Targeting of the mammalian nucleoporin p62 to the nuclear envelope in the yeast Saccharomyces cerevisiae and HeLa cells.
We have analyzed the sorting of the mammalian nucleoporin p62 in human culture cells and in the yeast Saccharomyces cerevisiae. To this end, gene fusions were generated that carry Aequorea victoria green fluorescence protein and defined portions of p62. Upon transient gene expression fluorescent fusion proteins were localized in HeLa cells. Likewise, fusion proteins were studied in S. cerevisiae using wild-type as well as mutant cells that cluster nuclear pore complexes. Our results demonstrate that evolutionarily distant organisms, such as humans and yeasts, recognize the same sequence elements of p62 for sorting to the nuclear envelope. Specifically, the entire sequence of p62 or its complete C-terminal domain targeted fusion proteins to the nuclear membranes. In contrast, truncations of the C-terminal domain or the N-terminal segment of p62 failed to associate with the nuclear envelope in either organism. In HeLa cells overexpression of several p62-containing fusion proteins resulted in nuclear fragmentation. The C-terminal domain of p62 caused this effect, and amino acid residues 477 to 525 were sufficient to induce aberrant nuclei. Thus, overexpression of 49 amino acid residues located at the C-terminal tail of p62 interferes with the nuclear integrity in human culture cells.
Barth
W
W
Physiology Department, McGill University, Montreal, QC, Canada.
Chatterjee
S
S
Stochaj
U
U
D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D009685:Nuclear Envelope / Q000378:metabolism*; D028861:Nuclear Pore Complex Proteins; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D014162:Transfection
D008164:Luminescent Proteins; D008562:Membrane Glycoproteins; D028861:Nuclear Pore Complex Proteins; D011993:Recombinant Fusion Proteins; C049312:nuclear pore protein p62; D049452:Green Fluorescent Proteins