Proximate sources of marine biodiversity

When temperature and other kinds of barrier divide formerly continuous populations and confine them to more restricted geographical areas, there is an evolutionary reaction that will, over time, result in the formation of endemic species. In such cases, an allopatric speciation process is considered to have taken place because reproductive isolation was caused by physical means instead of by natural selection. In contrast, when populations exist in a very high-diversity area and remain undivided by physical events, they exhibit a tendency to speciate by means of sympatry (or parapatry). This process, sometimes called competitive or ecological speciation, does involve reproductive isolation by means of natural selection. Populations that exist in geographical provinces bounded by physical barriers add to the overall diversity through the production of endemic species. This increase by species packing is relatively slow due to the very gradual tempo of the allopatric speciation process. Populations existing in centres of origin add to the general diversity through the production of species that are dominant in terms of their ability to spread over large parts of the world. It is proposed that such species are usually formed by sympatric speciation, a process that can be c. 20 times faster than species formation by allopatry. It is not suggested that sympatry is exclusive to centres of origin, nor that allopatry is confined to peripheral provinces. Both processes are widespread, but there do appear to be distinctive geographical concentrations. Considering that numbers of widespread species produced by centres of origin may eventually become subdivided by barriers, and thus give rise to descendants by allopatry, it is difficult to say how much of our present species diversity has come from one source or the other. Both speciation by sympatry from centres of origin and speciation by allopatry in peripheral provinces appear to be important sources of marine biodiversity.     

Resistin release by human adipose tissue explants in primary culture

Resistin, also known as Fizz3 or ADSF, is a protein found in murine adipose tissue and inflammatory lung exudates. The present studies found that resistin was released by explants of human adipose tissue but the release was quite variable ranging from 3 to 158 ng/g over 48 h. The release of resistin was 250% greater by explants of omental than by explants of human subcutaneous abdominal adipose tissue. Resistin release by adipocytes was negligible as compared to that by the non-fat cells of adipose tissue. Leptin formation by adipocytes was 8-fold greater than its formation by the non-fat cells, while the formation of PAI-1 by adipocytes was 38% of that by the non-fat cells. The conversion of glucose to lactate as well as the formation of PGE(2) and IL-8 was approximately 15% of that by the non-fat cells. In contrast the release of IL-6 and IL-1beta by adipocytes was 4-7% of that by the non-fat cells while the formation of resistin and IL-10 by adipocytes was 2% of that by non-fat cells. The release of adiponectin by explants ranged from 1000 to 5000 ng/g over 48 h but did not correlate with that of resistin. The present data suggest that resistin release by explants of human adipose tissue in primary culture is largely derived from the non-fat cells present in the explants.     

Reconsidering paternalism in clinical research

The ethical standards that regulate clinical research have multiple rationales. Among them is the need to protect potential subjects from making imprudent decisions, which extends beyond the soft paternalistic concern to protect people from making uninformed decisions to participate in trials. This article argues that a plausible risk/benefit restriction on clinical trials is presumptively justified by hard paternalism, which in turn is supported by a deeper fairness‐based rationale. This presumptive case for hard paternalism in research is not defeated by the alleged right to participate in clinical trials, by concerns about insult or status, by the need to conduct early phase trials that promise little to no benefit to participants, or by the recognition that some potential subjects are altruistically motivated.     

P-glycoprotein-overexpressing multidrug-resistant cells are resistant to infection by enveloped viruses that enter via the plasma membrane.

The multidrug resistance gene product P-glycoprotein confers drug resistance to tumor cells by acting as a transporter that blocks the entry into the cell of a great variety of drugs and hydrophobic peptides. In this study we find that in drug-resistant cells, the insertion of the influenza virus fusion protein (hemagglutinin-2) into the plasma membrane is blocked and that the fusion of the viral envelope with the plasma membrane of these cells is impaired. Multidrug-resistant cells display significant resistance to infection by envelope viruses that invade cells by fusion with the plasma membrane, but not to infection by pH-dependent viruses that penetrate cells by fusion with endocytic vesicles. These observations suggest that multidrug resistance phenomena may protect cells from infection by a large group of disease-causing viruses that includes human immunodeficiency virus, herpes simplex virus, and some cancer-inducing retroviruses.     

The mechanism of skeletal muscle myosin ATPase. Interaction of myosin active center with ATP and with ADP

The kinetics of the fluorescence enhancement and the transient release of H+ caused by the binding of ADP to the active center of myosin has been compared to that caused by myosin-ATP interaction. The results show that both the time courses of the fluorescence enhancement and the transient H+ release caused by ADP binding, like that caused by ATP hydrolysis in the initial burst, are monophasic exponential processes. The fact that the rates of these two processes are also equal suggests that they both reflect the same mechanistic event in the mechanism of ADP binding. The kinetics of ADP binding as measured by the fluorescence enhancement and the H+ release is different from that of ATP. This is in agreement with our previous finding that the enhancement of fluorescence and the transient release of H+, in the case of ATP, reflect the initial burst of ATP hydrolysis, whereas in the case of ADP, they represent a conformational change in the myosin-ADP complex. The magnitude of the H+ transient caused by the initial burst is approximately equal to that caused by ADP binding. The amplitude of the fluorescence enhancement caused by ADP binding is equal to one-third of that caused by the initial burst.     

Activation of Amiloride-Sensitive Sodium Transport in C6 Glioma Cells

We have characterized, in C6 cells, an amiloride-sensitive Na+ entry pathway that can exchange for H+. In this report we demonstrate that this cation-exchange system can be induced within 24-36 h by either serum removal or by dibutyryl cyclic AMP; however, these modes of induction are not additive and are manifest only after activation by serum. In these glioma cells we found that activation by serum can be mimicked in part by specific serum factors, i.e., epidermal growth factor and bradykinin. We attempted to characterize this activation process further using several cell biologic probes. We had previously shown that that activation process involves a calcium-dependent step with full activation obtained in the presence of the calcium ionophore A23187. The activation by serum was inhibited by preincubation with colchicine but not with dihydrocytochalasin B, suggesting a cytoskeletal involvement in the activation process. Activation by epidermal growth factor and bradykinin was found to be unaffected by colchicine, suggesting that other factors must be present in serum that confer sensitivity to colchicine. Incubation of the cells with phorbol myristoyl acetate results in the activation of amiloride-sensitive transport, suggesting that stimulation of protein kinase C may be integral to the activation process. Unlike the effects of serum, activation by phorbol myristoyl acetate is not inhibited by colchicine, indicating that this drug works in a way that bypasses the cytoskeletal-dependent step. Since diacylglycerol is the presumed endogenous activator of protein kinase C, we studied the effects of dioleylglycerol. This intermediate of phospholipid turnover was found to increase specifically the amiloride-sensitive sodium pathway.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inheritance of glutenin protein subunits of wheat

Summary The inheritance of the high-molecular-weight (HMW) glutenin protein subunits in hexaploid wheat has been investigated by using sodium dodecyl sulphate-polyacrylamide gel electrophoresis to examine the segregation of these subunits in 496 test-cross seeds. The parents of the f₁ hybrid were chosen so that the test-cross seeds segregated for all the HMW glutenin bands. Two glutenin subunits from one parent, believed to be controlled by genes on chromosome 1D, segregated as alternatives to two glutenin subunits from the other parent, a result that supports the assumption that these subunits are controlled by allelic genes at each of two loci that are very closely linked. Similar results were obtained for glutenin subunits believed to be controlled by chromosome IB, which suggests that these subunits are controlled also by allelic genes at each of two loci that are very closely linked. A single glutenin subunit band, believed to be controlled by chromosome 1A, segregated as an alternative to a single glutenin band from the other parent, except that one seed did not possess either band. It was concluded that these bands are controlled either by allelic genes or by nonallelic genes that are very closely linked.     

Action of con A-induced suppressor cells on a B hybridoma line

It has been shown that myeloma immunoglobulin production may be regulated by carrier-specific T_H or T_S cells, by idiotype reactive or alloreactive T cells, by antigen itself, or by immune complexes. However, regulation by nonspecific suppressor cells generated by CON A has not hitherto been shown in that system, possibly, it was proposed, due to the committed state of that cell line. We describe here a system in which immunoglobulin production by a hybridoma line is subject to regulation by CON A-activated spleen cells. This system allows us to show clearly that the suppression we observe here is not due to absorption of T_H factor since no such factors are required for antibody production by the hybridoma line. It could still, however, be argued that the suppression observed in these experiments is mediated by macrophages activated by T-cell products. This system has the advantage of being easily dissected to allow further analysis of the mechanism of nonspecific suppression.     

Mechanisms of monomeric and dimeric glycogenin autoglucosylation

Initiation of glucose polymerization by glycogenin autoglucosylation at Tyr-194 is required to prime de novo biosynthesis of glycogen. It has been proposed that the synthesis of the primer proceeds by intersubunit glucosylation of dimeric glycogenin, even though it has not been demonstrated that this mechanism is responsible for the described polymerization extent of 12 glucoses produced by the dimer. We reported previously the intramonomer glucosylation capability of glycogenin without determining the extent of autoglucopolymerization. Here, we show that the maximum specific autoglucosylation extent (MSAE) produced by the non-glucosylated glycogenin monomer is 13.3 ± 1.9 glucose units, similar to the 12.5 ± 1.4 glucose units measured for the dimer. The mechanism and capacity of the dimeric enzyme to carry out full glucopolymerization were also evaluated by construction of heterodimers able to glucosylate exclusively by intrasubunit or intersubunit reaction mechanisms. The MSAE of non-glucosylated glycogenin produced by dimer intrasubunit glucosylation was 16% of that produced by the monomer. However, partially glucosylated glycogenin was able to almost complete its autoglucosylation by the dimer intrasubunit mechanism. The MSAE produced by heterodimer intersubunit glucosylation was 60% of that produced by the wild-type dimer. We conclude that both intrasubunit and intersubunit reaction mechanisms are necessary for the dimeric enzyme to acquire maximum autoglucosylation. The full glucopolymerization capacity of monomeric glycogenin indicates that the enzyme is able to synthesize the glycogen primer without the need for prior dimerization.     

Studies on the mechanism of natural killer cell-mediated cytotoxicity. VII. functional comparison of human natural killer cytotoxic factors with recombinant lymphotoxin and tumor necrosis factor

The present study was undertaken to evaluate the possible contribution of other cytokines to the lytic activity of NKCF-containing supernatants. We compared some of the functional properties of human NKCF and purified recombinant human rLT and rTNF. It was found that the target cell specificity of rLT was quite different from NKCF in that rLT was neither species specific nor NK specific. Furthermore, antibodies against rLT did not affect the lytic activity of NKCF. These results demonstrate that LT does not significantly contribute to the lytic activity mediated by NKCF. The target specificity of rTNF was found to be related to that of NKCF with the exception of one NK-resistant cell line that was lysed by rTNF in a 20-hr 51Cr-release assay. However, rTNF was not toxic to any of the target cells tested as assessed by trypan blue exclusion in a 20-hr assay unless the targets were labeled with 51Cr. In contrast, NKCF did kill target cells as detected by trypan blue exclusion that were not labeled with 51Cr. Further analysis of this mechanistic difference in the lytic activity of rTNF and NKCF revealed that rTNF in combination with either cycloheximide or mitomycin C but not IFN-gamma could lyse unlabeled U937 target cells. In addition, pretreatment of U937 target cells with nonradioactive Na2CrO4 at concentrations equivalent to that used to 51Cr-labeled cells resulted in their susceptibility to lysis by rTNF as assessed by trypan blue exclusion. These findings suggest that lysis of several susceptible target cells in 20 hr by rTNF requires the presence of additional agents that may be sublethally toxic and/or inhibitory to macromolecular synthesis. Antibody inhibition studies revealed that anti-TNF mediated from partial to complete inhibition of lysis of U937 by unfractionated supernatants containing NKCF. However, fractionation of such supernatants on chromatofocusing columns yielded two distinct peaks of activity eluting in the pH range of 5 to 6 and 7 to 8. Anti-TNF could inhibit the acidic form of NKCF but not the neutral form. It is concluded that NKCF activity is mediated in part by TNF or an antigenically related molecule as well as some other distinct factor(s). The lack of consistent inhibition of NK CMC by anti-TNF suggests that TNF alone is not sufficient to mediate NK activity, or else it is inaccessible to the added antibody.     

Differential effect of zinc on teratogen-induced inhibition of pinocytosis by cultured rat yolk sac

Pinocytosis as measured by the uptake of 125I labelled PVP by the isolated cultured day 12 rat yolk sac was observed to be linear over a 4 h incubation period and to proceed at a rate of approximately 2.5 microliters/mg protein/h. Cadmium, anti-visceral yolk sac antibody (AVYS) and trypan blue all inhibited pinocytosis in a concentration-dependent fashion when added to the culture medium, although at low concentrations trypan blue was slightly stimulatory. The effect of zinc on the inhibition of pinocytosis by these three teratogens was studied. It was observed that zinc ameliorated the inhibitory effects of cadmium and AVYS, but had no effect on inhibition by trypan blue. These results indicate that the previously demonstrated protective action of zinc against cadmium-induced yolk sac dysfunction is not specific to that agent but extends to inhibition of pinocytosis by AVYS, and further suggest that, because of its refractoriness to zinc, trypan blue-induced inhibition of pinocytosis by yolk sac occurs by a mechanism different from that effected by cadmium and AVYS.     

External push and internal pull forces recruit curvature-sensing N-BAR domain proteins to the plasma membrane

Many of the more than 20 mammalian proteins with N-BAR domains control cell architecture and endocytosis by associating with curved sections of the plasma membrane. It is not well understood whether N-BAR proteins are recruited directly by processes that mechanically curve the plasma membrane or indirectly by plasma-membrane-associated adaptor proteins that recruit proteins with N-BAR domains that then induce membrane curvature. Here, we show that externally induced inward deformation of the plasma membrane by cone-shaped nanostructures (nanocones) and internally induced inward deformation by contracting actin cables both trigger recruitment of isolated N-BAR domains to the curved plasma membrane. Markedly, live-cell imaging in adherent cells showed selective recruitment of full-length N-BAR proteins and isolated N-BAR domains to plasma membrane sub-regions above nanocone stripes. Electron microscopy confirmed that N-BAR domains are recruited to local membrane sites curved by nanocones. We further showed that N-BAR domains are periodically recruited to curved plasma membrane sites during local lamellipodia retraction in the front of migrating cells. Recruitment required myosin-II-generated force applied to plasma-membrane-connected actin cables. Together, our results show that N-BAR domains can be directly recruited to the plasma membrane by external push or internal pull forces that locally curve the plasma membrane.     

Common pathway for the induction of hexose transport by insulin and stress

The effect of stress (heat shock, arsenite, or Semliki Forest virus [SFV] infection) on the induction of increased hexose transport has been compared with that of insulin. All four treatments increase the Vmax for transport by BHK cells three- to five-fold, with little effect (less than 40% decrease) on Km. Hydrogen peroxide and phenylarsine oxide (PAO) prevent the increase in hexose transport induced by stress treatments as effectively as they do that induced by insulin. Pinocytosis is not affected by any of the four treatments. On the other hand, the induction by insulin is sensitive to amiloride, whereas that by arsenite is not. Rat embryo fibroblasts, which respond poorly to insulin, respond well to arsenite, heat shock, or SFV infection. It is concluded that the stress response is mediated by certain compounds that may be common to those required for the action of insulin, but that those compounds act at a stage subsequent to the function of the insulin receptor.     

Endothelial cell-associated platelet-activating factor: a novel mechanism for signaling intercellular adhesion

The binding of neutrophils (polymorphonuclear leukocytes [PMNs]) to endothelial cells (ECs) presents special requirements in the regulation of intercellular adhesion. ECs that are stimulated by certain agonists, including thrombin and cytokines (tumor necrosis factor alpha, interleukin-1), generate molecular signals that induce the adhesion of PMNs (endothelial cell-dependent neutrophil adhesion). Our experiments demonstrate that the mechanism of binding induced by thrombin is distinct from that induced by the cytokines based on the time courses, the requirement for protein synthesis, and differential binding of HL60 promyelocytic leukemia cells to ECs activated by the two classes of agonists. The rapid EC-dependent PMN adhesion (initiated in minutes) that occurs when the ECs are stimulated by thrombin is temporally coupled with the accumulation of platelet-activating factor, a biologically active phosphoglyceride that remains associated with ECs and that activates PMNs by binding to a cell surface receptor. A portion of the newly synthesized platelet-activating factor (PAF) is on the EC surface, as demonstrated by experiments in which the rate of hydrolysis of PAF synthesized by activated ECs was accelerated by extracellular PAF acetylhydrolase. When ECs were treated with exogenous PAF they became adhesive for PMNs; the PMN binding was prevented by incubating the ECs with PAF acetylhydrolase or by treating the PMNs with competitive PAF receptor antagonists. Thus PAF associated with the EC plasma membrane induces PMN binding, an observation supported by experiments in which PAF in model membranes (liposomes) stimulated rapid PMN adhesion to ECs and to cell-free surfaces. In addition, competitive antagonists of the PAF receptor inhibited the binding of PMNs to ECs activated by thrombin and other rapidly acting agonists, but not to ECs activated by tumor necrosis factor alpha, indicating that PAF that is endogenously synthesized by ECs can mediate neutrophil adhesion. These experiments demonstrate a novel mechanism by which a cell-associated phospholipid, PAF, can serve as a signal for an intercellular adhesive event.