tagging. The protocol can be used to build libraries of cell clones, each expressing one endogenous protein tagged with a fluorophore such as the yellow fluorescent protein. Each round of library generation produces 100-200 cell clones and takes about 1 month. The protocol integrates procedures for high-throughput single-cell cloning using flow cytometry, high-throughput cDNA generation and 3' rapid amplification of cDNA ends, semi-automatic protein localization screening using fluorescent microscopy and freezing cells in 96-well format.     

Characterization of a plasma membrane H+-ATPase from the extremely acidophilic algaDunaliella acidophila

Summary Dunaliella acidophila is an unicellular green alga which grows optimally at pH 0–1 while maintaining neutral internal pH. A plasma membrane preparation of this algae has been purified on sucrose density gradients. The preparation exhibits vanadatesensitive ATPase activity of 2 mol P_i/mg protein/min, an activity 15 to 30-fold higher than that in the related neutrophilic speciesD. salina. The following properties suggest that the ATPase is an electrogenic plasma membrane H⁺ pump. (i) ATP induces proton uptake and generates a positive-inside membrane potential as demonstrated with optical probes. (ii) ATP hydrolysis and proton uptake are inhibited by vanadate, diethylstilbestrol, dicyclohexylcarbodiimide and erythrosine but not by molybdate, azide or nitrate. (iii) ATP hydrolysis and proton uptake are stimulated by fussicoccin in a pH-dependent manner as found for plants plasma membrane H⁺-ATPase. Unusual properties of this enzyme are: (i) theK _m for ATP is around 60 M, considerably lower than in other plasma membrane H⁺-ATPases, and (ii) the ATPase activity and proton uptake are stimulated three to fourfold by K⁺ and to a smaller extent by other monovalent cations. These results suggest thatD. acidophila possesses a vanadate-sensitive H⁺-ATPase with unusual features enabling it to maintain the large transmembrane pH gradient.     

Investigating a lotic microbial community following a severe detergent spill

A large non-ionic detergent spill affected the Yarqon stream, where water sampling was performed prior to the spill as a part of the stream’s routine sampling and during and after the event. Following the spill, a large foam layer was observed for about 3–4 days accompanied by death of all fauna in the stream. Despite a large quantity of freshwater that was introduced to the stream as an emergency measure, a drastic decrease in dissolved oxygen was also observed. A rapid reduction in bacterial diversity and richness, as measured by automated ribosomal intergenic spacer analysis, was also evident, as microbial assemblages changes accompanied pollutant exposure. However, this analysis showed that the microbial assemblages of the stream were quick to recover and became similar to pre-spill communities as early as a week after the spill. These findings suggest that bacterial assemblages are much more robust to large anthropogenic disturbances than expected.