Early localization of NPA58, a rat nuclear pore-associated protein, to the reforming nuclear envelope during mitosis.

We have studied the mitotic reassembly of the nuclear envelope, using antibodies to nuclear marker proteins and NPA58 in F-111 rat fibroblast cells. In earlier studies we have proposed that NPA58, a 58 kDa rat nuclear protein, is involved in nuclear protein import. In this report, NPA58 is shown to be localized on the cytoplasmic face of the envelope in interphase cells, in close association with nuclear pores. In mitotic cells NPA58 is dispersed in the cytoplasm till anaphase. The targeting of NPA58 to the reforming nuclear envelope in early telophase coincides with the recruitment of a well-characterized class of nuclear pore proteins recognized by the antibody mAb 414, and occurs prior to the incorporation of lamin B1 into the envelope. Significant protein import activity is detectable only after localization of NPA58 in the newly-formed envelope. The early targeting of NPA58 is consistent with its proposed role in nuclear transport.

Nucleocytoplasmic transport of proteins and control of cellular function.

Nucleocytoplasmic transport of proteins occurs through pores embedded in the nuclear envelope. Recent studies have defined the cytoplasmic factors required for signal-mediated import of nuclear proteins. Considerable progress has been made in understanding the mechanism of nuclear export of proteins by the identification of specific signal sequences needed for export. Regulatory molecules have been shown to adopt novel mechanisms to control their entry into the nucleus and thereby regulate their functions.

Effect of nuclear localization signal-receptor interaction on nuclear envelope-associated ATPase activity.

The possibility that interaction of the nuclear localization signal (NLS) with its pore receptor may directly stimulate nuclear envelope-associated ATPase activity and consequently provide energy for protein translocation across the pore has been studied. ATPase activity was assayed after cross-linking of the prototype NLS peptide with its pore receptor, or after preincubation of envelopes with NLS-albumin conjugates. Neither treatment enhanced enzyme activity. A more complex series of events may be required for energy-generation at the nuclear pore.

A conserved epitope on nuclear pore phosphoproteins reflects cell division status.

The nuclear pore complexes mediate the selective nuclear import of proteins in a signal- and energy-dependent process. We have earlier reported the characterization of a monoclonal antibody, Mab E2, that recognizes a novel class of nuclear pore phosphoproteins involved in signal-binding and protein transport. In the present study, we have analyzed the pattern of immunoreactivity of Mab E2 in cultured rat fibroblasts and have observed significant differences in the expression of epitopes in proliferating and quiescent cells. Furthermore, the common epitope recognized by Mab E2 is conserved across species, consistent with its essential role in nuclear protein import.