Beta-catenin is imported into the nucleus by binding directly to the nuclear pore machinery, similar to importin-beta/beta-karyopherin or other importin-beta-like import factors, such as transportin. These findings provide an explanation for how beta-catenin localizes to the nucleus without an NLS and independently of its interaction with TCF/LEF-1. This is a new and unusual mechanism for the nuclear import of a signal transduction protein. The lack of beta-catenin import activity in the presence of normal cytosol suggests that its import may be regulated by upstream events in the Wnt signaling pathway.

Abstract. 13-Catenin, a cytoplasmic protein known for its association with cadherin cell adhesion molecules, is also part of a signaling cascade involved in embryonic patterning processes such as the determination of the dorsoventral axis in Xenopus and determination of segment polarity in Drosophila. Previous studies suggest that increased cytoplasmic levels of 13-catenin correlate with signaling, raising questions about the need for interaction with cadherins in this process. We have tested the role of the 13-catenin-cadherin interaction in axis formation. Using [3-catenin deletion mutants, we demonstrate that significant binding to cadherins can be eliminated without affecting the signaling activity. Also, depletion of the soluble, cytosolic pool of 13-catenin by binding to overexpressed C-cadherin completely inhibited [3-catenin-inducing activity. We conclude that binding to cadherins is not required for [3-catenin signaling, and therefore the signaling function of [3-catenin is independent of its role in cell adhesion. Moreover, because 13-catenin signaling is antagonized by binding to cadherins, we suggest that cadherins can act as regulators of the intracellular [3-catenin signaling pathway.

Human cytoskeletal a-actinin cDNA was transfected into highly malignant simian virus 40-transformed BALB/c 3T3 (SVT2) cells that express 6-fold lower levels of a-actinin than nontransformed BALB/c 3T3 cells. SVT2 clones expressing various levels of a-actinin were isolated and their structure and tumorigenic properties were determined. Transfected SVT2 clones expressing a-actinin at levels found in nontumorigenic 3T3 cells displayed a flatter phenotype, a decreased ability to grow in suspension culture in soft agar, and a marked reduction in their ability to form tumors in syngeneic BALB/c mice and in athymic nude mice. Clones overexpressing a-actinin at the highest level (about 2-fold higher than 3T3 cells) were completely suppressed in their ability to form tumors in syngeneic BALB/c mice. The results suggest that a-actinin, an actin-crosslinking protein that is also localized in cell junctions, may have an effective suppressive ability on the transformed phenotype.Malignant transformation of cells is characterized by alterations in cell growth, adhesion, motility, and cell shape (1). Among the more conspicuous morphological alterations in transformed cells are a round phenotype and a decrease in the number of microfilament bundles (2, 3). The disorganization of the actin filaments in transformed cells is often associated with a reduced expression of microfilament proteins such as tropomyosin (4), gelsolin (5), and vinculin (6). These changes in microfilaments correlate with the ability of cells to grow in suspension in semisolid medium (7).Previous studies have demonstrated a reversion in the transformed phenotype of cancer cells in the presence of a variety of agents and a return to the original malignant behavior upon their removal (8). This "reverse transformation" includes a reversible modulation in cell shape and the organization of the cytoskeleton (9). However, the relationships between the changes in growth properties and the altered cell structure of transformed cells are still unknown (10). Do the characteristic changes in cell morphology of transformed cells result from alterations in the rate of growth or are changes in cell adhesion and cytostructure responsible for disruption of the adhesion-dependent growth control? In this study we addressed this question directly by modulating the expression ofa single actin-associated protein a-actinin in a tumorigenic cell line and determined the consequences of altered a-actinin expression on the phenotype of the cells. a-Actinin is an abundant cytoplasmic actin-binding protein (11) that crosslinks actin filaments in vitro and is found in both muscle and nonmuscle cells (12,13). In striated muscle cells, a-actinin is localized in Z-disks; in smooth muscle cells, it is in dense plaques; and in nonmuscle cells, a-actinin is found along microfilaments (14) and in adherens junctions (AJs), at sites where actin filaments attach to the membrane (15)(16)(17). The role of a-actinin in mediating actin attachment to the membrane is not completely understood. ...

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Actin filaments are major determinants of cell shape, motility and adhesion, which control important biological processes including embryonic development and wound healing. These processes are associated with changes in actin assembly, which is regulated by controlling the balance between polymerized and non-polymerized actin. To maintain a significant pool of non-polymerized actin, mechanism(s) linking actin synthesis to its state of polymerization were proposed. We have studied this relationship between actin synthesis and organization by modulating actin assembly using different drugs. Unassembled actin was increased in 3T3 cells using either the Clostridium botulinum C2 toxin, which ADP-ribosylates actin, or by latrunculin A, a Red Sea sponge product, which binds monomeric actin. The synthesis of actin was dramatically reduced in these cells owing to a concomitant decrease in actin RNA level. Similar results were obtained with HeLa cells grown in both monolayer and in suspension, suggesting that cell shape changes associated with drug treatment are not the primary cause for the effect on actin synthesis. In contrast, the scrape-loading of 3T3 cells with phalloidin, a stabilizer of polymerized actin that increased the level of assembled actin, resulted in elevated actin synthesis and RNA content. The expression of vinculin, a major component of adhesion plaques and cell-cell junctions, which is involved in actin-membrane associations, was altered in parallel with that of actin in cells treated with these drugs. The decrease in actin RNA resulted from destabilization of actin mRNA in cells where unassembled actin level was elevated. This is suggested by the unchanged transcription of actin in isolated nuclei from drug-treated cells, and by demonstrating that actin mRNA was degraded faster in cells after C2 toxin treatment than in control cells. This feedback control mechanism is mainly confined to the cytoplasm, as it remained active in enucleated cells. The results suggest the existence of an autoregulatory pathway for the expression of actin and other microfilament-associated proteins which is linked to the state of actin polymerization in the cell.

alpha-Actinin is an abundant actin crosslinking protein, also localized at adherens type junctions. In adhesion plaques, alpha-actinin can link the actin filaments to integrin via vinculin and talin, or directly by binding to the cytoplasmic domain of beta 1-integrin. The expression of alpha-actinin is rapidly elevated in growth-activated quiescent cells, and is reduced in SV40-transformed 3T3 cells and various differentiating cell types (reviewed by Gluck, U., Kwiatkowski, D. J. and Ben-Ze'ev, A. Proc. Nat. Acad. Sci. USA 90, 383–387, 1993). To study the effect of changes in alpha-actinin levels on cell behavior, alpha-actinin expression was elevated in 3T3 cells by transfection with a full-length human nonmuscle alpha-actinin cDNA. To suppress alpha-actinin levels, 3T3 cells were transfected with an antisense alpha-actinin cDNA construct. Cells overexpressing alpha-actinin by 40–60% displayed a significant reduction in cell motility, as demonstrated by their slower locomotion into an artificial wound, and by forming shorter phagokinetic tracks on colloidal gold-coated substrata. 3T3 cells in which the expression of alpha-actinin was reduced to 25–60% of control levels, after antisense alpha-actinin transfection, had an increased cell motility. Moreover, such alpha-actinin-deficient 3T3 cells formed tumors upon injection into nude mice. The results demonstrate that modulations in alpha-actinin expression can affect, in a major way, the motile and tumorigenic properties of cells, and support the view that decreased alpha-actinin expression could be a common regulatory pathway to malignant transformation of 3T3 cells.