Zinc-stimulated microtubule assembly and evidence for zinc binding to tubulin

• 1.1. Microtubule reassembly was studied in supernatant fluids from rat brain. Tubulin in extracts from zinc-deficient animals showed an impaired ability to repolymerize compared to extracts from controls; 10 μM zinc stimulated reassembly of tubulin in extracts from zinc-deficient animals.
• 2.2. Low zinc concentrations (250–900 μM zinc in the presence of 1 mM EGTA) stimulated reassembly oftubulin in brain extracts from control rats; similar concentrations of nickel had no effect whilst cobalt was inhibitory. In the absence of EGTA 20–40 μM zinc stimulated reassembly in brain extracts from normal rats.
• 3.3. Zinc-induced changes in reassembly were associated with changes in the free sulphydryl group content of the assembled crude microtubule protein; increased assembly was associated with a higher free sulphydryl group content, decreased assembly with a lower content.
• 4.4. ⁶⁵Zn was found to bind to tubulin. This binding was partly inhibited by N-ethylmaleimide.

     

Characteristics of the in vitro assembly of brain tubulin of Cyprinus carpio

• 1.1. Tubulin has been isolated from brain of carp (Cyprinus carpio), acclimated to summer temperatures (16–20°C), and its in vitro reassembly behavior has been characterized.
• 2.2. Among the striking properties of this tubulin preparation is the temperature profile showing a high level of polymerization at the environmental temperature of carp.
• 3.3. The critical tubulin concentration for assembly was 0.8 mg/ml, which was higher than mammalian tubulin purified by the cycle procedure.
• 4.4. The microtubular protein showed three high mol.wt component and a minor component of about 43,000 daltons was also found to copurify with tubulin.

     

Impaired microtubule assembly in brain from zinc-deficient pigs and rats

• 1.1. Microtubule reassembly was studied in brain extracts from pigs and rats deficient in zinc and from zinc-supplemented controls. Tubulin in extracts from zinc-deficient animals showed an impaired ability to repolymerize compared with extracts from control animals.
• 2.2. Crude microtubule protein isolated from zinc-deficient rat brain contained less free SH groups than that isolated from zinc-supplemented animals.
• 3.3. It is concluded that variation in zinc concentration close to the physiological range can influence microtubule assembly, and that zinc may have some function in microtubule polymerization in vivo.

     

Evidence that the 5' end of rabbit globin mRNA is hybridized with its 3' end

• 1.1. A radiopolyadenylated rabbit globin mRNA was treated with different concentrations of ribonuclease V₁ from cobra venom.
• 2.2. The enzymatic digests were chromatographed on an aminophenylboronate-agarose column, which specifically captured the cap structure i.e. n⁷G(5') ppp (5') NmP.
• 3.3. When the capture fragment was chromatographed on a Sephadex G-100 column, its size was smaller than the native molecule and also bore radioactivity, i.e. a poly(A) tail.
• 4.4. These results provide evidence that the 5' end (which encompasses the cap structure) of rabbit globin mRNA is hybridized and in close proximity to its 3' end.
• 5.5. We conclude that this conformation is required for messenger translation efficiency.

     

Characteristics of tubulin aggregation by tubulin-binding proteins from brain and by synthetic polycations

• 1.1. A basic fraction from brain cytosol was found to contain two or more tubulin-binding proteins, able to induce aggregation of tubulin accompanied by hydrolysis of GTP.
• 2.2. In some respects tubulin aggregation by the brain proteins was similar to its aggregation by polylysine.
• 3.3. The burst of GTP hydrolysis accompanying tubulin aggregation by polylysine had the following characteristics: enhanced by salt and abolished by low temperature; not stoichiometric with the amount of tubulin precipitated and actually maximal at relatively low polylysine concentration; uncoupled temporally from aggregation, which occurred over a much shorter interval.

     

Thrombocyte aggregation in rainbow trout

• 1.1. Thrombocytes from mature rainbow trout (Sahmo gairdneri) aggregated in vitro after addition of ADP, ATP, collagen, epinephrine, 5-hydroxytryptamine or thrombin to thrombocyte-rich plasma.
• 2.2. Thrombocyte aggregation was dose dependent and could be inhibited by preincubating the thrombocyte-rich plasma with adenosine, acetylsalicylic acid or prostaglandin E₁.
• 3.3. Thrombocytes released ADP and ATP when aggregated by 10 μM epinephrine. This release was blocked by 1 mM adenosine.
• 4.4. Electron microscopic observations of thrombocytes revealed them to contain numerous microtubules and electron-dense vesicles. In addition a possible shape change associated with thrombocyte aggregation was noted.

     

Origin of kinetochore microtubules in Chinese hamster ovary cells

We have attempted to determine whether chromosomal microtubules arise by kinetochore nucleation or by attachment of pre-existing microtubules. The appearance of new microtubules was investigated in vivo on kinetochores to which microtubules had not previously been attached. The mitotic apparatus of Chinese hamster ovary cells was reconstructed in three dimensions from 0.25 m thick serial sections, and the location of chromosomes, kinetochore outer disks, centrioles, virus-like particles and microtubules determined. Central to the interpretation of these data is a synchronization scheme in which cells entered Colcemid arrest without forming mitotic microtubules. Cells were synchronized by the excess thymidine method and exposed to 0.3 g/ml Colcemid for 8 h. Electron microscopic examination showed that this Colcemid concentration eliminated all microtubules. Mitotic cells were collected by shaking off, and cell counts showed that over 95% of the cells were in interphase when treatment began and thus were arrested without the kinetochores having been previously attached to microtubules. Cells were then incubated in fresh medium and fixed for high voltage electron microscopy at intervals during recovery. — In early stages of recovery, short microtubules were observed near and in contact with kinetochores and surrounding centrioles. Microtubules were associated with kinetochores facing away from centrosomes and far from any centrosomal microtubules, and thus were not of centrosomal origin. At a later stage of recovery, long parallel bundles of microtubules, terminating in the kinetochore outer disk, extended from kinetochores both toward and away from centrosomes. Because microtubules had never been attached to kinetochores, the possibility that kinetochore microtubles were initiated by microtubule stubs resistant to Colcemid was eliminated. Therefore we conclude that mammalian kinetochores can initiate microtubules in vivo, thus serving as microtubule organizing centers for the mitotic spindle, and that formation of kinetochore-microtubule bundles is not dependent on centrosomal activity.     

Mechanism of neurofibrillary degeneration in Alzheimer's disease

Neurofibrillary degeneration associated with the formation of intraneuronal neurofibrillary tangles of paired helical filaments (PHF) and 2.1 nm τ filaments is one of the most characteristic brain lesions of Alzheimer's disease. The major polypeptides of PHF are the microtubule associated protein, τ. τ, in PHF is present in abnormally phosphorylated forms. In addition to the PHF, the abnormal τ is present in soluble non-PHF form in the alzheimer's disease brain. The level of τ in Alzheimer's disease neocortex is severalfold higher than in aged control brain, and this increase is in the form of the abnormally phosphorylated protein. The abnormally phosphorylated τ does not promote the assembly of tubulin into microtubules in vitro, and it inhibits the normal τ-stimulated microtubule assembly. After in vitro dephosphorylation both PHF and non-PHF abnormal τ stimulate the assembly of tubulin into microtubules. The activities of phosphoseryl/phosphothreonyl protein phosphatase 2A and nonreceptor phosphotyrosyl phosphatase(s) are decreased in AD brain. It is suggested that

1. A defect(s) in the protein phosphorylation/dephosphorylation system is one of the early events in the neurofibrillary pathology in AD;
2. A decrease in protein phosphatase, activities, at least in part, allows the hyperphosphorylation of τ; and
3. Abnormal phosphorylation and polymerization of τ into PHF most probably lead to a breakdown of the microtubule system and consequently to neuronal degeneration.

     

Quantification of Dynamic Protein Interactions and Phosphorylation in LPS Signaling Pathway by SWATH-MS

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Highlights

• •Intensive investigation of dynamic interactions in MyD88, TRAF6, and NEMO complexes.
• •Mechanistic insights into IRAKs proteins' assembly.
• •A signal amplification mechanism for MyD99-denpendent TLR signaling disclosed by stoichiometry of complexes.
• •Quantitative measurement of multiple phosphorylation sites on the key components in TLR signaling pathway.

     

The effects of cooling the egg on the respiratory movements of the hatching fowl,Gallus g. domesticus,with a note on vocalization

• 1.1. The ambient temperature of pipped eggs of the domestic fowl was reduced from 39 to 20°C for a period of 2 hr.
• 2.2. In the majority of embryos the amplitude of respiration more than doubled during the first hour, and in each embryo the frequency fell to a minimum value by the end of cooling.
• 3.3. When the eggs were rewarmed the respiratory frequency usually returned to normal within a period of 1 hr. Vocal activity was often stimulated and was accompanied by a marked increase in the amplitude of respiration.
• 4.4. These results are discussed in relation to the development of thermoregulation, and are compared with results obtained elsewhere in a similar study on the quail.

     

Organ specific changes in energy metabolism due to anaerobiosis in the sea mussel Mytilus edulis (L.)

• 1.1. Anaerobic energy metabolism was investigated in different organs of Mytilus edulis and the whole animal.
• 2.2. Succinate accumulates to high levels in most organs but remains low in the hemolymph.
• 3.3. After 16 hours propionate accumulation is observed in all organs. Experimental evidence is not sufficient yet to point out organs that produce more propionate than others.
• 4.4. Acetate is a minor end product.
• 5.5. Acetate and propionate are found in the hemolymph in amounts equal to those in the organs.
• 6.6. Animals incubated in oxygen-free seawater accumulate more end products than animals exposed to air, in the form of volatile fatty acids that are excreted into the incubation water.
• 7.7. Alanine and glutamine increase in the posterior adductor muscle. Aspartate decreases in the total animal, posterior adductor muscle and gills, while in the hemolymph decrease in alanine, asparagine, serine, threonine and proline are observed.

     

Factors regulating the functioning of the in vitro perfused aglomerular kidney of the anglerfish,Lophius piscatorius L.

• 1.1. Kidneys of Lophius were perfused from the renal portal vein with a Ringer's solution.
• 2.2. Mammalian and piscine neurohypophysial hormones (in doses of 20–500 ng/kg body wt) did not affect the rate of urine production or the urinary concentration of inorganic ions.
• 3.3. The rate of urine production and the urinary concentration of magnesium and sodium ions varied with the concentration of magnesium in the perfusate.
• 4.4. The rate of urine production was positively correlated with urine magnesium concentration (r = 0.83 ± 0.04) and negatively correlated with that of sodium (r = −0.40).
• 5.5. The urinary concentration of sodium ions varied inversely with that of magnesium ions (r = −0.89).
• 6.6. Ouabain treatment (0.1–0.8 mM/l) reduced the rate of urine production by over 60% and altered, to varying extents, the pattern of electrolyte excretion. A simple model for the mode of formation of urine by the aglomerular kidney, based on the present results and other observations is suggested.

     

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Identification of Novel Protein Targets of Dimethyl Fumarate Modification in Neurons and Astrocytes Reveals Actions Independent of Nrf2 Stabilization

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Highlights

• •Dimethyl fumarate covalently modifies cysteine residues in neurons and astrocytes.
• •Cofilin-1, tubulin and collapsin response mediator protein 2 (CRMP2) are targets.
• •DMF-modified cofilin-1 reduces actin-severing ability, preserving filamentous actin.

     

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Breast,hips, and buttocks revisited: Honest fatness for honest fitness

This paper comments on: Low, B. S., Alexander, R. D., and Noonan, K.M. Human hips, breast, and buttocks: Is fat deceptive? Ethology and Sociobiology 8: 249-247, 1987. In it I argue that:

• 1.1. Sexual selection has probably not been the most important selection pressure on
• 2.female human body shape.
• 3.2. Male humans in different cultures find different aspects of the female body attractive
• 4.and therefore are unlikely to have exerted consistent directional sexual selection on
• 5.the female body.
• 6.3. Breast size is not correlated with lactation success.
• 7.4. Visible hip width is not correlated with parturition success.
• 8.5. Women would lower their fitness if they tried to deceive men about their internal
• 9.pelvic dimensions.
• 10.6. There are many alternative hypothesis to explain the existence of fat onwomen's
• 11.breast, hips, and buttocks.

     

Relationship between brain trehalase activity and trehalosema during direct and diapausing development in Pieris brassicae

• 1.1. Brain trehalase specific activity and trehalosemia were measured during the end of the developmental life cycle in non-diapausing and diapausing insects.
• 2.2. During non-diapausing development, trehalosemia reached maximum values at the beginning of pupal life. Then a constant decrease was observed up to the end of adult life.
• 3.3. The specific activity of brain trehalase was maximum when the insects were in active feeding periods, minimum activity appearing during moulting phases.
• 4.4. During diapausing development, trehalosemia was very high at the beginning of pupal life, particularly when insects were exposed to wintering conditions.
• 5.5. When diapause was broken, trehalosemia fell, announcing adult emergence.
• 6.6. Brain trehalase activity showed the same qualitative variations as in non-diapausing larvae, but with rather lower values.
• 7.7. During pupal life, brain trehalase activity decreased markedly during the long period necessary to obtain diapause breakdown.
• 8.8. Wintering conditions allow a progressive increase of brain trehalase activity, which preceded the fall of trehalosemia.