Induction of a regular nuclear lattice by overexpression of NuMA.
Transient overexpression of nuclear mitotic apparatus protein (NuMA) in HeLa cells results in ordered lattices which can fill the nucleus and which are stable to detergent extraction. Electron microscopy reveals a quasi-hexagonal organization with an average spacing between the vertices of approximately 170 nm and short 6-nm-diameter rods connecting the vertices. Overexpression of a NuMA construct with an in-frame addition in the coiled-coil domain shows hexagons with the spacing increased by 42% while constructs with deletions in the coiled-coil domain yield hexagons with the spacing decreased by 40 and 19%. NuMA constructs truncated at residue 2005 or 2030 in the tail domain cause a drastic reorganization of nuclear components with relocation of the DNA, histone H1, and nucleoli to the nuclear rim. A construct lacking the head and much of the coiled-coil region also affects nuclear organization. In contrast, NuMA constructs truncated at residue 1950 or 1935 which lack the nuclear localization signal display normal nuclear structure but form cytoplasmic aggregates which also display hexagonal organization. Immunoelectron microscopy confirms that the nuclear lattices are built from NuMA. We discuss the importance of the different domains of NuMA for building the ordered in vivo lattices and whether NuMA could play a structural role in the architecture of the normal interphase nucleus.
Gueth-Hallonet
C
C
Department of Biochemistry, Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, Goettingen, 37077, Germany.
Wang
J
J
Harborth
J
J
Weber
K
K
Osborn
M
M
D034961:Antigens, Nuclear; D018797:Cell Cycle Proteins; D002467:Cell Nucleus / Q000378:metabolism / Q000648:ultrastructure*; D006367:HeLa Cells; D006801:Humans; D007150:Immunohistochemistry; D034521:Nuclear Matrix-Associated Proteins; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D010455:Peptides / Q000378:metabolism; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D034961:Antigens, Nuclear; D018797:Cell Cycle Proteins; C050045:NUMA1 protein, human; D034521:Nuclear Matrix-Associated Proteins; D009687:Nuclear Proteins; D010455:Peptides; D016271:Proto-Oncogene Proteins c-myc; D011993:Recombinant Fusion Proteins