Tomographic reconstruction of treponemal cytoplasmic filaments reveals novel bridging and anchoring components

An understanding of the involvement of bacterial cytoplasmic filaments in cell division requires the elucidation of the structural organization of those filamentous structures. Treponemal cytoplasmic filaments are composed of one protein, CfpA, and have been demonstrated to be involved in cell division. In this study, we used electron tomography to show that the filaments are part of a complex with a novel molecular organization that includes at least two distinct features decorating the filaments. One set of components appears to anchor the filaments to the cytoplasmic membrane. The other set of components appears to bridge the cytoplasmic filaments on the cytoplasmic side, and to be involved in the interfilament spacing within the cell. The filaments occupy between 3 and 18% of the inner surface of the cytoplasmic membrane. These results reveal a novel filamentous molecular organization of independent filaments linked by bridges and continuously anchored to the membrane

The de novo centriole assembly pathway in HeLa cells: cell cycle progression and centriole assembly/maturation

It has been reported that nontransformed mammalian cells become arrested during G1 in the absence of centrioles (Hinchcliffe, E., F. Miller, M. Cham, A. Khodjakov, and G. Sluder. 2001. Science. 291:1547–1550). Here, we show that removal of resident centrioles (by laser ablation or needle microsurgery) does not impede cell cycle progression in HeLa cells. HeLa cells born without centrosomes, later, assemble a variable number of centrioles de novo. Centriole assembly begins with the formation of small centrin aggregates that appear during the S phase. These, initially amorphous “precentrioles” become morphologically recognizable centrioles before mitosis. De novo–assembled centrioles mature (i.e., gain abilities to organize microtubules and replicate) in the next cell cycle. This maturation is not simply a time-dependent phenomenon, because de novo–formed centrioles do not mature if they are assembled in S phase–arrested cells. By selectively ablating only one centriole at a time, we find that the presence of a single centriole inhibits the assembly of additional centrioles, indicating that centrioles have an activity that suppresses the de novo pathway

Kinetochores Use a Novel Mechanism for Coordinating the Dynamics of Individual Microtubules

SummaryChromosome alignment during mitosis is frequently accompanied by a dynamic switching between elongation and shortening of kinetochore fibers (K-fibers) that connect kinetochores and spindle poles [1, 2]. In higher eukaryotes, mature K-fibers consist of 10–30 kinetochore microtubules (kMTs) whose plus ends are embedded in the kinetochore [1–3]. A critical and long-standing question is how the dynamics of individual kMTs within the K-fiber are coordinated [1–5]. We have addressed this question by using electron tomography to determine the polymerization/depolymerization status of individual kMTs in the K-fibers of PtK1 and Drosophila S2 cells. Surprisingly, we find that the plus ends of two-thirds of kMTs are in a depolymerizing state, even when the K-fiber exhibits net tubulin incorporation at the plus end [6–8]. Furthermore, almost all individual K-fibers examined had a mixture of kMTs in the polymerizing and depolymerizing states. Therefore, although K-fibers elongate and shrink as a unit, the dynamics of individual kMTs within a K-fiber are not coordinated at any given moment. Our results suggest a novel control mechanism through which attachment to the kinetochore outer plate prevents shrinkage of kMTs. We discuss the ramifications of this new model on the regulation of chromosome movement and the stability of K-fibers

Tomographic reconstruction of treponemal cytoplasmic filaments reveals novel bridging and anchoring components

An understanding of the involvement of bacterial cytoplasmic filaments in cell division requires the elucidation of the structural organization of those filamentous structures. Treponemal cytoplasmic filaments are composed of one protein, CfpA, and have been demonstrated to be involved in cell division. In this study, we used electron tomography to show that the filaments are part of a complex with a novel molecular organization that includes at least two distinct features decorating the filaments. One set of components appears to anchor the filaments to the cytoplasmic membrane. The other set of components appears to bridge the cytoplasmic filaments on the cytoplasmic side, and to be involved in the interfilament spacing within the cell. The filaments occupy between 3 and 18% of the inner surface of the cytoplasmic membrane. These results reveal a novel filamentous molecular organization of independent filaments linked by bridges and continuously anchored to the membrane

Releasing the spindle assembly checkpoint without tension

Eukaryotic cells have evolved a spindle assembly checkpoint (SAC) that facilitates accurate genomic segregation during mitosis by delaying anaphase onset in response to errors in kinetochore microtubule attachment. In contrast to the well-studied molecular mechanism by which the SAC blocks anaphase onset, the events triggering SAC release are poorly understood. Papers in this issue by Uchida et al. (Uchida, K.S.K., K. Takagaki, K. Kumada, Y. Hirayama, T. Noda, and T. Hirota. 2009. J. Cell Biol. 184:383–390) and Maresca and Salmon (Maresca, T.J., and E.D. Salmon. 2009. J. Cell Biol. 184:373–381) make an important advance by demonstrating that SAC release depends on molecular rearrangements within the kinetochore rather than tension-produced stretch between sister kinetochores

The M\u3csub\u3er\u3c/sub\u3e-value of chloroplast coupling factor 1

The Mr of spinach chloroplast coupling factor 1 has been determined by sedimentation equilibrium and by light scattering to be 400 000 ± 24 600 and 407 000 ± 20 000, respectively. These values differ substantially from that obtained previously (325 000) and are consistent with an α3β3γδε{lunate} subunit stoichiometry. © 1983

Regulation of Kainate Receptor Subunit mRNA by Stress and Corticosteroids in the Rat Hippocampus

Kainate receptors are a class of ionotropic glutamate receptors that have a role in the modulation of glutamate release and synaptic plasticity in the hippocampal formation. Previous studies have implicated corticosteroids in the regulation of these receptors and recent clinical work has shown that polymorphisms in kainate receptor subunit genes are associated with susceptibility to major depression and response to anti-depressant treatment. In the present study we sought to examine the effects of chronic stress and corticosteroid treatments upon the expression of the mRNA of kainate receptor subunits GluR5-7 and KA1-2. Our results show that, after 7 days, adrenalectomy results in increased expression of hippocampal KA1, GluR6 and GluR7 mRNAs, an effect which is reversed by treatment with corticosterone in the case of KA1 and GluR7 and by aldosterone treatment in the case of GluR6. 21 days of chronic restraint stress (CRS) elevated the expression of the KA1 subunit, but had no effect on the expression of the other subunits. Similarly, 21 days of treatment with a moderate dose of corticosterone also increased KA1 mRNA in the dentate gyrus, whereas a high corticosterone dose has no effect. Our results suggest an interaction between hippocampal kainate receptor composition and the hypothalamic-pituitary-adrenal (HPA) axis and show a selective chronic stress induced modulation of the KA1 subunit in the dentate gyrus and CA3 that has implications for stress-induced adaptive structural plasticity