Degradation of GFP-labelled POM121, a non-invasive sensor of nuclear apoptosis, precedes clustering of nuclear pores and externalisation of phosphatidylserine.
The nuclear pore membrane protein POM121 is specifically degraded during apoptosis by a caspase-3-dependent process enabling early detection of apoptosis in living cells expressing POM121-GFP. Here we further investigated temporal aspects of apoptotic degradation of POM121-GFP. We demonstrate that decreased POM121-GFP fluorescence precedes annexin V-labelling of apoptotic cells. This indicates that degradation of the nuclear pore complex starts prior to redistribution of plasma membrane phosphatidylserine, which serves as a signal for phagocytotic elimination of apoptotic cells. Furthermore, a caspase-resistant GFP-labelled mutant of POM121 resisted degradation even in late apoptosis and was detected in clustered nuclear pores. Thus, it can be concluded that loss of POM121-GFP is a specific sensor of the activation of caspase-3-dependent proteolysis at the nuclear pores.
Beckman
M
M
Department of Neurochemistry and Neurotoxicology, Stockholm University, SE-106 91 Stockholm, Sweden.
Kihlmark
M
M
Iverfeldt
K
K
Hallberg
E
E
D000818:Animals; D017304:Annexin A5 / Q000378:metabolism; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000502:physiology*; D006224:Cricetinae; D003412:Cricetulus; D004789:Enzyme Activation; D006868:Hydrolysis; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D009154:Mutation; D022022:Nuclear Pore / Q000378:metabolism*; D016716:PC12 Cells; D010718:Phosphatidylserines / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D013997:Time Factors
D017304:Annexin A5; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; C094157:POM121 protein, human; C511307:POMZP3 protein, human; D010718:Phosphatidylserines; D011994:Recombinant Proteins; D020169:Caspases