Tpr, a large coiled coil protein whose amino terminus is involved in activation of oncogenic kinases, is localized to the cytoplasmic surface of the nuclear pore complex.

Abstract

Abstract. From a panel of monoclonal antibodiesraised against fractions of rat liver nuclear envelopes (NEs), we have identified an antibody, RL30, which reacts with novel nuclear pore complex (NPC) antigens that are not O-glycosylated. By immunofluorescence staining of cultured cells, RL30 reacts exclusively with the NE in a punctate pattern that largely coincides with that of identified NPC proteins. RL30 labels only the cytoplasmic surface of the NPC in immunogold electron microscopy, predominantly in peripheral regions nearby the cytoplasmic ring. In immunoblots of isolated rat liver NEs and cultured rat cells, RL30 recognizes a 265-kD band, as well as a series of 175-265-kD bands in rat liver NEs that are likely to be proteolytic products of p265. Sequencing of peptides from the 175-and 265-kD RL30 antigens of rat liver revealed that they are both closely related to human Tpr, a protein whose amino-terminal 150-250 amino acids appear in oncogenic fusions with the kinase domains of the met, trk, and raf protooncogenes.We found that in vitro translation of human Tpr mRNA yields a major 265-kD band. Considered together, these data indicate that the 265-kD RL30 antigen in the NPC is the rat homologue of Tpr. Interestingly, Tpr contains an exceptionally long predicted coiled coil domain (,x,1600 amino acids). The localization and predicted structure of Tpr suggest that it is a component of the cytoplasmic fibrils of the NPC implicated in nuclear protein import. Immunofluorescence microscopy shows that during NPC reassembly at the end of mitosis, Tpr becomes concentrated at the NE significantly later than O-linked glycoproteins, including p62. This indicates that reassembly of the NPC after mitosis is a stepwise process, and that the Tpr-containing peripheral structures are assembled later than p62. N 'UCLEOCYTOPLASMIC transport is carried out by nuclear pore complexes (NPCs)' (reviewed by Forbes, 1992;Gerace, 1992;Pant6 and Aebi, 1993;Fabre and Hurt, 1994), large supramolecular structures that span the nuclear envelope (NE) at regions where the inner and outer nuclear membranes are joined. The NPC contains aqueous channels that provide conduits for nonselective passive diffusion of ions, metabolites, and other small molecules (Peters, 1986). Most proteins and RNAs are too large to diffuse through these channels at physiologically significant rates, and instead are actively transported through a

Role of TFG sequences outside the coiled-coil domain in TRK-T3 oncogenic activation

“…Several thyroid TRK oncogenes have been isolated in our laboratory and named TRK, TRK-T1, TRK-T2 and TRK-T3. TRK oncogene is activated by TPM3, encoding the nonmuscle tropomyosin (MartinZanca et al, 1986); TRK-T1 and TRK-T2 are both activated by TPR gene, encoding a filamentous protein of the nuclear pore complex (Byrd et al, 1994); in TRK-T3 oncogene, the activating portion is contributed by TFG, a novel gene on chromosome 3 encoding a protein of yet unknown function (Greco et al, 1995). All the activating sequences contain dimerization/oligomerization domains playing a crucial role in oncogenic activation by inducing constitutive, ligand-independent tyrosine kinase activity.…”

Regulation of the urokinase-type plasminogen activator gene by the oncogene Tpr-Met involves GRB2

“…The receptor for HGF/SF was identi®ed as the normal counterpart of an oncogene, Tpr-Met, that arose in cell culture, by a spontaneous chromosomal translocation after treatment with a carcinogen (Park et al, 1986;Cooper et al, 1984). In Tpr-Met, the leucine-zipper dimerization domain of the nuclear envelope protein Tpr (Byrd et al, 1994) is linked to the intracellular moiety of the HGF/SF receptor, Met, containing the tyrosine kinase domain (reviewed in Comoglio, 1993). Thus, while Met dimerizes and becomes active as a signal transducer upon ligand binding, Tpr-Met spontaneously dimerizes through the leucine-zipper domain of Tpr, in the absence of the ligand, and is constitutively active (Rodrigues and Park, 1993).…”

The C-terminal domain of myosin-like protein 1 (Mlp1p) is a docking site for heterogeneous nuclear ribonucleoproteins that are required for mRNA export

“…In mammalian cells, the translocated promoter region (Tpr) protein was originally identified as a component of the filaments that extend from the NPC into the nuclear interior (16,17). Tpr is a 270-kDa protein consisting mostly of an ␣-helical coiled-coil structure with a carboxyl-terminal globular domain (18,19). Recently, Tpr has been localized to both the nuclear basket of the NPC and discrete foci within the nucleus (20).…”