Astral microtubules (MTs) emanating from the mitotic apparatus (MA) during anaphase are required for stimulation of cytokinesis in eggs. We have used green fluorescent protein-labeled EB1 to observe MT dynamics during mitosis and cytokinesis in normal sea urchin eggs. Analysis of astral MT growth rates during anaphase shows that MTs contact the polar cortex earlier than the equatorial cortex after anaphase onset but that a normal cleavage furrow is not induced until contact with MTs has been achieved throughout the cortex. To assess the role of MT dynamics in initiation of cytokinesis, we used a collection of small molecule drugs to affect dynamics. Hexylene glycol resulted in rapid astral elongation due to decreased MT catastrophe and precocious furrowing. Taxol suppressed MT dynamics but did not inhibit furrow induction when the MA was manipulated toward the cortex. Urethane resulted in short, highly dynamic astral MTs with increased catastrophe that also stimulated furrowing upon being brought into proximity to the cortex. Our findings indicate that astral MT contact with the cortex is necessary for furrow initiation but that the dynamic state of astral MTs does not affect their competency to stimulate furrowing. INTRODUCTIONCytokinesis is the process by which the cytoplasm is divided at the end of mitosis to form two daughter cells. Much has been learned about the actomyosin ring that generates the contractile force required for cytokinesis; however, many questions remain regarding mechanisms that specify the place and time for contractile ring assembly and activation. The cleavage furrow normally bisects the mitotic apparatus (MA) at the plane of the former metaphase plate (Wilson, 1928), and microtubules (MTs) of the MA are essential for communication of the contractile signals (Beams and Evans, 1940). Several models have been proposed to describe the initiation of cytokinesis, each of which attributes the source of the signal to a different MT population within the MA (Balasubramanian et al., 2004;Burgess and Chang, 2005). In one of the most well studied cell types, the cleaving echinoderm egg, positioning of the furrow is determined by equatorial stimulation from two asters and not by the bundled MTs of the spindle midzone, kinetochores, or by the polar astral MTs (Rappaport, 1996), which may play a stimulatory role in other cell types (Burgess and Chang, 2005;D'Avino et al., 2005;Glotzer, 2005). However, the nature, degree, and extent of contact between the astral MTs and the cortex required for stimulation of cytokinesis is not yet determined.Changes in MT dynamics facilitate structural reorganization of the MA during cell division. At the metaphaseanaphase transition astral MTs elongate due to decreased catastrophe or increased rescue, kinetochore MTs shorten to segregate the chromosomes, and spindle midzone MTs grow to promote separation of the spindle poles (DeBrabander et al., 1986;Snyder and Mullins, 1993;Zhai et al., 1995;Mallavarapu et al., 1999;Rusan et al., 2002;Piehl et al., 2004). Anaphase o...
Neonates are more susceptible than adults to viral and bacterial diseases. We hypothesized that plasmacytoid dendritic cells, the cells that provide large amounts of IFN-α in response to Toll-like receptor 9 (TLR9) agonists, are defective in neonates. To assess the intrinsic functionality of plasmacytoid dendritic cells from neonates we compared IFN-α production by plasmacytoid dendritic cells derived from neonates versus adults in both whole blood and in purified plasmacytoid dendritic cells. TLR9-stimulation of whole blood from adults and neonates resulted in comparable amounts of IFN-α production. However, we observed small but significant differences in IFN-α production from purified CD123+ plasmacytoid dendritic cells from neonates after stimulation with the TLR9 ligand CpG-DNA. Furthermore, we assessed surface expression of co-stimulatory molecules on plasmacytoid dendritic cells after stimulation. While purified CD123+ plasmacytoid dendritic cells from adults up-regulated co-stimulatory molecules CD80 and CD86 with IL-3 alone those from neonates required the addition of CpG-DNA to reach adult levels. Therefore, the intrinsic deficiencies of neonatal plasmacytoid dendritic cells can be mitigated by TLR9 agonists. These results are consistent with the observation that vaccines that effect strong adjuvant activity on dendritic cells can induce protective responses in neonates.Neonates are more susceptible than adults to severe disease following exposure to viruses and bacteria (1). A lack of pre-existing memory T-and B-cells as well as diminished T-cell responses (2), reduced numbers of dendritic cells (DC), and a bias toward Th2 immunity (3) are contributors, but likely don't entirely explain this observation. One hypothesis is that neonatal DC are functionally immature. One manifestation of such immaturity may be that neonatal DC respond suboptimally to pathogens via Toll-like receptor (TLR) agonist stimulation and therefore are lacking the ability to provide a link between innate and adaptive immune responses (4). Unlike myeloid DC, plasmacytoid DC (pDC) express high levels of the pathogen-recognizing receptor TLR9 (4). Once stimulated with TLR9 agonists such as viral (5,6) as well as bacterial DNA (7,8) pDC produce large amounts of IFN-α. In addition, pDC can stimulate allogeneic T-cells and pDC exposed to virus are capable of directly priming T-cells in mice (9). Furthermore, influenza-exposed pDC can stimulate antigen-experienced influenza-specific CD4 and CD8 T-cell clones to produce IFN-γ suggesting a role for pDC in stimulation of memory T-cell responses (10).We hypothesized that DC from the plasmacytoid subset in human neonates were functionally defective. Previous studies of DC either in vivo or using mixed cell populations that contained nonDC (11,12) showed that neonatal DC may be defective while more recent studies using purified populations (13,14) defects. Therefore, to distinguish intrinsic deficiencies from extrinsic factors we studied responses in whole blood and in highly purifie...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
customersupport@researchsolutions.com
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.