Xl. Zhu et al., CARBOXYL-TERMINUS OF MITOSIN IS SUFFICIENT TO CONFER SPINDLE POLE LOCALIZATION, Journal of cellular biochemistry, 66(4), 1997, pp. 441-449
Mitosin is a nuclear protein of 3,113 amino acids which has been shown
to associate with the mitotic apparatus, especially the kinetochore,
during mitosis. In this paper we further confirmed its association wit
h the spindle poles in normal monkey kidney CV1 cells by indirect immu
nofluorescence microscopy. When the carboxyl portion of mitosin contai
ning amino acids 2,094-3,113 (named mitosin-pTN) was stably expressed
in rat fibroblast Rat2 cells using a tetracycline-inducible system, st
rong spindle pole association was observed in addition to expected cen
tromere localization. The same results were achieved in Chinese hamste
r ovary (CHO) cells. On the other hand, mitosin-pTC containing amino a
cids 2,756-3,113 was not targeted to spindle poles. Use of the FLAG ep
itope [Hopp et al., 1988] genetically fused to each amino terminus of
these mutants eliminated possible artifacts due to antibody cross-reac
tion, since the spindle pole localization of wild-type mitosin was con
firmed with a FLAG-tagged mutant by an antibody (anti-FLAG M2 monoclon
al antibody) irrelevant to antibodies to mitosin. Our data also sugges
ted a possible interaction of mitosin with the spindle microtubules. I
nteraction of mitosin with the major parts of the mitotic apparatus fu
rther implies an important role in mitosis. (C) 1997 Wiley-Liss, Inc.