2002
DOI: 10.1002/cm.10032
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Inducing precocious anaphase in cultured mammalian cells

Abstract: The spindle checkpoint, which prevents anaphase onset upon spindle damage or incorrect chromosome alignment, presents a problem for experimental analysis of protein function in anaphase and cytokinesis. This is because the functional disruption of many proteins before anaphase onset can activate this checkpoint, preventing anaphase and subsequent cell cycle events. This paper compares new and old methods of overriding the spindle checkpoint in prometaphase mammalian tissue culture cells. Cell Motil. Cytoskelet… Show more

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Cited by 38 publications
(31 citation statements)
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“…Thus, CeMCAK and HCP-1/2 do not seem to disrupt kineto-chore structure generally and could be involved more directly in checkpoint assembly or function. Indeed, the precocious anaphase observed in hcp-1/2(RNAi) embryos is characteristic of other spindle checkpoint proteins (Gorbsky et al, 1998;Canman et al, 2002) and is consistent with the role of hcp-1/2 in checkpoint function.…”
Section: Kinetochore Proteins and Spindle Checkpoint Functionsupporting
confidence: 74%
“…Thus, CeMCAK and HCP-1/2 do not seem to disrupt kineto-chore structure generally and could be involved more directly in checkpoint assembly or function. Indeed, the precocious anaphase observed in hcp-1/2(RNAi) embryos is characteristic of other spindle checkpoint proteins (Gorbsky et al, 1998;Canman et al, 2002) and is consistent with the role of hcp-1/2 in checkpoint function.…”
Section: Kinetochore Proteins and Spindle Checkpoint Functionsupporting
confidence: 74%
“…This is because microinjection of Mad1F10 substantially shortens the delay. Mad1F10 lacks the kinetochore targeting domain of Mad1 but retains the Mad2 binding domain (Canman et al, 2002a(Canman et al, , 2002bChung and Chen, 2002), which is nearly identical to the Mad2 binding domain in Cdc20 (Luo et al, 2002). In vitro, Mad2 forms complexes exclusively with either Mad1 or Cdc20 and does not form a Mad1-Mad2-Cdc20 ternary complex (Sironi et al, 2001).…”
Section: Hypothermic Conditions Results In Decreased Tension and A Chementioning
confidence: 99%
“…GH cells were studied because of their relative high levels of MAD2 and high transfection efficiency. MAD2DC has been reported to override the function of MAD2, therefore, it is regarded as the dominantnegative form of MAD2 (Canman et al, 2002). As shown in Figure 4A, HA protein was detected in cells transfected with MAD2DC but not in the vector control cells (see the arrow).…”
Section: Inactivation Of Mad2 Leads To Downregulation Of Mek/erk Pathwaymentioning
confidence: 94%
“…For plasmid construction, a 1.5-kb fragment containing HA-tagged truncated form of human MAD2 cDNA (MAD2DC) was cloned into pLenti6/V5-D-TOPO expression vector by TOPO cloning reaction (Invitrogen, Carlsbad, CA, USA). MAD2DC lacks the C-terminal 10 amino acids that is necessary for the function of MAD2 (Canman et al, 2002). Cisplatin was added 48 h post-transfection.…”
Section: Transfectionmentioning
confidence: 99%