Interaction of an Overexpressed gamma-Tubulin with Microtubules In Vivo and In Vitro.

gamma-Tubulin is an ubiquitous MTOC (microtubule-organizing center) component essential for the regulation of microtubule functions. A 1.8 kb cDNA coding for gamma-tubulin was isolated from CHO cells. Analysis of nucleotide sequence predicts a protein of 451 amino acids, which is over 97% identical to human and Xenopus gamma-tubulin. When CHO cells were transiently transfected with the gamma-tubulin clone, epitope-tagged full-length, as well as truncated polypeptides (amino acids 1-398 and 1-340), resulted in the formation of cytoplasmic foci of various sizes. Although one of the foci was identified as the centrosome, the rest of the dots were not associated with any other centrosomal components tested so far. The pattern of microtubule organization was not affected by induction of such gamma-tubulin-containing dots in transfected cells. In addition, the cytoplasmic foci were unable to serve as the site for microtubule regrowth in nocodazole-treated cells upon removal of the drug, suggesting that gamma-tubulin-containing foci were not involved in the activity for microtubule formation and organization. Using the monomeric form of Chlamydomonas gamma-tubulin purified from insect Sf9 cells (), interaction between gamma-tubulin and microtubules was further investigated by immunoelectron microscopy. Microtubules incubated with gamma-tubulin monomers in vitro were associated with more gold particles conjugated with gamma-tubulin than in controls where no exogenous gamma-tubulin was added. However, binding of gamma-tubulin to microtubules was not extensive and was easily lost during sample preparation. Although gamma-tubulin was detected at the minus end of microtubules several times more frequently than the plus end, the majority of gold particles were seen along the microtubule length. These results contradict the previous reports (; ), which might be ascribed to the difference in the level of protein expression in transfected cells.

[The role of the cytoskeletal elements during the return to the cell center of their nuclei displaced by centrifugation]. / Rol' élementov tsitoskeleta v protsesse vozvrashcheniia v tsentr kletok ikh iader, smeshchennykh tsentrifugirovaniem.

Living cells of the monolayer cultures of embryonal pig kidney epithelium (PKE-cells) and of embryonal bovine tracheal cells (FBT-cells) were ultracentrifuged at 20,000g. The centrifugal force was directed parallel to the surface of the culture slides. Just after centrifugation, the cellular nuclei were displaced to the centrifugal parts of cells. Centrifuged slides with cells were returned to the normal culture conditions, and 22 h later the nuclei were seen to restore their central position in the cells. The motion of the nuclei to the cell center was rather chaotic both in direction and speed. The speed of this motion never exceeded several microns per hour. After nocodasole treatment (0.1-10 mkg/ml) of the cells or in a hypotonic medium, the distance of nuclear dislocation during centrifugation was longer, and the nuclei returned to the cell centers faster than in the control ones. After cytochalasin B treatment (2 mkg/ml), the nuclei moved to the cell centers somewhat more slowly than they did in the control cells. Thus, the establishment of the central position of nuclei in the cells takes place in the absence of microtubules or intermediate filaments. Probably, the central position of nuclei depends mainly on the action part of the cytoskeleton.

Diagnosis of rheumatism by means of universal criteria, obtained by the cybernetic method.

With a view to establishing standardized criteria for the diagnosis of rheumatism suitable for widespread use in polyclinics and in large-scale epidemiological investigations, 3000 adults and children with rheumatism or one of seven similar diseases were subjected to multiple screening in eight centres in the USSR using a specially prepared standard card and standard methods. The information obtained was processed by cybernetic methods. Following determination of the sensitivity, specificity, and informational value of 486 signs of rheumatism, 68 of the most indicative signs were selected as criteria of rheumatism. These were included in the final table and weighted according to their indicational value both individually and in groups as syndromes. Using the table it is possible by simple calculation, without using a computer, to arrive at an objective evaluation of the reliability of the clinical diagnosis of rheumatism in patients with one of a group of eight similar diseases. The final diagnostic table was tested on 562 patients selected epidemiologically or from polyclinics or clinics and was shown to result in a high degree of diagnostic precision (higher than 95%).