The fine structure of the kinetochore of a mammalian cell in vitro

The chromosomes of Chinese hamster cells were examined with the electron microscope and the following observations were made concerning the structure and organization of the kinetochore. — The kinetochore consists of a dense core 200–300 Å in diameter surrounded hy a less dense zone 200–600 Å wide. The dense core consists of a pair of axial fibrils 50–80 Å in diameter which may be coiled together in a cohelical manner. The less dense zone about the axial elements is composed of numerous microfibrils which loop out at right angles to the axial fibrils. Together the structures comprise a lampbrush-like filament which extends along the surface of each chromatid. Some sections suggested that two such filaments may be present on each chromatid. The fine structure of kinetochores associated with spindle filaments was essentially the same as those free of filaments. The structure and organization of the kinetochore of these mammalian cells was compared to that of lampbrush chromosomes of certain amphibian oöcytes, dipteran polytene chromosome puffs, and the synaptinemal complex seen during meiotic prophase.The authors also wish to thank Dr. Arthur Cole of the Department of Physics for the use of his electron microscope facilities and for his helpful criticism.     

Centrophilin: a novel mitotic spindle protein involved in microtubule nucleation

A novel protein has been identified which may serve a key function in nucleating spindle microtubule growth in mitosis. This protein, called centrophilin, is sequentially relocated from the centromeres to the centrosomes to the midbody in a manner dependent on the mitotic phase. Centrophilin was initially detected by immunofluorescence with a monoclonal, primate-specific antibody (2D3) raised against kinetochore-enriched chromosome extract from HeLa cells (Valdivia, M. M., and B. R. Brinkley. 1985. J. Cell Biol. 101:1124-1134). Centrophilin forms prominent crescents at the poles of the metaphase spindle, gradually diminishes during anaphase, and bands the equatorial ends of midbody microtubules in telophase. The formation and breakdown of the spindle and midbody correlates in time and space with the aggregation and disaggregation of centrophilin foci. Immunogold EM reveals that centrophilin is a major component of pericentriolar material in metaphase. During recovery from microtubule inhibition, centrophilin foci act as nucleation sites for the assembly of spindle tubules. The 2D3 probe recognizes two high molecular mass polypeptides, 180 and 210 kD, on immunoblots of whole HeLa cell extract. Taken together, these data and the available literature on microtubule dynamics point inevitably to a singular model for control of spindle tubule turnover.     

Role of PII proteins in nitrogen fixation control of Herbaspirillum seropedicae strain SmR1

Background  

The PII protein family comprises homotrimeric proteins which act as transducers of the cellular nitrogen and carbon status in prokaryotes and plants. In Herbaspirillum seropedicae, two PII-like proteins (GlnB and GlnK), encoded by the genes glnB and glnK, were identified. The glnB gene is monocistronic and its expression is constitutive, while glnK is located in the nlmAglnKamtB operon and is expressed under nitrogen-limiting conditions.     

Plasma gonadotrophins and progesterone concentrations during various degrees of underfeeding in pregnant mice

Daily reduction of the normal (ad libitum) food consumption by as little as 35% significantly reduced (P less than 0.05) the percentage of mice with implantation sites at Days 7 and 9 of gestation. Underfeeding decreased body weight and reduced the weight of the ovaries and uterus. Plasma progesterone was decreased (P less than 0.05) as dietary intake was restricted and was associated with regression of the corpora lutea. No significant alterations in the plasma values of LH and FSH were observed in mice underfed between Days 1 and 9 of pregnancy. The decrease in plasma progesterone in the absence of reduced LH values may indicate that progesterone secretion between Days 5 and 9 of gestation is not controlled solely by LH.     

Cytoplasmic microtubules in transformed mouse x nontransformed human cell hybrids: correlation with in vitro growth.

Patterns of cytoplasmic microtubules in somatic cell hybrids between transformed mouse cells and nontransformed human skin fibroblasts were examined using antitubulin antibodies as an immunofluorescent probe. Nontransformed cells have been shown to exhibit an extensive cytoplasmic microtubule complex (CMTC), while in transformed cells, this complex is greatly diminished. The hybrid populations contained both types of cells. In addition, they contained cells with previously undescribed intermediate CMTC phenotypes. The percentage of each phenotype present in hybrid populations was determined for sixteen hybrid clones. Seven clones were found which appeared transformed on the basis of their CMTC pattern. The others were comprised of various proportions of all the cell types described. Repeated quantitation of the proportions of these types in the hybrid populations showed them to be stable with time in culture. Growth in vitro of the hybrid clones was assayed by determining their saturation densities, their plating efficiencies on plastic and their colony-forming abilities in soft agar. In vitro growth of a cell population was found to be directly proportional to the percentage of cells in the population which showed the greatly diminished CMTC pattern which has been described for transformed cells. This is strong evidence that a greatly reduced CMTC is associated with transformed behavior, especially the increased capacity of transformed cells for in vitro growth.     

An improved culture system for secondary palatal elevation

Summary An organ culture system devised for studying the development of the secondary palate was modified so that it retained high partial pressures of oxygen and supported total anterior and posterior palatal elevation. The cultured tissues appeared healthy as judged by histological examination. Medium was continuously recirculated through the culture system in which Day 13 embryonic mouse heads, with the brain and tongue removed, were totally submerged and suspended. The medium was constantly gassed via hollow fiber devices. A motor-driven stirrer, run at a low rate, agitated the medium so that the boundary layer surrounding the tissue was dispersed. Embryonic mouse heads were cultured in each of four media: Eagle's basal medium, Ham's F-12 medium, Fitton-Jackson's modified BGJb medium, and Waymouth's MB 752/1 medium. Elevation of the palate in both anterior and posterior regions with excellent tissue viability was achieved in all heads grown in BGJb medium. This work was supported by NIH Post-doctoral Fellowship 2F32 DE05038-03 to C. A. L.