Combined Pulldown and Time-Lapse Microscopy Studies for Determining the Role of Rap1 in the Crosstalk Between Integrins and Cadherins

2013 ◽  
pp. 177-195 ◽  
Author(s):  
Luca Goitre ◽  
Saverio Francesco Retta
Keyword(s):  
Time Lapse ◽  
Time Lapse Microscopy

scholarly journals SpoIIB Localizes to Active Sites of Septal Biogenesis and Spatially Regulates Septal Thinning during Engulfment in Bacillus subtilis

2000 ◽  
Vol 182 (4) ◽  
pp. 1096-1108 ◽  
Author(s):  
Ana R. Perez ◽  
Angelica Abanes-De Mello ◽  
Kit Pogliano
Keyword(s):  
Bacillus Subtilis ◽  
Fusion Protein ◽  
Mother Cell ◽  
Active Sites ◽  
Time Lapse ◽  
Formation Pathway ◽  
Time Lapse Microscopy ◽  
Division Site ◽  
Proposal A

ABSTRACT A key step in the Bacillus subtilis spore formation pathway is the engulfment of the forespore by the mother cell, a phagocytosis-like process normally accompanied by the loss of peptidoglycan within the sporulation septum. We have reinvestigated the role of SpoIIB in engulfment by using the fluorescent membrane stain FM 4-64 and deconvolution microscopy. We have found thatspoIIB mutant sporangia display a transient engulfment defect in which the forespore pushes through the septum and bulges into the mother cell, similar to the situation in spoIID,spoIIM, and spoIIP mutants. However, unlike the sporangia of those three mutants, spoIIB mutant sporangia are able to complete engulfment; indeed, by time-lapse microscopy, sporangia with prominent bulges were found to complete engulfment. Electron micrographs showed that in spoIIB mutant sporangia the dissolution of septal peptidoglycan is delayed and spatially unregulated and that the engulfing membranes migrate around the remaining septal peptidoglycan. These results demonstrate that mother cell membranes will move around septal peptidoglycan that has not been completely degraded and suggest that SpoIIB facilitates the rapid and spatially regulated dissolution of septal peptidoglycan. In keeping with this proposal, a SpoIIB-myc fusion protein localized to the sporulation septum during its biogenesis, discriminating between the site of active septal biogenesis and the unused potential division site within the same cell.

scholarly journals Opposing actions of septins and Sticky on Anillin promote the transition from contractile to midbody ring

2013 ◽  
Vol 203 (3) ◽  
pp. 487-504 ◽  
Author(s):  
Nour El Amine ◽  
Amel Kechad ◽  
Silvana Jananji ◽  
Gilles R.X. Hickson
Keyword(s):  
Essential Role ◽  
Time Lapse ◽  
Maturation Process ◽  
S2 Cells ◽  
Contractile Ring ◽  
Time Lapse Microscopy ◽  
Drosophila Melanogaster S2 Cells ◽  
C Terminus ◽  
N Terminus

During cytokinesis, closure of the actomyosin contractile ring (CR) is coupled to the formation of a midbody ring (MR), through poorly understood mechanisms. Using time-lapse microscopy of Drosophila melanogaster S2 cells, we show that the transition from the CR to the MR proceeds via a previously uncharacterized maturation process that requires opposing mechanisms of removal and retention of the scaffold protein Anillin. The septin cytoskeleton acts on the C terminus of Anillin to locally trim away excess membrane from the late CR/nascent MR via internalization, extrusion, and shedding, whereas the citron kinase Sticky acts on the N terminus of Anillin to retain it at the mature MR. Simultaneous depletion of septins and Sticky not only disrupted MR formation but also caused earlier CR oscillations, uncovering redundant mechanisms of CR stability that can partly explain the essential role of Anillin in this process. Our findings highlight the relatedness of the CR and MR and suggest that membrane removal is coordinated with CR disassembly.

scholarly journals Distinct roles of septins in cytokinesis: SEPT9 mediates midbody abscission

2010 ◽  
Vol 191 (4) ◽  
pp. 741-749 ◽  
Author(s):  
Mathew P. Estey ◽  
Caterina Di Ciano-Oliveira ◽  
Carol D. Froese ◽  
Margaret T. Bejide ◽  
William S. Trimble
Keyword(s):  
Cell Division ◽  
Time Lapse ◽  
Vesicle Tethering ◽  
Daughter Cells ◽  
Time Lapse Microscopy ◽  
Early Stages ◽  
Mechanistic Insight ◽  
The Family ◽  
Insight Into

Septins are a family of GTP-binding proteins implicated in mammalian cell division. Most studies examining the role of septins in this process have treated the family as a whole, thus neglecting the possibility that individual members may have diverse functions. To address this, we individually depleted each septin family member expressed in HeLa cells by siRNA and assayed for defects in cell division by immunofluorescence and time-lapse microscopy. Depletion of SEPT2, SEPT7, and SEPT11 causes defects in the early stages of cytokinesis, ultimately resulting in binucleation. In sharp contrast, SEPT9 is dispensable for the early stages of cell division, but is critical for the final separation of daughter cells. Rescue experiments indicate that SEPT9 isoforms containing the N-terminal region are sufficient to drive cytokinesis. We demonstrate that SEPT9 mediates the localization of the vesicle-tethering exocyst complex to the midbody, providing mechanistic insight into the role of SEPT9 during abscission.

Experience of using time-lapse microscopy in IVF and ICSI programs

2020 ◽  
pp. 47-50
Author(s):  
N. V. Saraeva ◽  
N. V. Spiridonova ◽  
M. T. Tugushev ◽  
O. V. Shurygina ◽  
A. I. Sinitsyna
Keyword(s):  
Pregnancy Rate ◽  
Embryo Transfer ◽  
Selection Procedure ◽  
Time Lapse ◽  
Single Embryo Transfer ◽  
Main Group ◽  
Clinical Pregnancy ◽  
Control Group ◽  
Time Lapse Microscopy

In order to increase the pregnancy rate in the assisted reproductive technology, the selection of one embryo with the highest implantation potential it is very important. Time-lapse microscopy (TLM) is a tool for selecting quality embryos for transfer. This study aimed to assess the benefits of single-embryo transfer of autologous oocytes performed on day 5 of embryo incubation in a TLM-equipped system in IVF and ICSI programs. Single-embryo transfer following incubation in a TLM-equipped incubator was performed in 282 patients, who formed the main group; the control group consisted of 461 patients undergoing single-embryo transfer following a traditional culture and embryo selection procedure. We assessed the quality of transferred embryos, the rates of clinical pregnancy and delivery. The groups did not differ in the ratio of IVF and ICSI cycles, average age, and infertility factor. The proportion of excellent quality embryos for transfer was 77.0% in the main group and 65.1% in the control group (p = 0.001). In the subgroup with receiving eight and less oocytes we noted the tendency of receiving more quality embryos in the main group (р = 0.052). In the subgroup of nine and more oocytes the quality of the transferred embryos did not differ between two groups. The clinical pregnancy rate was 60.2% in the main group and 52.9% in the control group (p = 0.057). The delivery rate was 45.0% in the main group and 39.9% in the control group (p > 0.050).

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