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A Forer

Publications and source records attributed to A Forer.

At least 55 records · Page 3Linked to original sources
PubMed1984

Composition of the isolated mitotic apparatus: analysis by polyacrylamide gel electrophoresis.

The mitotic apparatus (MA) from zygotes of the sea urchin Strongylocentrotus purpuratus were isolated and the components of the isolated MA were studied using one-dimensional polyacrylamide gel electrophoresis. MA were isolated using two different stabilization media, hexylene glycol and glycerol-dimethylsulphoxide ( MTM ), because previous studies have shown that MA isolated with these solutions have different properties. Both MA have at least 15-20 bands on gels. Whereas a few bands might be unique to one method of isolation, most bands are the same in hexylene glycol MA and MTM MA. Both kinds of MA differ from the general cytoplasm. Detailed descriptions of the appearances of the gels are given. The main difference between the two kinds of MA is in the relative intensities of the bands found in both MA. The physical and chemical properties of MA change during storage. After various durations of storage MA were extracted with 0.5 M KCl: more proteins were extracted from freshly isolated MA than from stored MA. There were changes in extractibility which occurred at the same time as the physiological changes, but the particular changes in hexylene glycol MA were not the same as those in MTM MA. Thus, there is no common denominator that would allow us to say that loss of extractibility is associated with changes in a certain protein of the isolated MA.

Animals↗
PubMed1983

Action spectrum for changes in spindle fibre birefringence after ultraviolet microbeam irradiations of single chromosomal spindle fibres in crane-fly spermatocytes.

Single chromosomal spindle fibres in Nephrotoma suturalis (crane-fly) spermatocytes in metaphase and anaphase were irradiated with monochromatic ultraviolet light focussed to a 2 micrometer spot. In cells in both metaphase and anaphase either the birefringence of the irradiated spindle fibre was altered in the irradiated region, or there was no change, depending on the dose and wavelength of ultraviolet light used for the irradiation. When there was an area of reduced birefringence (ARB), it moved poleward regardless of whether the associated chromosome moved poleward. When cells were irradiated in early metaphase they remained in metaphase until the ARB reached the pole. In some cells irradiated in late metaphase the chromosomes began anaphase before the ARB reached the pole; in many such cells anaphase was abnormal in that all six half-bivalents separated at the start of anaphase but none moved polewards. In all cases the ARB moved poleward at the same speed as subsequent chromosome movement; that is, at about 0.8 micrometer/min. In cells irradiated in anaphase, spindle fibre birefringence was reduced independently of blockage of chromosome movement. Because birefringence and movement were altered independently there were four classes of results: (1) in some cases there was no effect on the movement of the chromosome associated with the irradiated spindle fibre and no effect on the birefringence of the irradiated spindle fibre. (2)In some cases, primarily with 260 nm wavelength light, there was no effect on the movement of the chromosome associated with the irradiated spindle fibre and there was an effect on the birefringence of the irradiated spindle fibre. (3) In some cases, primarily with 290 nm wavelength light, there was an effect on the movement of the chromosome associated with the irradiated spindle fibre and no effect on the birefringence of the irradiated spindle fibre. (4) In some cases, primarily with 270 nm and 280 nm wavelength light, there was an effect on the movement of the chromosomes associated with the irradiated spindle fibre and there was an effect on the birefringence of the irradiated spindle fibre. The action spectrum for reducing spindle fibre birefringence in crane-fly spermatocytes had two peaks, one at 260 nm and the other, less sensitive, at 280 nm. For irradiations at 270 nm, 280 nm and 290 nm, five to fifty times more energy was needed to reduce spindle fibre birefringence than to stop chromosome movement, but for irradiations at 260 nm five times less energy was needed to reduce spindle fibre birefringence than to stop chromosome movement. The action spectrum for reducing spindle fibre birefringence is quite different from that for stopping chromosome movement.

Anaphase↗
PubMed1983

Prometaphase forces towards opposite spindle poles are not independent: an on/off control system is identified by ultraviolet microbeam irradiations.

Individual spindle fibres in prometaphase spermatocytes of the cricket, Neocurtilla hexadactyla, were irradiated with an ultraviolet microbeam. The stretched heteromorphic bivalent (X2Y) contracted to about 75% of its pre-irradiation length after irradiation of either of its two oppositely directed spindle fibres (i.e., the X2 or Y spindle fibre). The X2Y bivalent also contracted after irradiation of the connection between the kinetochores of the univalent X1 chromosome and the Y chromosome but it did not contract after irradiation of autosomal spindle fibres or of the spindle fibre of the X1 univalent sex chromosome. The spindles sometimes shrank after irradiation, but contraction of the X2Y bivalent was independent of spindle shrinkage. The data strongly suggest that the oppositely directed forces on a bivalent are not independent. One reason is that the X2Y contractions were asymmetrical: during contraction one kinetochore remained fixed in position while the other kinetochore (facing the opposite pole) moved towards the equator. In 75% of the cases the stationary kinetochore was associated with the irradiated spindle fibre. Thus, we suggest that the irradiation of a spindle fibre produces a state analogous to rigor in the irradiated spindle fibre and, because of effects on the control system, produces relaxation of tension in the oppositely directed non-irradiated spindle fibre, so that the kinetochore associated with the non-irradiated spindle fibre moves towards the equator. These experiments have identified a control system that coordinates force production to opposite poles.

Animals↗
PubMed1981

Analysis of chromosome movement in crane fly spermatocytes by ultraviolet microbeam irradiation of individual chromosomal spindle fibres. II. Action spectra for stopping chromosome movement and for blocking ciliary beating and myofibril contractions.

Chromosome-to-pole movement in crane fly spermatocytes was temporarily blocked by ultraviolet light focussed to a 4-micrometer-diameter spot on single chromosomal spindle fibres. Since similar irradiation of the interzonal region did not alter chromosome-to-pole movement, this effect was specific to spindle fibres. The action spectrum for blocking chromosome movement in this specific way had two peaks, one at 270 nm and one at 290 nm. To block movement, irradiations with 280-nm-wavelength light required two to four times more energy than irradiations with 270- or 290-nm-wavelength light. Action spectra were obtained for blocking ciliary beating and for blocking myofibril contraction. The action spectrum for blocking ciliary beating had a broad peak, between 260 nm and 280 nm, whilst that for blocking myofibril contraction had two peaks, at 270 and 290 nm, just like that for blocking chromosome movement. We discuss the similarities and differences in the various action spectra, and we compare the action spectra to absorption spectra of spindle components and to other action spectra (e.g., that for depolymerizing actin-containing filaments). Absorption spectra were obtained for ultraviolet light passing through spindle fibres as well as for ultraviolet light passing through the interzone.

Animals↗
PubMed1981

Autosomal spindle fibres influence subsequent sex-chromosome movement in crane-fly spermatocytes.

In meiosis-I crane-fly spermatocytes 3 autosomal half-bivalents move to each pole in anaphase while the 2 sex-chromosomal univalents remain at the equator. The sex chromosomes move to opposite poles only after the autosomes reach the poles; the sex chromosomes start to move polewards about 25 min after the autosomal half-bivalents have begun to move. We irradiated portions of single autosomal spindle fibres with an ultraviolet microbeam and found that these irradiation altered the subsequent sex-chromosome movements. Two effects were observed. In one, one of the sex chromosomes did not move at all; the sex cin after the autosomal half-bivalents have begun to move. We irradiated portions of single autosomal spindle fibres with an ultraviolet microbeam and found that these irradiation altered the subsequent sex-chromosome movements. Two effects were observed. In one, one of the sex chromosomes did not move at all; the sex cin after the autosomal half-bivalents have begun to move. We irradiated portions of single autosomal spindle fibres with an ultraviolet microbeam and found that these irradiation altered the subsequent sex-chromosome movements. Two effects were observed. In one, one of the sex chromosomes did not move at all; the sex chromosome that remained at the equator would normally have moved to the pole associated with the irradiated autosomal spindle fibre. In the second, both sex chromosomes moved to the same pole, always that of the non-irradiated side. These effects occurred whether or not autosomal anaphase movement was blocked by the irradiation. There was no wavelength dependence for altering sex-chromosome movements. Sex-chromosome movements were altered only when at least one sex-chromosomal spindle fibre was adjacent to the irradiated autosomal spindle fibre; when neither sex chromosome had a spindle fibre adjacent to the irradiated autosomal spindle fibres the chromosomes always moved normally. Irradiation of sex-chromosomal spindle fibres during sex-chromosomal anaphase showed short blockages of movement (usually 5-8 min), and then complete recovery. Direct irradiation of sex-chromosomal spindle fibres (without irradiating autosomal spindle fibres) when the autosomes were in anaphase but the sex chromosomes were in metaphase never caused abnormal sex-chromosome movements. These results eliminate the possibility that when we irradiated autosomal spindle fibres that were adjacent to sex-chromosomal spindle fibres the sex-chromosomal spindle fibres were irradiated inadvertently and were unable to recover from the damage. We suggest that the irradiations of autosomal spindle fibres alter a control system involved in "turning on' sex-chromosomal spindle fibre motors, rather than directly altering the motors. We suggest that interactions between spindle fibres are somehow involved in this control system.

Animals↗
PubMed1980

Experimental determinations of tubulin in the in vivo mitotic apparatus of sea urchin zygotes.

Mitotic apparatus (MA) were isolated from zygotes of a sea urchin (Strongylocentrotus purpuratus), using hexylene glycol (pH 6.4) as lysing-stabilizing agent. Protein was measured in the MA pellet and in the remainder of the cell lysate (using the Lowry procedure). Tubulin was measured in the MA pellet and in the remainder of the cell lysate (using microdensitometry of stained gels after sodium dodecyl sulphate - polyacrylamide gel electrophoresis). From these data we calculated the maximum possible amounts of tubulin in the isolated MA and in the MA in vivo; in these calculations we assumed that all the tubulin in the cell is associated with the MA, and we assumed that, as reported in the literature, the MA lose 90% of their dry matter during the isolation. We conclude that tubulin probably comprises less than 7% of the protein in the in vivo MA, and, even if there are very large errors, tubulin is considerably less than half the protein in the MA.

Animals↗
PubMed1979

Actin in spindles of Haemanthus katherinae endosperm. I. General results using various glycerination methods.

We have studied actin-containing filaments in spindles in Haemanthus endosperm cells glycerinated by various methods; the actin-containing filaments were identified by their reaction with rabbit skeletal muscle heavy meromyosin (HMM) to form 'decorated' filaments. Actin-containing filaments in the spindle were seen in amongst microtubules in bundles (both non-kinetochore microtubule bundles and kinetochore microtuble bundles) and were also seen not associated with microtubules. There were very few extra-spindle actin-containing filaments in these cells. Actin-containing filaments seemed to interact with microtubules, because the filaments remained close to and parallel to microtubules even when the microtubules were sharply curved. Because of the close association between microtubules and actin-containing filaments we could not identify all the actin-containing filaments present in microtubule bundles: microtubules obscured actin-containing filaments. We studied Haemanthus endosperm cells as they were glycerinated. For some of these observations we used phase-contrast microscopy. Glycerination caused the cells to shrink, initially, and this was followed by rapid expansion, but the cells did not expand to as large a volume as before glycerination. Spindle structure was maintained despite these changes in cell size. Evidences for this are that relative chromosome positions were maintained during glycerination, that spindle birefringence was maintained during glycerination, and that individual chromosomal spindle fibres remained birefringent during glycerination. Electron-microscopic observations supported this in that kinetochore microtubule bundles and non-kinetochore microtubule bundle were maintained during glycerination, as was the helical arrangement of spindle ribosomes into polyribosomes. One-step glycerination procedures were used (cells were treated with mixtures containing 25% glycerol, Triton-X-100 and HMM), and such procedures might be of general use. Living cells were embedded in fibrin clots in making light-microscopic observations; this procedure, too, might be of general use.

Actins↗
PubMed1979

Actin in spindles of Haemanthus katherinae endosperm. II. Distribution of actin in chromosomal spindle fibres, determined by analysis of serial sections.

We have studied the arrangements of actin-containing filaments in 13 bundles of kinetochore microtubules in glycerinated, heavy meromyosin-treated Haemanthus endosperm cells: 7 bundles were in a cell at anaphase, and 6 were in a cell at metaphase. Actin-containing filaments were present in each of the 13 bundles of kinetochore microtubules: they were in amongst the microtubules in the bundle and seemed to be associated with the microtubules. Actin-containing filaments in each bundle seemed to terminate at the kinetochores. Actin-containing filaments associated with the kinetochore microtubules were of consistent polarity (the arrowheads pointed towards the kinetochores) whereas those associated with other microtubles and those not associated with microtubules did not have consistent polarity (some pointed towards the spindle pole, others pointed away from it). Roughly, there were as many individual stretches of actin-containing filaments identified per bundle of kinetochore microtubules as there were microtubules which terminated at the kinetochore. These data suggest that actin-containing filaments in spindles have a functional role. We used 2 glycerination procedures in our studies (one for each cell), and neither seemed to disrupt the basic microtubule arrangements: the arrangements of spindle microtubules seen after glycerination of Haemanthus endosperm were identical to those described previously by others in non-glycerinated glutaraldehyde-fixed Haemanthus endosperm. Thus we argue that spindle structure is not disrupted by the procedures, and therefore that the arrangements of actin-containing filaments are not artifacts of the glycerination procedures. The only difference between microtubules in glycerinated cells and microtubules in untreated cells is that there seem to be fewer in the glycerinated cells. The possible role of actin-containing filaments in the spindle is discussed.

Actins↗
PubMed1979

Temperature effects on anaphase chromosome movement in the spermatocytes of two species of crane flies (Nephrotoma suturalis Loew and Nephrotoma ferruginea Fabricius).

Using phase-contrast cinemicrography on living crane fly (Nephrotoma suturalis Loew and Nephrotoma ferruginea Fabricius) spermatocytes, we have studied the effects of a range of temperatures (6--30 degrees C) on the anaphase I chromosome-to-pole movements of both autosomes and sex chromosomes. In contrast to previous work we have been able to study chromosome-to-pole velocities of autosomes without concurrent pole-to-pole elongation. In these cells we found that the higher the temperature, the faster was the autosomal chromosomes movement. From reviewing the literature we find that the general pattern of the effects of temperature on chromosome movement is similar whether or not pole-to-pole elongation occurs simultaneously with the chromosome-to-pole movement. Changes in cellular viscosities calculated from measurements of particulate Brownian movement do not seem to be able to account for the observed velocity differences due to temperature. Temperature effects on muscle contraction speed, flagellar beat frequency, ciliary beat frequency, granule flow in nerves, and chromosome movement have been compared, as have the activation energies for the rate-limiting steps in these motile systems: no distinction between possible mechanisms of force production is possible using these comparisons. The data show that even the different autosomes within single spermatocytes usually move at different speeds. These velocity differences cannot simply be related to chromosome size as the autosomes are visually indistinguishable. The sex chromosomes start their anaphase poleward movement after that of the autosomes, and move more slowly (by a factor of about 4), but their velocities appear to be affected by temperature in the same fashion as those of the autosomes. The interval between the onset of autosome anaphase and sex chromosome anaphase is also affected by temperature: the higher the temperature, the shorter the interval between the 2 stages. We have observed abnormalities in sex chromosome segregation, which may be due to temperature, but have not determined what the exact temperature shift conditions are that cause these abnormalities.

Anaphase↗
PubMed1978

Induction of cleavage in nucleated and enucleated frog eggs by injection of isolated sea-urchin mitotic apparatus.

Mitotic apparatus (MA) were isolated in glycerol-dimethylsulphoxide solution (MTME) from zygotes of sea urchins (Stronglyocentrotus purpuratus). Freshly isolated MA were stored in 1/10 strength MTME for varying periods of time and were then injected into unfertilized frog (Rana pipiens) eggs. These injections induced 40-60% of the recipient frog eggs to initiate cleavage, resulting in the formation of blastula cell clusters. The cleavage-inducing activity of MA stored in 1/10 MTME at room temperature decreased with time of storage in 1/10 strength MTME, and disappeared by about 6 h. There was no change in the ultrastructure of MA during storage. MA isolated and stored in MTME at room temperature had a constant level of cleavage-inducing activity during the first 48 h of storage, but this activity slowly declined upon further storage; almost no activity was left after 3 weeks. MA isolated in hexylene glycol (HG) and immediately transferred into MTME were compared with MA isolated in MTME; both MA had the same cleavage-inducing activity on the day of isolation, after which the MA isolated in HG quickly lost activity. On the other hand, MA isolated and stored in HG had little cleavage-inducing activity when tested 3 h following isolation. Cleavage-inducing agent (CIA) isolated from frog brains induced cleavage and blastula formation when injected into nucleated frog eggs, but had no such activity when injected into enucleated frog eggs. MA isolated in MTME induced cleavage and blastula formation in enucleated frog eggs as well as in nucleated frog eggs. Cytological examination revealed that blastula cells which developed from MA-injected enucleated eggs contained Feulgennegative nuclei, whereas cells which developed from CIA-injected nucleated eggs contained Feulgen-positive nuclei. These results suggest that sea-urchin nuclear materials participate in mitosis in frog eggs. Isolated MA which had been stored in MTME for 3 weeks and which exhibited little cleavage-inducing activity were injected together with frog brain CIA into either normal or enucleated eggs; normal recipient eggs cleaved with significantly higher frequencies (70%) than those injected with CIA alone (40%). Furthermore, enucleated eggs injected with CIA alone failed to cleave, while those injected with MA and CIA together cleaved with significant frequencies (overall 29%). This result suggests a cooperative interaction between CIA and the inactivated MA to restore the cleavage-inducing activity of MA.

Animals↗
PubMed1976

Spindle birefringence of isolated mitotic apparatus analysed by pressure treatment.

Sea-urchin zygote mitotic apparatus (MA) isolated in a glycerol/dimethylsulphoxide medium were treated with pressure. Pressure treatment had no effect on spindle birefringence when MA were in full-strength isolation medium. After placing MA in quarter-strength isolation medium, pressures of 4-0 X 10(3)-1-8 X 10(4) lbf in.-2 (2 X 76 X 10(4)-I X 24 X 10(5) k N m-2) for 15 min caused reduction of birefringence which occurred in 2 steps: firstly 20-30% of the birefringence was lost, and then, at higher pressures, the rest of the birefringence was lost. Electron microscopy suggested that pressure-induced changes were in non-microtubule material. Pressure treatment had no effect on MA isolated with hexylene glycol when the MA were pressurized in hexylene glycol; but pressure treatment did cause loss of birefringence when MA isolated in hexylene glycol were transferred immediately into glycerol/dimethylsulphoxide medium and were subsequently treated with pressure (after dilution into quarter-strength glycerol/dimethyl-sulphoxide). We discuss the differences in response between isolated MA and in vivo MA, and we discuss the possibility that 2 components contribute to MA birefringence.

Animals↗
PubMed1976

Spindle birefringence of isolated mitotic apparatus analysed by treatments with cold, pressure, and diluted isolation medium.

Mitotic apparatus (MA) were isolated from sea-urchin zygotes using glycerol-dimethyl-sulphoxide. Cold treatment had no effect on MA birefringence when MA were in isolation medium, but caused a 10-15% reduction of MA birefringence when MA were in quarter-strength isolation medium. Pressure treatment also caused a reduction in MA birefringence, but the cold and pressure treatments were not additive, suggesting that both treatments affected the same MA component. MA were not stable in quarter-strength isolation medium, and birefringence gradually decayed, with a half-life of about 40 h. Electron microscopy after cold treatment, or after decay of 55% of the MA birefringence showed abundant, normal-looking microtubules, suggesting that alterations in non-microtubule components cause the reductions in birefringence. Addition of EGTA eliminates the effect of cold treatment, suggesting that Ca2+ has a role in maintenance of spindle structure. We discuss possible reasons why isolated MA do not respond to cold treatment like MA in vivo.

Animals↗