Modulation of Ca2+ dependent protease activity in fish and invertebrates by weak low-frequency magnetic fields

Results of experiments addressing the effects of weak low-frequency magnetic fields on intracellular Ca²⁺ dependent proteinases (calpains) from invertebrates and fish are discussed. Exposure of live animals to weak low-frequency magnetic fields with parameters chosen to induce the resonance of Ca²⁺ ions led to a significant decrease of calpain activity in the animals investigated. The physical factor studied also caused partial loss of activity in preparations of Ca²⁺ dependent proteinases obtained from invertebrates and fish. The phenomenon discovered is in accordance with the interference model of the effect of weak low-frequency magnetic fields on biological objects.     

Mannose-6-phosphate receptors (MPR 300 and 46) from the highly evolved invertebrate <Emphasis Type="Italic">Asterias rubens</Emphasis> (Echinodermate): biochemical and functional characterization of MPR 46 protein

Mammalian mannose 6-phosphate receptors (MPR 300 and 46) mediate transport of lysosomal enzymes to lysosomes. Recent studies established that the receptors are conserved throughout vertebrates. Although we purified the mollusc receptors and identified only a lysosomal enzyme receptor protein (LERP) in the Drosophila melanogaster, little is known about their structure and functional roles in the invertebrates. In the present study, we purified the putative receptors from the highly evolved invertebrate, starfish, cloned the cDNA for the MPR 46, and expressed it in mpr^(−/−) mouse embryonic fibroblast cells. Structural comparison of starfish receptor sequences with other vertebrate receptors gave valuable information on its extensive structural homology with the vertebrate MPR 46 proteins. The expressed protein efficiently sorts lysosomal enzymes within the cells establishing a functional role for this protein. This first report on the invertebrate MPR 46 further confirms the structural and functional conservation of the receptor not only in the vertebrates but also in the invertebrates.     

Role of gangliosides in the processes of calcium ion transport across the membranes of nerve cells

It has been demonstrated that the content of extracellular Ca in the nervous system is inversely related to the content of gangliosides. The results obtained on invertebrates, lower and higher vertebrates indicate that the highest content of extracellular Ca is typical of the nervous tissue of invertebrates, whereas the lower one--of the nervous tissue of higher vertebrates (mammals). Ganglioside content, on the contrary, is the highest in the brain tissue of the higher vertebrates (mammals and birds), being significantly lower in lower vertebrates; no gangliosides was found at all in the nervous tissue of protostomian invertebrates. The highest ganglioside content in the organism of vertebrates is characteristic to the surface membranes of the nervous cells, especially in the region of synapses. Functional significance of the inverse relationship between the content of extracellular Ca and gangliosides is discussed from the standpoint of one of the authors (R. Veh) who postulated the existence of calcium--ganglioside buffer in the vicinity of the surface of the nervous cells.     

Predator impacts on stream benthic prey

The impact that predators have on benthic, macroinvertebrate prey density in streams is unclear. While some studies show a strong effect of predators on prey density, others show little or no effect. Two factors appear to influence the detection of predator impact on prey density in streams. First, many field studies have small sample sizes and thus might be unable to detect treatment effects. Second, streams contain two broad classes of predators, invertebrates and vertebrates, which might have different impacts on prey density for a variety of reasons, including availability of refuge for prey and prey emigration responses to the two types of predators. In addition, predatory vertebrates have more complex prey communities than predatory invertebrates; this complexity might reduce the impact that predatory vertebrates have on prey because of indirect effects. I conducted a meta-analysis on the results of field studies that manipulate predator density in enclosures to determine (1) if predators have a significant impact on benthic prey density in streams, (2) if the impacts that predatory invertebrates and vertebrates have differ, and (3) if predatory vertebrates have different impacts on predatory prey versus herbivorous prey. The results of the meta-analysis suggest that on average predators have a significant negative effect on prey density, predatory invertebrates have a significantly stronger impact than predatory vertebrates, and predatory vertebrates do not differ in their impact on predatory versus herbivorous invertebrate prey. Three methodological variables (mesh size of enclosures, size of enclosures, and experimental duration) were examined to determine if cross correlations exist that may explain the differences in impact between predatory invertebrates and vertebrates. No correlation exists between mesh size and predator impact. Over all predators, no correlation exists between experimental duration and predator impact; however, within predatory invertebrates a correlation does exist between these variables. Also, a correlation was found between enclosure size and predator impact. This correlation potentially explains the difference in impact between predatory invertebrates and predatory vertebrates. Results of the meta-analysis suggest two important areas for future research: (1) manipulate both types of predators within the same system, and (2) examine their impacts on the same spatial scale.     

Protochordate immunity. I. Primary immune response of the tunicate Ciona intestinalis to vertebrate erythrocytes

Serum from the tunicate Ciona intestinalis contains a low titer, natural agglutinin for a variety of vertebrate erythrocytes. Primary injections of 8.5 × 10³ human erythrocytes or 7.0 × 10² duck erythrocytes into the tunic tissues or perivisceral cavity were agglutinated and then cleared by phagocytosis within 24 hr. Higher concentrations of human or duck erythrocytes injected into the tunic tissues were encapsulated. Concomitant with clearance or encapsulation, serum agglutinin levels decreased and then returned to normal within 24 hr. Tunicates are classified taxonomically with the vertebrates in the phylum Chordata. Since phagocytosis and encapsulation reactions are typical of most invertebrates, the results of this study suggest that tunicate internal defense mechanisms are more closely allied to invertebrates than to vertebrates.     

Molecular evolution of intracellular Ca<Superscript>2+</Superscript>-dependent proteases

The structural features and evolutionary interrelationships of the intracellular Ca²⁺-dependent cysteine enzymes calpains, proteases of the family C2 (EC 3.4.22.17), are considered. A variety of identified sequences of calpains and calpain-like polypeptides found in organisms of different taxons, from the protozoa to mammals, are described. Calpains of the major evolutionary groups, typical and atypical, are classified by the analysis of their phylogenetic tree and are differentiated due to the presence of the calmodulin-like Ca²⁺-binding domain. It is shown that, along with enzymes having “advanced” characteristics (heterodimeric structure, presence of tissue-specific isoforms and splice variants, regulation by the endogenous inhibitor calpastatin, and others), higher organisms contain homologues of calpains of lower eukaryotes. A high degree of homology of the catalytic domain of calpains and the variable structure of other functional domains indicate that calpains are implicated in various physiological processes with the retention of their regulatory role.     

Enzymes of cyclic nucleotide metabolism in invertebrate and vertebrate sperm.

Sperm from several invertebrates contained guanylate cyclase activity several-hundred-fold greater than that in the most active mammalian tissues; the enzyme was totally particulate. Activity in the presence of Mn²⁺ was up to several hundred-fold greater than with Mg²⁺ and was increased 3–10-fold by Triton X-100. Sperm from several vertebrates did not contain detectable guanylate cyclase. Sperm of both invertebrates and vertebrates contained roughly equal amounts of Mn²⁺-dependent adenylate cyclase activity; in invertebrate sperm, this enzyme was generally several hundred-fold less active than guanylate cyclase. Adenylate cyclase was particulate, was unaffected by fluoride, and was generally greater than 10-fold more active with Mn²⁺ than with Mg²⁺. Invertebrate sperm contained phosphodiesterase activities against 1.0 μm cyclic GMP or cyclic AMP in amounts greater than mammalian tissues. Fish sperm, which did not contain guanylate cyclase, had high phosphodiesterase activity with cyclic AMP as substrate but hydrolyzed cyclic GMP at a barely detectable rate. In sea urchin sperm, phosphodiesterase activity against cyclic GMP was largely particulate and was strongly inhibited by 1.0% Triton X-100. In contrast, activity against cyclic AMP was largely soluble and was weakly inhibited by Triton. The cyclic GMP and cyclic AMP contents of sea urchin sperm were in the range of 0.1–1 nmol/g. Sea urchin sperm homogenates possessed protein kinase activity when histone was used as substrate; activities were more sensitive to stimulation by cyclic AMP than by cyclic GMP.⁵     

Is vitellogenin an ancestor of apolipoprotein B-100 of human low-density lipoprotein and human lipoprotein lipase?

Vitellogenin, an ancient animal protein, is the major yolk protein of eggs, where it is used as a food source during embryogenesis. Here it is shown that vitellogenins, including those from the invertebrates Caenorhabditis elegans and Drosophila melanogaster, contain domains that are homologous with parts of apolipoprotein B-100 (apoB-100) of human low-density lipoprotein and human lipoprotein lipase. As vitellogenins are likely to have been used by invertebrates during embryogenesis well before the circulation of lipids appeared in vertebrates, it is suggested that copies of a precursor gene, serving a function similar to vitellogenin, were modified to code for part of apoB-100 and lipoprotein lipase in vertebrates. In addition to providing a link between invertebrates and vertebrates for proteins involved in lipid transport, these homologies suggest new functions for vitellogenin other than being a yolk food for the developing embryo.     

The family of metazoan metal-independent {beta}-galactoside-binding lectins: structure, function and molecular evolution

Animal metal-independent ß-galactoside-binding lectinswere initially found in vertebrates, but they have recentlybeen isolated from much lower invertebrates, such as nematodeand sponge, as well. Further, an eosinophilic lysophospholipaseassociated with various inflammatory reactions was very recentlyfound to be a new member of this protein family. It appearsthat ß-galactoside-binding lectins and some non-lectinproteins form a superfamily whose members are widely distributedfrom vertebrates to invertebrates. From the viewpoints of proteinarchitecture, the superfamily members can be subdivided intothree types; i.e. ‘proto type’ (the relatively well-studied14 kDa lectins), ‘chimera type’ (29–35 kDalectins also known as     

环境污染对几类水生无脊椎动物内分泌功能扰乱的研究现状

近年来，在环境毒理学这门边缘学科中又诞生了一个新的领域，即环境污染对内分泌功能的扰乱。研究发现，许多人工合成的杀虫剂和工业化合物能够扰乱脊椎动物的内分泌功能，这些化合物也存在于水环境中。近年来，这些环境有机污染物是否对水生无脊椎动物的内分泌功能同样具有扰乱作用成了环境内分泌学这个新领域的热点之一。由于近年来的研究侧重于腔肠动物、轮虫、软体动物、甲壳动物及棘皮动物，因此，本文主要介绍有关环境污染物对这几类水生无脊椎动物内分泌功能扰乱的研究进展。另外，对环境污染对水生无脊椎动物内分泌扰乱这个研究热点的现状以及今后的发展方向进行了评述。在从事环境污染对无脊椎动物内分泌功能影响的研究时，研究者必须意识到无脊椎动物和脊椎动物在内分泌机制上的差异，不可随意地在这两大类动物类群之间互相引伸研究结果。     

Adrenergic receptor homologies in vertebrate and invertebrate species examined by DNA hybridization

The deduced protein sequences of the mammalian adrenergic receptors (ARs) suggest that these proteins have evolved by several ancient gene duplication events. To investigate in what species these events may have occurred DNA fragments encoding the family of adrenergic receptors from human (beta 1AR and alpha 2AR) and hamster (beta 2AR and alpha 1AR) were used to detect homologous sequences in other vertebrates, invertebrates and unicellular organisms by Southern blot hybridization analysis. Sequences homologous to hamster beta 2AR were detected in lower vertebrates, invertebrates and Dictyostelium, but not in yeast or bacteria. Within vertebrates, sequences strongly homologous to human beta 1AR and human platelet alpha 2AR were confined to the higher vertebrates only. In the invertebrates, only Drosophila contained sequences homologous to hamster alpha 1AR. Our results suggest that non-mammalian species may contain receptors homologous to the mammalian adrenergic receptors and that the sequences homologous to human beta 2AR have been the most strongly conserved.     

Life is a huge compromise: Is the complexity of the vertebrate immune-neuroendocrine system an advantage or the price to pay?

Recent advances in comparative immunology have established that invertebrates produce hypervariable molecules probably related to immunity, suggesting the possibility of raising a specific immune response. “Priming” and “tailoring” are terms now often associated with the invertebrate innate immunity. Comparative immunologists contributed to eliminate the idea of a static immune system in invertebrates, making necessary to re-consider the evolutive meaning of immunological memory of vertebrates. If the anticipatory immune system represents a maximally efficient immune system, why can it be observed only in vertebrates, especially in consideration that molecular hypervariability exists also in invertebrates? Using well-established theories concerning the evolution of the vertebrate immunity as theoretical basis we analyze from an Eco-immunology-based perspective why a memory-based immune system may have represented an evolutive advantage for jawed vertebrates. We hypothesize that for cold-blooded vertebrates memory represents a complimentary component that flanks the robust and fundamental innate immunity. Conversely, immunological memory has become indispensable and fully exploited in warm-blooded vertebrates, due to their stable inner environment and high metabolic rate, respectively.     

Comparative sensitivity of cholinesterases in vertebrates and invertebrates to highly specific organophosphorus inhibitors,diisopropyl fluorophosphate (DFP) and (2-ethoxymethyl phosphoryl thioethyl) ethyl (methyl) sulphonium sulphomethylate (GD-42)

The review presents the data on a comparative reactivity of 68 cholinesterase preparations from various organs and tissues in a number of vertebrates and invertebrates based on the sensitivity to the two highly specific and most studied organophosphorus inhibitors, diisopropyl fluorophosphates (DFP) and (2-ethoxymethyl phosphoryl thioethyl) ethyl (methyl) sulphonium sulphomethylate (GD-42). An analysis of the data obtained suggests a great diversity of enzymologic characteristics of cholinesterase preparations in vertebrates and invertebrates observed even in closely related enzymes in animals at virtually the same level of evolutionary development.     

Calpain 8/nCL-2 and Calpain 9/nCL-4 Constitute an Active Protease Complex,G-Calpain,Involved in Gastric Mucosal Defense

Calpains constitute a superfamily of Ca²⁺-dependent cysteine proteases, indispensable for various cellular processes. Among the 15 mammalian calpains, calpain 8/nCL-2 and calpain 9/nCL-4 are predominantly expressed in the gastrointestinal tract and are restricted to the gastric surface mucus (pit) cells in the stomach. Possible functions reported for calpain 8 are in vesicle trafficking between ER and Golgi, and calpain 9 are implicated in suppressing tumorigenesis. These highlight that calpains 8 and 9 are regulated differently from each other and from conventional calpains and, thus, have potentially important, specific functions in the gastrointestinal tract. However, there is no direct evidence implicating calpain 8 or 9 in human disease, and their properties and physiological functions are currently unknown. To address their physiological roles, we analyzed mice with mutations in the genes for these calpains, Capn8 and Capn9. Capn8^−/− and Capn9^−/− mice were fertile, and their gastric mucosae appeared normal. However, both mice were susceptible to gastric mucosal injury induced by ethanol administration. Moreover, the Capn8^−/− stomach showed significant decreases in both calpains 9 and 8, and the same was true for Capn9^−/−. Consistent with this finding, in the wild-type stomach, calpains 8 and 9 formed a complex we termed “G-calpain,” in which both were essential for activity. This is the first example of a “hybrid” calpain complex. To address the physiological relevance of the calpain 8 proteolytic activity, we generated calpain 8:C105S “knock-in” (Capn8^CS/CS) mice, which expressed a proteolytically inactive, but structurally intact, calpain 8. Although, unlike the Capn8^−/− stomach, that of the Capn8^CS/CS mice expressed a stable and active calpain 9, the mice were susceptible to ethanol-induced gastric injury. These results provide the first evidence that both of the gastrointestinal-tract-specific calpains are essential for gastric mucosal defense, and they point to G-calpain as a potential target for gastropathies caused by external stresses.     

Role of calpains in diabetes mellitus-induced cataractogenesis: A mini review

Premature visual impairment due to lens opacification is a debilitating characteristic of untreated diabetes. Lens opacification is primarily due to the insolubilization of crystallins, proteins essential for lens optical properties, and recent studies have suggested that a major cause of this insolubilization may be the unregulated proteolysis of crystallins by calpains. These are intracellular cysteine proteases whose activation requires the presence of calcium (Ca²⁺) and elevated levels of lens Ca²⁺ is a condition associated with both diabetic cataractogenesis and other forms of the disorder. A number of calpains have been identified in the lens, including calpain 2, calpain 10 and two isozymes of calpain 3:Lp82 and Lp85. The use of animal hereditary cataract models have suggested that calpain 2 and/or Lp82 may be the major calpains involved in murine cataractogenesis with contributions from calpain 10 and Lp85. However, calpain 2 appears to be the major calpain involved in murine diabetic cataractogenesis and the strongest candidate of the calpains for a role in human types of cataractogenesis. Here, we present an overview of recent evidence on which these observations are based with an emphasis on the ability of calpains to proteolyse lens crystallins and calpain structural features, which appear to be involved in the Ca²⁺-mediated activation of these enzymes. (Mol Cell Biochem 261: 151–159, 2004)     

Collected and self‐secreted building materials and their contributions to compression and tension structures

Animals that build structures have only two ways of obtaining their building materials: collecting them from the environment or secreting them. We identify a clear pattern among invertebrates: collected materials dominate in structures that bear loads predominantly in compression, whereas self‐secreted materials dominate in structures that bear loads in tension. Among vertebrates, by contrast, structures in tension are much less common and may make use of either self‐secreted or collected materials. Two possible explanations are examined for the difference between invertebrates and vertebrates. It may be that there has been evolutionary constraint on bodily secretions of vertebrates. Alternatively, the cost‐effectiveness of self‐secreted building materials diminishes with the load they bear in tension, and most vertebrates, because of their characteristically greater weight compared with invertebrates, exceed the viability threshold. Invertebrates have adapted biochemically varied secretions from a variety of glands to perform specialized mechanical roles. However, vertebrates have secretions with the potential to become building materials, in particular keratin, but only mucus‐like secretions have evolved as building materials. The cost of suspended structures is predicted to become relatively greater with increased weight, whilst it will also become more difficult to find supports rigid enough to resist the forces they impose. This conforms to the relatively small size and small number of vertebrate structures in tension. The much larger, heavier suspended structures built by some social insects demonstrate that ecological factors may have important effects on the determination of weight thresholds. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112 , 625–639.     

Presynaptic modulation of sensory afferents in the invertebrate and vertebrate nervous system

• 1.1. Ultrastructural examination of the central terminals of sensory afferent neurons in both invertebrates and vertebrates demonstrates that the synapses that form the substrate for presynaptic inhibition and facilitation are almost universally present.
• 2.2. Presynaptic modulation of afferent input acts in many ways which tailor the inflow of sensory information to the behaviour of the animal, effectively providing a means of turning this on and off, or of combining information of the same or different modalities to refine responsiveness or clarify ambiguity.
• 3.3. Presynaptic modulation may act in several different roles on the same afferent.
• 4.4. A comparison of the mechanisms of presynaptic inhibition in different animals demonstrates the likelihood of a variety of common mechanisms,several of which may act simultaneously on the same terminal.These include changes in the conductance of the afferent membrane to Cl^-, K⁺and Ca²⁺ions, in addition to less well understood mechanisms that directly affect transmitter release.
• 5.5.A single transmitter can produce several effects on a terminal through the same or different receptors.
• 6.6. Ultrastructural studies of afferent terminals reveal that only a proportion of boutons on a given afferent may receive presynaptic input and that this may depend on the region of the nervous system in which these are found or on the identity of the postsynaptic neurons contacted.
• 7.7. The synaptic relationships of afferent terminals can be complex. In invertebrates different types of presynaptic neuron may interact synaptically,as may postsynaptic dendrites in vertebrates.
• 8.8. Axons presynaptic to afferent terminals in vertebrates frequently synapse also with dendrites postsynaptic to the afferents.
• 9.9. In both invertebrates and vertebrates reciprocal interactions between afferents and postsynaptic neurons are seen.
• 10.10. Ultrastructural immunocytochemistry reveals the likely dominance of GABA as an agent of presynaptic inhibition but also demonstrates the possible presence of other transmitters some of whose roles are less completely understood.

     

Molecular evolution of the NF-κB signaling system

The mechanisms of innate immunity in vertebrates show certain overall resemblances to immune mechanisms of insects. Two hypotheses have been proposed to explain these resemblances. (1) According to the evolutionary continuity hypothesis, innate immune mechanisms evolved in the common ancestor of vertebrates and insects and have been conserved since that time. (2) In the independent-evolution hypothesis, the mechanisms of innate immunity in vertebrates evolved independently from invertebrate immune mechanisms. Phylogenetic analysis of five gene families (Pelle, Rel, IkappaB, Toll, and TRAF) whose members are involved in NF-kappaB signaling in vertebrates and insects were used to decide between these hypotheses. The phylogenies of the Rel and TRAF families strongly supported independent evolution of immune functions in vertebrates and invertebrates, and, except for a possible case in the Pelle family, orthologous molecules having immune functions in both vertebrates and invertebrates were not found. The results suggest that NF-kappaB represents an ancient, generalized signaling system that has been co-opted for immune system roles independently in vertebrate and insect lineages.     

Insect herbivory and vertebrate grazing impact food limitation and grasshopper populations during a severe outbreak

1. Competition between herbivores often plays an important role in population ecology and appears strongest when densities are high or plant production is low. Phytophagous insects are often highly abundant, but relatively few experiments have examined competition between vertebrates and phytophagous insects. 2. In grassland systems worldwide, grasshoppers are often the dominant phytophagous insect, and livestock grazing is a dominant land use. For this study, a novel experiment was conducted examining competition between vertebrates and invertebrates, where both grasshopper densities and sheep grazing were manipulated inside 10‐m² caged mesocosms during a grasshopper outbreak. We examined how grasshopper densities and the timing of vertebrate herbivory affected grasshopper densities, if the effects of vertebrates on survival and reproduction changed with grasshopper density, and how a naturally occurring grasshopper outbreak affected grasshopper populations in the following year. 3. Densities of grasshoppers at the site peaked at 130 m^–2. Food‐limited competition was stronger in treatments with higher grasshopper densities and repeated or late livestock herbivory, leading to reduced survival, femur length, and functional ovarioles, a measure of future reproduction. Strong food‐limited density‐dependent reproduction and survival led to reduced hatching densities in 2001. 4. As competition was typically stronger with high grasshopper densities than with livestock grazing, competition from vertebrates could be relatively less important for phytophagous insect population dynamics during outbreaks. The experiment provides insights into how competition between insect and vertebrate herbivores influences insect population dynamics, and indicates that severe outbreaks can rapidly subside with strong competition from vertebrate and insect herbivores.     

基因倍增和脊椎动物起源

有机体基因复制导致基因复杂性增加及其和脊椎动物起源的关系已经成为进化生物学研究的热点。20世纪70年代由Ohno提出后经Holland等修正的原始脊索动物经两轮基因组复制产生脊椎动物的假设目前已被广泛接受。脊椎动物起源和进化过程中发生过两轮基因组复制的主要证据有三点：（1）据估计脊椎动物基因组内编码基因数目大约相当于果蝇、海鞘等无脊椎动物的4倍;原口动物如果蝇和后口动物如头索动物文昌鱼的基因组大都只有单拷贝的基因,而脊椎动物的基因组则通常有4个同属于一个家族的基因。（2）无脊椎动物如节肢动物、海胆和头索动物文昌鱼都只有一个Hox基因簇,而脊椎动物除鱼类外,有7个具有Hox基因簇,其余都具有4个Hox基因簇。（3）基因作图证明,不但在鱼类和哺乳动物染色体广大片段上基因顺序相似,而且有证据显示哺乳动物基因组不同染色体之间存在相似性。据认为第一次基因倍增发生在脊椎动物与头索动物分开之后,第二次基因倍增发生在有颌类脊椎动物和无颌类脊椎动物分开以后。但是,基因是逐个发生倍增,还是通过基因组内某些DNA片段抑或整个基因组的加倍而实现的,目前还颇有争议。