Calorimetric studies on monomeric and polymeric actin

Differential scanning calorimetry of polymeric F-actin at pH 8.0 showed that the polymer had a concentration-independent thermal profile with a single transition temperature of 81 degrees C. In contrast, the thermal profile of G-actin was concentration-dependent, and although it resembled the F-actin profile at lower concentrations, it was found to have a more complex profile at higher protein concentrations.     

An actin-destabilizing factor is present in human plasma

Summary Plasma and serum of humans or experimental animals contain a factor which destabilizes F-actin. The factor has no DNAse or thrombin activity and after incubation with F-actin does not modify the position of the actin band on a SDS polyacrylamide gel. Hence it probably depolymerizes F-actin.Acknowledgment. This work was supported by the Swiss National Science Foundation, grant No.3.692-0.76. We thank Drs M. Crippa and C.A. Bouvier for measuring the DNAse and thrombin activities. The technical help of Mrs M. Redard and A. De Almeida and the photographic work of Mr J.-C. Rumbeli and Mr E. Denkinger are gratefully acknowledged.     

An actin-destabilizing factor is present in human plasma.

Plasma and serum of humans or experimental animals contain a factor which destabilizes F-actin. The factor has no DNAse or thrombin activity and after incubation with F-actin does not modify the position of the actin band on a SDS polyacrylamide gel. Hence it probably depolymerizes F-actin.     

Effect of phosphorylation of myosin light chains on interaction of heavy meromyosin with regulated F-actin in ghost fibers

Summary The binding of phosphorylated heavy meromyosin to regulated F-actin in ghost fibers at high Ca²⁺ concentration increases, and at low Ca²⁺ concentration decreases, the anisotropy of intrinsic tryptophan fluorescence of F-actin. The effect is opposite to the effect of the binding of dephosphorylated heavy meromyosin.     

Pinitol,a larval growth inhibitor forHeliothis zea in soybeans

Summary A search for insect growth inhibitors in methanol extracts of soybean leaves resulted in isolation of pinitol. Pinitol caused a 50% reduction in weight gain (ED₅₀) ofHeliothis zea larvae at about 0.7% when added to a synthetic diet. Although myo-inositol is a normal component of the insect diet, it also caused growth inhibition at higher concentrations; ED₅₀4%.The authors are indebted to R.J. Molyneux for a reference sample of pinitol and to J. Baker for aid with the bioassay. Reference to a company and/or product named by the Department is only for purposes of information and does not imply approval or recommendation of the product to the exclusion of others which may also be suitable.     

Effect of phosphorylation of myosin light chains on interaction of heavy meromyosin with regulated F-actin in ghost fibers

The binding of phosphorylated heavy meromyosin to regulated F-actin in ghost fibers at high Ca2+ concentration increases, and at low Ca2+ concentration decreases, the anisotropy of intrinsic tryptophan fluorescence of F-actin. The effect is opposite to the effect of the binding of dephosphorylated heavy meromyosin.     

The turnover of F-actin-bound ADP in vivo

Summary A procedure for estimating the rate of turnover of F-actin-bound ADP in vivo is described. A turnover rate of 0.88 h^–1 was determined for mouse muscle F-actin. The validity of the method when used to estimate the turnover rate of F-actin per se is discussed in relation to the possible exchange of F-actin-bound ADP.Acknowledgments. The invaluable technical assistance of Mr L. Carrington is gratefully acknowledged.     

The neuronal p35 activator of Cdk5 is a novel F-actin binding and bundling protein

The neuronal Cdk5 activator p35 is involved in a multitude of neuronal activities, including cytoskeletal organization. We show here that p35 directly interacts with filamentous actin (F-actin) but not with monomeric actin (G-actin). Through binding, p35 induces the formation of actin bundles and stabilizes F-actin against dilution-induced depolymerization. p35 forms intermolecular self-associations, suggesting that p35 cross-links actin filaments into bundles via its intermolecular self-association. p35 dimerization and association with F-actin occur at the N-terminal region that is absent in the calpain-cleaved product p25, indicating that such p35 properties are lost by its truncation induced under neurotoxic conditions. Using p35 phosphorylated by Cdk5 and a mutational approach, we demonstrate that the phosphorylation of p35 promotes its homodimerization and p35-induced formation of F-actin bundles. In addition, the phosphorylation regulates p35 distribution to microtubule and actin cytoskeletons. Together, these observations define a novel function for p35 in cytoskeletal regulation.     

Disruption of sperm release from insect testes by cytochalasin and β-actin mRNA mediated interference

Release of sperm bundles from moth testes is controlled by the local circadian oscillator. The mechanism which restricts migration of sperm bundles to a few hours each day is not understood. We demonstrate that a daily cycle of sperm release is initiated by the migration of folded apyrene sperm bundles through a cellular barrier at the testis base. These bundles have conspicuous concentrations of actin filaments at their proximal end. Inhibition of actin polymerization by cytochalasin at a specific time of day inhibited sperm release from the testis. Likewise, application of double-stranded actin RNA specifically inhibited sperm release. This RNA-mediated interference (RNAi) lowered the pool of actin mRNA in tissues involved in sperm release. The decline in mRNA levels resulted in the selective depletion of F-actin from the tip of apyrene sperm bundles, suggesting that this actin may be involved in the initiation of sperm release. Combined results of RNAi experiments at physiological, cellular and molecular levels identified unique cells that are critically involved in the mechanism of sperm release.Received 11 April 2003; received after revision 2 May 2003; accepted 27 May 2003     

Radioimmunoassay of some hormones simultaneously measured in serum and breast cyst fluid

Summary Blood and breast cyst fluid were drawn simultaneously for hormonal determination. There was no difference between serum and cyst fluid values of PRL and TSH. A significant difference was noted for LH (p<0.01) and FSH (p<0.05), serum concentrations being higher than cyst fluid concentrations.The authors are indebted to Miss J. Meister, E. Dullaert and J. Beck for their skillful technical help. We are also indebted to the National Institutes of Health, NIAMDD and the World Health Organization, International Laboratory for Biological Standards.     

Freeze tolerance in the frog,Rana sylvatica

Summary Wood frogs survive extracellular freezing at moderate subzero temperatures (–4°C) for at least 11 days. Freezing survival is aided by the accumulation of high concentrations of glucose as a cryoprotectant in blood and tissues. Glucose production was accompanied by a rapid decline in liver, but not muscle, glycogen levels suggesting that liver is the organ controlling cryoprotectant synthesis.Acknowledgments. This study was supported by grants to L.M.G. from the Kroc Foundation (Santa Ynez, California) and from the National Institute of Dental Research (Grant No. DE-03987). The authors wish to thank K. Yorko, M. Shakin, J. Finan and Mrs N. Manivannan for their technical and secretarial assistance.Acknowledgments. I thank Dr J. Ballantyne, Dr F. Schueler and I. McMurray for help with frog collections and Dr. W. Schmid, Dr J. Bogart and Dr F. Cook for helpful discussions. Supported by an N.S.E.R.C. operating grant and by a grant from the Atkinson Charitable Foundation.     

Swiprosin-1 modulates actin dynamics by regulating the F-actin accessibility to cofilin

Membrane protrusions, like lamellipodia, and cell movement are dependent on actin dynamics, which are regulated by a variety of actin-binding proteins acting cooperatively to reorganize actin filaments. Here, we provide evidence that Swiprosin-1, a newly identified actin-binding protein, modulates lamellipodial dynamics by regulating the accessibility of F-actin to cofilin. Overexpression of Swiprosin-1 increased lamellipodia formation in B16F10 melanoma cells, whereas knockdown of Swiprosin-1 inhibited EGF-induced lamellipodia formation, and led to a loss of actin stress fibers at the leading edges of cells but not in the cell cortex. Swiprosin-1 strongly facilitated the formation of entangled or clustered F-actin, which remodeled the structural organization of actin filaments making them inaccessible to cofilin. EGF-induced phosphorylation of Swiprosin-1 at Ser183, a phosphorylation site newly identified using mass spectrometry, effectively inhibited clustering of actin filaments and permitted cofilin access to F-actin, resulting in actin depolymerization. Cells overexpressing a Swiprosin-1 phosphorylation-mimicking mutant or a phosphorylation-deficient mutant exhibited irregular membrane dynamics during the protrusion and retraction cycles of lamellipodia. Taken together, these findings suggest that dynamic exchange of Swiprosin-1 phosphorylation and dephosphorylation is a novel mechanism that regulates actin dynamics by modulating the pattern of cofilin activity at the leading edges of cells.     

Seasonal changes in the critical arousal temperature of the marsupialSminthopsis crassicaudata correlate with the thermal transition in mitochondrial respiration

Summary During periods of torporSminthopsis crassicaudata, a dasyurid marsupial, regulated its body temperature above about 16.3°C in summer and 13.0°C in winter. Animals with lower body temperatures were unable to arouse. Liver, heart and brain mitohondrial succinate:cytochrome c reductase showed a thermal transition at 16°C in summer and at 12.5°C in winter. Thus the lowest regulated body temperature was just above the temperature where changes were detected in mitochondrial respiration.Acknowledgments. This work was supported by a Flinders University Research Scholarship Award to F. Geiser. We thank Prof. J. H. Bennett for the supply of animals and M. O'Driscoll for the animal maintenance.     

The turnover of F-actin-bound ADP in vivo.

A procedure for estimating the rate of turnover of F-actin-bound ADP in vivo is described. A turnover rate of 0.88 h-1 was determined for mouse muscle F-actin. The validity of the method when used to estimate the turnover rate of F-actin per se is discussed in relation to the possible exchange of F-actin-bound ADP.     

Calcite growth under controlled diffusion

Summary The growth of calcite was studied in a gelatin-gel medium under variable environmental conditions by 2 different methods. The results suggest that the organic matrix, the temperature, the diffusion fluctuation depending on ionic concentrations, and the presence of additives exert a fine control on the evolution of single crystals, polycrystalline aggregates, and highly structured concretions of calcite.Presented at the 22nd Canadian Conference on Pharmaceutical Research, Montreal (Quebec), Canada, 16 May 1975.Acknowledgments. This work was supported, by the Medical Research Council of Canada. — The authors are indebted to Dr J. M. Dorlot, Department of Metallurgy, Ecole Polytechnique, Montreal, for the use of the scanning electron microscope and thanks are due to Mr J. Claudinon for his technical assitance.     

The actin-binding domain of actin filament-associated protein (AFAP) is involved in the regulation of cytoskeletal structure

Actin filament-associated protein (AFAP) plays a critical role in the regulation of actin filament integrity, formation and maintenance of the actin network, function of focal contacts, and cell migration. Here, we show that endogenous AFAP was present not only in the cytoskeletal but also in the cytosolic fraction. Depolymerization of actin filaments with cytochalasin D or latrunculin A increased AFAP in the cytosolic fraction. AFAP harbors an actin-binding domain (ABD) in its C-terminus. AFAPΔABD, an AFAP mutant with selective ABD deletion, was mainly in the cytosolic fraction when overexpressed in the cells, which was associated with a disorganized cytoskeleton with reduced stress fibers, accumulation of F-actin on cellular membrane, and formation of actin-rich small dots. Cortactin, a well-known podosome marker, was colocalized with AFAPΔABD in these small dots at the ventral surface of the cell, indicating that these small dots fulfill certain criteria of podosomes. However, these podosome-like small dots did not digest gelatin matrix. This may be due to the reduced interaction between AFAPΔABD and c-Src. When AFAPΔABD-transfected cells were stimulated with phorbol ester, they formed podosome-like structures with larger sizes, less numerous and longer life span, in comparison with wild-type AFAP-transfected cells. These results indicate that the association of AFAP with F-actin through ABD is crucial for AFAP to regulate cytoskeletal structures. The AFAPΔABD, as cytosolic proteins, may be more accessible to the cellular membrane, podosome-like structures, and thus be more interactive for the regulation of cellular functions.     

Tubes,randomness, and Brownian motions: or,how engineers learned to start worrying about electronic noise

In this paper, we examine the pioneering research on electronic noise—the current fluctuations in electronic circuit devices due to their intrinsic physical characteristics rather than their defects—in Germany and the U.S. during the 1910s–1920s. Such research was not just another demonstration of the general randomness of the physical world Einstein’s work on Brownian motion had revealed. In contrast, we stress the importance of a particular engineering context to electronic noise studies: the motivation to design and improve high-gain thermionic-tube amplifiers for radio and wired communications. Engineering scientists’ endeavors to understand electronic noise started in 1918, when Walter Schottky at Siemens formulated a theory of “shot noise,” current fluctuations owing to the random emissions of discrete electrons in a tube. Schottky’s theory was revised and experimentally tested at Siemens, General Electric, and AT&T during the 1920s, leading to the discoveries of several other types of noise and an increasing interest in the thermal fluctuations in electronic circuits. In 1925–1928, J.B. Johnson and Harry Nyquist at Bell Labs developed a theory of thermal noise for any electrical resistor at a non-zero temperature. Although these studies were initiated to chart the fundamental performance limit of electronic technology, they ended up assisting the empirical determination of individual electronic components’ characteristics.     

连续性血液净化治疗对重症急性胰腺炎患者内皮细胞通透性的影响

目的探讨连续性血液净化治疗对重症急性胰腺炎(severeacutepancreatitis,SAP)患者内皮细胞的作用.方法人脐静脉内皮细胞(HumanUmbilicalVeinEndothelialCells,HUVEC)按实验分组分别用16例健康对照组血清,38例 SAP患者连续性血液净化(ContinuousBloodPurificantion,CBP)治疗前、治疗6h、治疗20h血清体外干预脐静脉内皮细胞5h.transwell小室观察内皮细胞通透性变化,激光共聚焦显微镜观察 F actin应力微丝的表达及分布.结果 SAP患者 CBP治疗前内皮细胞通透性较健康对照组明显增高(P<0.01),CBP治疗6h、治疗20h后通透性均降低(P<0.05),但治疗20h组与治疗6h组相比,差异无显著性(P>0.05).激光共聚焦结果显示,与健康对照组相比,CBP治疗前 SAP患者内皮细胞 F actin应力微丝的数量及密度均明显增加,CBP治疗6h后,F actin应力微丝形成减少,治疗20h后减少更加显著.结论重症急性胰腺炎患者内皮细胞通透性明显增加,CBP治疗可以显著降低 SAP患者内皮细胞通透性,其机制可能与调节 F actin应力微丝形成与分布有关     

Anti-feedant activity of the polyacetylene,phenylheptatriyne (PHT), from the Asteraceae toEuxoa messoria (Lepidoptera: Noctuidae)

Summary Phenylheptatriyne (PHT), a polyacetylene from various species of Asteraceae reduced feeding and weight gain of larvae of the polyphagous insectEuxoa messoria when incorporated into an artificial diet at concentrations of 10–300 ppm. These results suggest a role as insect antifeedants for the widely distributed polyacetylenes of the Asteraceae.Acknowledgment. This work was supported by NSERC and Agriculture Canada (E.M.R.) We thank Dr R. J. Byers (Agriculture Canada) for eggs ofEuxoa. To whom reprint requests should be addressed.     

A cold-active salmon goose-type lysozyme with high heat tolerance

The Atlantic salmon (Salmo salar) goose-type lysozyme gene was isolated and revealed alternative splicing within exon 2 affecting the signal peptide-encoding region. The lysozyme was produced in Escherichia coli, and the recombinant enzyme showed a high specific lytic activity that was stimulated by low or moderate concentrations of mono- or divalent cations. Relative lytic activities of 70 and 100% were measured at 4°C and 22°C, respectively, and there was no detectable activity at 60°C. However, 30% activity was retained after heating the enzyme for 3 h at 90°C. This unique combination of thermal properties was surprising since the salmon goose-type lysozyme contains no cysteines for protein structure stabilization through disulphide bond formation. The results point to a rapid reversal of inactivation, probably due to instant protein refolding. Received 14 August 2007; received after revision 07 September 2007; accepted 12 September 2007