Nuclear import of metallothionein requires its mRNA to be associated with the perinuclear cytoskeleton.
The influence of mRNA localization on metallothionein-1 protein distribution was studied by immunocytochemistry. We used Chinese hamster ovary cells that had been transfected with either a native metallothionein-1 gene construct or metallothionein-1 5'-untranslated region and coding sequences linked to the 3'-untranslated region from glutathione peroxidase. The change in the 3'-untranslated region caused the delocalization of the mRNA with a loss of the perinuclear localization and association with the cytoskeleton. Clones were selected which expressed similar levels of metallothionein-1 protein, as assessed by radioimmunoassay. The results showed that loss of metallothionein-1 mRNA localization was associated with a loss of metallothionein-1 protein localization, most notably with a lack of metallothionein-1 protein in the nucleus of synchronized cells which were beginning to synthesize DNA. This indicates that the association of metallothionein-1 mRNA with the cytoskeleton around the nucleus is essential for efficient shuttling of the protein into the nucleus during the G(1) to S phase transition. This is the first demonstration of a physiological role for perinuclear mRNA localization and we propose that such localization may be important for a wide range of nuclear proteins, including those that shuttle between nucleus and cytoplasm in a cell cycle dependent manner.
Levadoux
M
M
Rowett Research Institute, Greenburn Road, Bucksburn, Aberdeen AB21 9SB, United Kingdom.
Mahon
C
C
Beattie
J H
JH
Wallace
H M
HM
Hesketh
J E
JE
D020413:3' Untranslated Regions / Q000235:genetics; D000818:Animals; D001692:Biological Transport; D016466:CHO Cells; D002453:Cell Cycle; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism*; D006224:Cricetinae; D003593:Cytoplasm / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism*; D005979:Glutathione Peroxidase / Q000235:genetics / Q000378:metabolism; D007150:Immunohistochemistry; D008668:Metallothionein / Q000235:genetics / Q000378:metabolism*; D018613:Microscopy, Confocal; D012333:RNA, Messenger / Q000378:metabolism*; D016196:S Phase; D013936:Thymidine / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
D020413:3' Untranslated Regions; D012333:RNA, Messenger; D008668:Metallothionein; D005979:Glutathione Peroxidase; D013936:Thymidine