Evaluating the involvement of autolysosomes in the nuclear translocation of fluorescent proteins

During the construction of control mCherry-labelled HeLa cells, we unexpectedly observed red fluorescence in cell nuclei and therefore investigated the mechanisms underlying the transport of fluorescent proteins (FPs) into the nuclei. We confirmed that mCherry and mCherry/EGFP tandem FPs were translocated into the nucleus and found that FPs are taken up by autophagosomes and translocated into the nucleus after entering lysosomes. However, pharmacological inhibition of autophagosome-lysosome fusion and syntaxin 17 knockdown decreased the nuclear translocation of mCherry in HeLa cells, but not in HepG2 cells, indicating that autophagy may also be involved in the nuclear translocation of FPs. Electron microscopy revealed that autolysosomes fused with the nuclear envelope and continued into the nucleus in HeLa cells but not in HepG2 cells, indicating that autophagy is involved in the nuclear translocation of FPs in HeLa cells. In addition, immunotransmission electron microscopy revealed that FPs can be transported directly into the nucleus through the nuclear pore complex. Our results suggest that autophagy is involved in the intracellular degradation and nuclear translocalisation of FPs.