A nonerythroid isoform of protein 4.1R interacts with the nuclear mitotic apparatus (NuMA) protein

Red blood cell protein 4.1 (4.1R) is an 80-kD erythrocyte phosphoprotein th at stabilizes the spectrin/actin cytoskeleton. In nonerythroid cells, multi ple 4.1R isoforms arise from a single gene by alternative splicing and pred ominantly code for a 135-kD isoform. This isoform contains a 209 amino acid extension at its NH2 terminus (head piece; HP). Immunoreactive epitopes sp ecific for HP have been detected within the cell nucleus, nuclear matrix, c entrosomes, and parts of the: mitotic apparatus in dividing cells. Using a yeast two-hybrid system, in vitro binding assays, coimmunolocalization, and coimmunoprecipitation studies, we show that a 135-kD 4.1R isoform specific ally interacts with the nuclear mitotic apparatus (NuMA) protein. NuMA and 4.1R partially colocalize in the interphase nucleus of MDCK cells and redis tribute to the spindle poles early in mitosis, Protein 4.1R associates with NuMA in the interphase nucleus and forms a complex with spindle pole organ izing proteins, NuMA, dynein, and dynactin during cell division. Overexpres sion of a 135-kD isoform of 4.1R alters the normal distribution of NuMA in the interphase nucleus. The minimal sequence sufficient for this interactio n has been mapped to the amino acids encoded by exons 20 and 21 of 4.1R and residues 1788-1810 of NuMA. Our results not only suggest that 4.1R could, possibly, play an important role in organizing the nuclear architecture, mi totic spindle, and spindle poles, but also could define a novel role for it s 22-24-kD domain.