Figure 4. CERKL-HA (isoform a, 532 amino acids) shows a dynamic subcellular localization in COS-7 transfected cells. A: Several cells per field showed uniform distribution of CERKL in the cytosol and the nucleus, with clear exclusion from the nucleoli. B: A similar pattern but with strong accumulation in the nucleoli (nuclei, counter-stained with DAPI, appear in blue). C and D: In most cells, CERKL was absent from the nucleus and instead accumulated in clusters, preferentially in the perinuclear region (A). These two patterns were cell-specific and could be observed in the same field (D). E: CERKL localized to several membranous subcellular compartments, mainly ER and Golgi. The markers used were calnexin for ER, GM130 for Golgi, EEA1 for endosomes and MitoTracker for mitochondria. F: R257X truncated CERKL localized preferentially in the nuclei, although it was also detected in the ER. Arrows highlight the relevant CERKL localizations, as immunodetected with an anti-HA monoclonal antibody. Scale bar corresponds to 10 μm. The same results were obtained for all the CERKL isoforms, irrespective of the epitope used, HA or GFP (data not shown). Abbreviations: endoplasmic reticulum (ER), endosomes (End), mitochondria (Mit).