guanosine-triphosphate and 5-((4-6-dichloro-1-3-5-triazin-2-yl)amino)fluorescein

guanosine-triphosphate has been researched along with 5-((4-6-dichloro-1-3-5-triazin-2-yl)amino)fluorescein* in 1 studies

Other Studies

1 other study(ies) available for guanosine-triphosphate and 5-((4-6-dichloro-1-3-5-triazin-2-yl)amino)fluorescein

Article	Year
Incorporation and turnover of labeled exogenous tubulin in the mitotic spindles of Chaetopterus oocytes and HeLa cells. Cell motility and the cytoskeleton, 1986, Volume: 6, Issue:2 The incorporation of tubulin into mitotic spindles in situ was studied by incubating permeabilized mitotic cells in solutions containing [3H]GTP-labeled or dichlorotriazinylamino fluorescein (DTAF)-labeled tubulin. Metaphase HeLa cells or spindle-containing "minicells" from Chaetopterus oocytes were lysed in a microtubule-assembly buffer plus 0.5% Nonidet P-40, 1 mg/ml 120,000g supernatant mammalian brain tubulin, and [3H]GTP. After different periods of incubation, mitotic spindles were isolated in 2 M-glycerol-containing assembly buffer and separated from unbound counts by centrifugation through a 4 M-glycerol cushion; 3H counts per mg protein increase linearly for 8-12 min and then reach a plateau or steady state in both Chaetopterus oocytes and HeLa cells. Addition of 4 mM CaCl2 blocks or reverses incorporation. Little or no [3H]GTP is incorporated if exogenous tubulin or lysed cells are omitted from the assembly mixture. To measure the loss rate of [3H]GTP-tubulin from mitotic spindles, cells were incubated in tubulin plus [3H]GTP for 30 min, and a 20-fold excess of cold GTP (2 mM) was added. Samples were removed after incubation for different periods, and spindles were isolated as described above and counted for 3H content. [3H]GTP is lost from spindles at a rate of about 16%/min until a new steady state is reached in about 8 min. These results are consistent with an incorporation and turnover of [3H]GTP-tubulin in spindle microtubules of these lysed-cell models. The location of this newly incorporated tubulin in the spindle was investigated by incorporating fluorescent DTAF-tubulin into mitotic spindles of these lysed cell types.(ABSTRACT TRUNCATED AT 250 WORDS) Topics: Animals; Annelida; Cell Membrane Permeability; Detergents; Fluoresceins; Fluorescent Dyes; Guanosine Diphosphate; Guanosine Triphosphate; HeLa Cells; Humans; Kinetics; Microscopy, Electron; Microtubules; Octoxynol; Oocytes; Polyethylene Glycols; Spindle Apparatus; Tubulin	1986

Article

Year

Incorporation and turnover of labeled exogenous tubulin in the mitotic spindles of Chaetopterus oocytes and HeLa cells.

Cell motility and the cytoskeleton, 1986, Volume: 6, Issue:2

The incorporation of tubulin into mitotic spindles in situ was studied by incubating permeabilized mitotic cells in solutions containing [3H]GTP-labeled or dichlorotriazinylamino fluorescein (DTAF)-labeled tubulin. Metaphase HeLa cells or spindle-containing "minicells" from Chaetopterus oocytes were lysed in a microtubule-assembly buffer plus 0.5% Nonidet P-40, 1 mg/ml 120,000g supernatant mammalian brain tubulin, and [3H]GTP. After different periods of incubation, mitotic spindles were isolated in 2 M-glycerol-containing assembly buffer and separated from unbound counts by centrifugation through a 4 M-glycerol cushion; 3H counts per mg protein increase linearly for 8-12 min and then reach a plateau or steady state in both Chaetopterus oocytes and HeLa cells. Addition of 4 mM CaCl2 blocks or reverses incorporation. Little or no [3H]GTP is incorporated if exogenous tubulin or lysed cells are omitted from the assembly mixture. To measure the loss rate of [3H]GTP-tubulin from mitotic spindles, cells were incubated in tubulin plus [3H]GTP for 30 min, and a 20-fold excess of cold GTP (2 mM) was added. Samples were removed after incubation for different periods, and spindles were isolated as described above and counted for 3H content. [3H]GTP is lost from spindles at a rate of about 16%/min until a new steady state is reached in about 8 min. These results are consistent with an incorporation and turnover of [3H]GTP-tubulin in spindle microtubules of these lysed-cell models. The location of this newly incorporated tubulin in the spindle was investigated by incorporating fluorescent DTAF-tubulin into mitotic spindles of these lysed cell types.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Annelida; Cell Membrane Permeability; Detergents; Fluoresceins; Fluorescent Dyes; Guanosine Diphosphate; Guanosine Triphosphate; HeLa Cells; Humans; Kinetics; Microscopy, Electron; Microtubules; Octoxynol; Oocytes; Polyethylene Glycols; Spindle Apparatus; Tubulin

1986