DEDD association with cytokeratin filaments correlates with sensitivity to apoptosis.
The cytokeratin 8/18 (CK8/18) cytoskeleton network is an early target for caspase cleavage during apoptosis. Recent reports suggest that the highly conserved and ubiquitous death effector domain containing DNA binding protein (DEDD) plays a role in the recruitment of procaspase-9 and -3 at this CK8/18 scaffold. DEDD interacts with both the CK8/18 intermediate filament network and procaspase-3 and -9. It is suggested that the CK8/18 fibrils may provide a scaffold for the proximity-induced autocleavage and activation of procaspase-9 in close association with caspase-3.We addressed this issue by investigating DEDD staining patterns in various cell lines and by correlating these expression patterns with the sensitivity of these cell lines for roscovitine-induced apoptosis. We showed that in some cell lines DEDD revealed a bright filamentous staining pattern in others DEDD staining was weak and diffusely distributed in the cytoplasm of the cells. The difference in staining patterns was irrespective of the phosphorylation status of the cytokeratin filaments. In cells showing a filamentous staining pattern, DEDD was strongly associated with the CK8/18 cytokeratin filaments as evidenced by double immunofluorescence and its resistance to extraction with Triton X-100. Subcellular fractionation indicates that DEDD co-purifies with CK18, which corroborates a strong association of DEDD and the cytokeratin network. DEDD was either mono- or diubiquinated. Cells showing a filamentous DEDD distribution are more apoptosis-prone as evidenced by the rapid appearance of M30 CytoDeath-positive cells after induction of apoptosis. The sensitivity towards apoptosis is irrespective of the procaspase-3 content of the cells. Our data support the notion that DEDD-mediated accumulation of procaspases at the cytokeratin scaffold leads to an increase in the local concentration, which renders cells more apoptosis-prone.
Schutte
Bert
B
Department of Molecular Cell Biology, Research Institute Growth & Development, GROW, University of Maastricht, P.O. Box 616, 6200, MD, Maastricht, The Netherlands. bert.schutte@molcelb.unimaas.nl
Henfling
Mieke
M
Ramaekers
Frans C S
FC
D017209:Apoptosis / Q000187:drug effects; D018938:Caco-2 Cells; D004268:DNA-Binding Proteins / Q000378:metabolism* / Q000502:physiology*; D053418:Death Domain Receptor Signaling Adaptor Proteins / Q000378:metabolism* / Q000502:physiology*; D019008:Drug Resistance, Neoplasm; D015870:Gene Expression; D045325:HCT116 Cells; D019073:HT29 Cells; D006367:HeLa Cells; D006801:Humans; D007633:Keratins / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology; D014018:Tissue Distribution; D025801:Ubiquitin / Q000378:metabolism
C116096:DEDD protein, human; D004268:DNA-Binding Proteins; D053418:Death Domain Receptor Signaling Adaptor Proteins; D025801:Ubiquitin; D007633:Keratins