Abstract
AbstractCytokinesis, the last step during cell division, is a highly coordinated process that involves the relay of signals from both the outside and inside of the cell. We have a basic understanding of how cells regulate internal events, but how cells respond to extracellular cues is less explored. In a systematic RNAi screen of G proteinācoupled receptors (GPCRs) and their effectors, we found that some GPCRs are involved in cytokinesis. RNAi knockdown of these GPCRs caused increased binucleated cell formation, and live cell imaging showed that most formed midbodies but failed at the abscission stage. OR2A4 (olfactory receptor, family 2, subfamily A, member 4) localized to cytokinetic structures in cells and its knockdown caused cytokinesis failure at an earlier stage, likely due to effects on the actin cytoskeleton. Identifying the downstream components that transmit GPCR signals during cytokinesis will be the next step and we show that GIPC1 (GIPC PDZ domain containing family, member 1), an adaptor protein for GPCRs, may play a part. RNAi knockdown of GIPC1 significantly increased binucleated cell formation. Understanding the molecular details of GPCRs and their interaction proteins in cytokinesis regulation will give us important clues about GPCRs signaling as well as how cells communicate with their environment during division. Ā© 2012 Wiley Periodicals, Inc
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🏛️ Research Organizations (ROR)
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📊 Figures
Figure 1
Systematic screen for cytokinesis inhibition with siRNAs targeting GPCRs and effector proteins
Schematic flow chart illustrates our systematic GPCR RNAi screen. HeLa cells were plated in 384 well plates before they were transfected with GPCR siRNAs at ICCB-Longwood. Cells were incubated for 72 ...
Figure 2
Knockdown of several GPCRs causes binucleated and multinucleated cell formation
(A) Control or GPCR RNAi cells were stained with TAMRA-NHS (shown in red) to visualize whole HeLa cells, and DAPI (shown in green) for DNA. Images were taken using an OPERA high content confocal micro...
Figure 3
GPCR knockdowns cause cytokinesis failure at early (OR2A4) and late stages (other GPCRs)
(A) Schematic illustration of cytokinesis failure after RNAi depletion of the 12 hit GPCRs. 11/12 GPCR knockdowns show cytokinesis failure at the abscission stage, where midbody bridges were formed be...
Figure 4
OR2A4 localizes to cytokinetic structures and affects the actin cytoskeleton
(A) Western blots with OR2A4 antibody show the knockdown of OR2A4. Tubulin and Actin were used as loading controls. (B) OR2A4 localizes to the spindle poles (red arrows) and chromosomes (green arrow) ...
Figure 5
GIPC1 RNAi knockdown causes cytokinesis inhibition
(A) Western blots with GIPC1 antibody show knockdown efficiency of GIPC1 RNAi. Tubulin was used as loading control. (B) Real-Time quantitative PCR shows knockdown efficiency of GIPC1 RNAi. 28S RNA was...
Figure images are served from the NIH/NLM PubMed Central Open Access Subset or Europe PMC; copyright remains with the publishers and authors.
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