scholarly journals Ultrastructural analysis of intracellular membrane and microtubule behavior during mitosis of Drosophila S2 cells

2017 ◽  
Author(s):  
Anton Strunov ◽  
Lidiya V. Boldyreva ◽  
Evgeniya N. Andreyeva ◽  
Gera A. Pavlova ◽  
Julia V. Popova ◽  
...  
Keyword(s):  
Ultrastructural Level ◽  
S2 Cells ◽  
Intracellular Membrane ◽  
Genetic Determinants ◽  
Microtubule Organization ◽  
Mitotic Apparatus ◽  
Transmission Electron ◽  
Chromosome Alignment ◽  
Drosophila S2 Cells ◽  
Cell Mitosis

AbstractS2 cells are one of the most widely used Drosophila melanogaster cell lines for molecular dissection of mitosis using RNA interference (RNAi). However, a detailed and complete description of S2 cell mitosis at the ultrastructural level is still missing. Here, we analyzed by transmission electron microscopy (TEM) a random sample of 144 cells undergoing mitosis, focusing on intracellular membrane and microtubule (MT) behavior. This unbiased approach allowed us to discover that S2 cells exhibit a characteristic behavior of intracellular membranes, involving the formation of a quadruple nuclear membrane in early prometaphase and its disassembly during late prometaphase. After nuclear envelope disassembly, the mitotic apparatus becomes encased by a discontinuous network of ER membranes that associate with mitochondria preventing their diffusion into the spindle area. We also observed a peculiar metaphase spindle organization. We found that kinetochores with attached k-fibers are almost invariably associated with lateral MT bundles that can be either interpolar bundles or k-fibers connected to a different kinetochore. This spindle organization is likely to favor chromosome alignment at metaphase and subsequent segregation during anaphase. In summary, we describe several previously unknown features of membrane and microtubule organization during S2 cell mitosis. The genetic determinants of these mitotic features of can now be investigated using an RNAi-based approach, which is particularly easy and efficient in S2 cells

scholarly journals Ultrastructural analysis of mitotic Drosophila S2 cells identifies distinctive microtubule and intracellular membrane behaviors

BMC Biology ◽  
2018 ◽  
Vol 16 (1) ◽  
Author(s):  
Anton Strunov ◽  
Lidiya V. Boldyreva ◽  
Evgeniya N. Andreyeva ◽  
Gera A. Pavlova ◽  
Julia V. Popova ◽  
...  
Keyword(s):  
Ultrastructural Analysis ◽  
S2 Cells ◽  
Intracellular Membrane ◽  
Drosophila S2 Cells

Centrosome Fine Ultrastructure of the Osteocyte Mechanosensitive Primary Cilium

2012 ◽  
Vol 18 (6) ◽  
pp. 1430-1441 ◽  
Author(s):  
R.E. Uzbekov ◽  
D.B. Maurel ◽  
P.C. Aveline ◽  
S. Pallu ◽  
C.L. Benhamou ◽  
...  
Keyword(s):  
Primary Cilium ◽  
Ultrastructural Level ◽  
Mutual Orientation ◽  
Positive Staining ◽  
Microtubule Organization ◽  
Alpha Tubulin ◽  
Transmission Electron ◽  
Mother And Daughter ◽  
Mother Centriole ◽  
Daughter Centriole

AbstractThe centrosome is the principal microtubule organization center in cells, giving rise to microtubule-based organelles (e.g., cilia, flagella). The aim was to study the osteocyte centrosome morphology at an ultrastructural level in relation to its mechanosensitive function. Osteocyte centrosomes and cilia in tibial cortical bone were explored by acetylated alpha-tubulin (AαTub) immunostaining under confocal microscopy. For the first time, fine ultrastructure and spatial orientation of the osteocyte centrosome were explored by transmission electron microscopy on serial ultrathin sections. AαTub-positive staining was observed in 94% of the osteocytes examined (222/236). The mother centriole formed a short primary cilium and was longer than the daughter centriole due to an intermediate zone between centriole and cilium. The proximal end of the mother centriole was connected with the surface of daughter centriole by striated rootlets. The mother centriole exhibited distal appendages that interacted with the cell membrane and formed a particular structure called “cilium membrane prolongation.” The primary cilium was mainly oriented perpendicular to the long axis of bone. Mother and daughter centrioles change their original mutual orientation during the osteocyte differentiation process. The short primary cilium is hypothesized as a novel type of fluid-sensing organelle in osteocytes.

Suppression of  -N-acetylglucosaminidase in the N-glycosylation pathway for complex glycoprotein formation in Drosophila S2 cells

Glycobiology ◽  
2008 ◽  
Vol 19 (3) ◽  
pp. 301-308 ◽  
Author(s):  
Y. K. Kim ◽  
K. R. Kim ◽  
D. G. Kang ◽  
S. Y. Jang ◽  
Y. H. Kim ◽  
...  
Keyword(s):  
S2 Cells ◽  
Drosophila S2 Cells ◽  
Glycosylation Pathway

scholarly journals Dissection of a Hypoxia-induced, Nitric Oxide–mediated Signaling Cascade

2009 ◽  
Vol 20 (18) ◽  
pp. 4083-4090 ◽  
Author(s):  
Pascale F. Dijkers ◽  
Patrick H. O'Farrell
Keyword(s):  
Nitric Oxide ◽  
Signal Transduction ◽  
Signaling Cascade ◽  
S2 Cells ◽  
Signal Transduction System ◽  
Signaling Systems ◽  
Calcium Dependent ◽  
Drosophila S2 Cells ◽  
Oxide Synthase ◽  
Multiple Signaling

Befitting oxygen's key role in life's processes, hypoxia engages multiple signaling systems that evoke pervasive adaptations. Using surrogate genetics in a powerful biological model, we dissect a poorly understood hypoxia-sensing and signal transduction system. Hypoxia triggers NO-dependent accumulation of cyclic GMP and translocation of cytoplasmic GFP-Relish (an NFκB/Rel transcription factor) to the nucleus in Drosophila S2 cells. An enzyme capable of eliminating NO interrupted signaling specifically when it was targeted to the mitochondria, arguing for a mitochondrial NO signal. Long pretreatment with an inhibitor of nitric oxide synthase (NOS), L-NAME, blocked signaling. However, addition shortly before hypoxia was without effect, suggesting that signaling is supported by the prior action of NOS and is independent of NOS action during hypoxia. We implicated the glutathione adduct, GSNO, as a signaling mediator by showing that overexpression of the cytoplasmic enzyme catalyzing its destruction, GSNOR, blocks signaling, whereas knockdown of this activity caused reporter translocation in the absence of hypoxia. In downstream steps, cGMP accumulated, and calcium-dependent signaling was subsequently activated via cGMP-dependent channels. These findings reveal the use of unconventional steps in an NO pathway involved in sensing hypoxia and initiating signaling.

scholarly journals Drosophila kinesin-8 stabilizes the kinetochore–microtubule interaction

2018 ◽  
Vol 218 (2) ◽  
pp. 474-488 ◽  
Author(s):  
Tomoya Edzuka ◽  
Gohta Goshima
Keyword(s):  
Yeast Cell ◽  
Ectopic Expression ◽  
Cell Types ◽  
Binding Activity ◽  
S2 Cells ◽  
Tubulin Binding ◽  
Chromosome Alignment ◽  
Normal Frequency ◽  
Directed Motility

Kinesin-8 is required for proper chromosome alignment in a variety of animal and yeast cell types. However, it is unclear how this motor protein family controls chromosome alignment, as multiple biochemical activities, including inconsistent ones between studies, have been identified. Here, we find that Drosophila kinesin-8 (Klp67A) possesses both microtubule (MT) plus end–stabilizing and –destabilizing activity, in addition to kinesin-8's commonly observed MT plus end–directed motility and tubulin-binding activity in vitro. We further show that Klp67A is required for stable kinetochore–MT attachment during prometaphase in S2 cells. In the absence of Klp67A, abnormally long MTs interact in an “end-on” fashion with kinetochores at normal frequency. However, the interaction is unstable, and MTs frequently become detached. This phenotype is rescued by ectopic expression of the MT plus end–stabilizing factor CLASP, but not by artificial shortening of MTs. We show that human kinesin-8 (KIF18A) is also important to ensure proper MT attachment. Overall, these results suggest that the MT-stabilizing activity of kinesin-8 is critical for stable kinetochore–MT attachment.

scholarly journals Evidence for a rapid cold hardening response in cultured Drosophila S2 cells

2019 ◽  
Vol 223 (2) ◽  
pp. jeb212613 ◽  
Author(s):  
Emily A. W. Nadeau ◽  
Nicholas M. Teets
Keyword(s):  
Cold Hardening ◽  
S2 Cells ◽  
Rapid Cold Hardening ◽  
Drosophila S2 Cells
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