Peroxisomal targeting signals (PTSs) are recognized by predominantly cytosolic receptors, Pex5p and Pex7p. The fate of these PTS receptors following their interactions on the peroxisomal membrane with components of docking and putative translocation complexes is unknown. Using both novel and multiple experimental approaches, we show that human Pex5p does not just bind cargo and deliver it to the peroxisome membrane, but participates in multiple rounds of entry into the peroxisome matrix and export to the cytosol independent of the PTS2 import pathway. This unusual shuttling mechanism for the PTS1 receptor distinguishes protein import into peroxisomes from that into most other organelles, with the exception of the nucleus.

Human nm23 has been implicated in suppression of metastasis in various cancers, but the underlying mechanism of such activity has not been fully understood. Using Drosophila tracheal system as a genetic model, we examined the function of the Drosophila homolog of nm23, the awd gene, in cell migration. We show that loss of Drosophila awd results in dysregulated tracheal cell motility. This phenotype can be suppressed by reducing the dosage of the chemotactic FGF receptor (FGFR) homolog, breathless (btl), indicating that btl and awd are functionally antagonists. In addition, mutants of shi/dynamin show similar tracheal phenotypes as in awd and exacerbate those in awd mutant, suggesting defects in vesicle-mediated turnover of FGFR in the awd mutant. Consistent with this, Btl-GFP chimera expressed from a cognate btl promoter-driven system accumulate at high levels on tracheal cell membrane of awd mutants as well as in awd RNA duplex-treated cultured cells. Thus, we propose that awd regulates tracheal cell motility by modulating the FGFR levels, through a dynamin-mediated pathway.[Keywords: awd/nm23; btl/FGFR; shi/dynamin; tracheal development; Drosophila; cell migration] Supplemental material is available at http://www.genesdev.org.

We show that localized expression of the integrin α 3 protein is regulated at the level of RNA localization by the human homologue of Drosophila Muscleblind, MLP1/MBLL/MBNL2, a unique Cys 3 His zinc-finger protein. This is supported by the following observations: MLP1 knockdown abolishes localization of integrin α 3 to the adhesion complexes; MLP1 is localized in adhesion plaques that contain phospho-focal adhesion kinase; this localization is microtubule-dependent; integrin α 3 transcripts colocalize with MLP1 in distinct cytoplasmic loci; integrin α 3 transcripts are physically associated with MLP1 in cells and MLP1 binds to a specific ACACCC motif in the integrin α 3 3′ untranslated region (UTR) in vitro; and a green fluorescent protein (GFP) open reading frameintegrin α 3 3′ UTR chimeric gene directs GFP protein localization to distinct cytoplasmic loci near the cell periphery, which is dependent on MLP1 and is mediated by the ACACCC motif but is independent of the integrin α 3 signal peptide.We have recently identified a gene that is differentially expressed in many tumours. The cognate protein was named MLP1 for Muscleblind-like protein 1 (GenBank accession #AF491866) because of its high degree of homology with the Drosophila Muscleblind (Mbl) protein. The Drosophila mbl gene is involved in regulatory processes that are associated with the terminal differentiation of photoreceptors and muscles 1,2 . The Mbl protein is characterized by novel Cys 3 His zinc-finger motifs, CX 7 CX 6 CX 3 H-CX 7 CX 4 CX 3 H. Highly conserved homologues of mbl are found from worms and insects to mammals. Three human Mbl-like proteins -named EXP/MBNL1, MBLL/MBNL2 and MBXL/CHCR/MBNL3 -have been partially characterized 3-7 . MLP1 is the same as MBLL/MBNL2 and, as the former name was first used in the GenBank database, we will continue to use it throughout this report for brevity. These three proteins display RNA-binding activity towards transcripts that contain expanded trinucleotide repeats in myotonic dystrophy muscle cells, presumably sequestering the target RNAs 3-5 . These observations, however, did not shed light on the normal cellular functions of this family of proteins, although a role in mRNA processing has been implicated 7 . The developmental functions of Drosophila Mbl do not include early proliferation or differentiation of the muscle precursors 2 . In mbl mutants, muscles detach from the epidermis, indicating a defect in integrin-mediated adhesion. In addition, Mbl protein is found associated with the Zband that is rich in actin. Therefore, these mbl homologues may represent a unique group of regulatory genes that control the cellular functions that are related to cell-matrix interactions and cytoskeletal organization.

The tumor suppressor VHL (von Hippel-Lindau protein) serves as a negative regulator of hypoxia-inducible factor-␣ subunits. However, accumulated evidence indicates that VHL may play additional roles in other cellular functions. We report here a novel hypoxia-inducible factor-independent function of VHL in cell motility control via regulation of fibroblast growth factor receptor 1 (FGFR1) endocytosis. In VHL null tumor cells or VHL knock-down cells, FGFR1 internalization is defective, leading to surface accumulation and abnormal activation of FGFR1. The enhanced FGFR1 activity directly correlates with increased cell migration. VHL disease mutants, in two of the mutation hot spots favoring development of renal cell carcinoma, failed to rescue the above phenotype. Interestingly, surface accumulation of the chemotactic receptor appears to be selective in VHL mutant cells, since other surface proteins such as epidermal growth factor receptor, platelet-derived growth factor receptor, IGFR1, and c-Met are not affected. We demonstrate that 1) FGFR1 endocytosis is defective in the VHL mutant and is rescued by reexpression of wild-type VHL, 2) VHL is recruited to FGFR1-containing, but not EGFR-containing, endosomal vesicles, 3) VHL exhibits a functional relationship with Rab5a and dynamin 2 in FGFR1 internalization, and 4) the endocytic function of VHL is mediated through the metastasis suppressor Nm23, a protein known to regulate dynamin-dependent endocytosis.The von Hippel-Lindau disease is an inherited disorder that manifests in tumor formation in multiple organs (1, 2). The disease is characterized by highly vascularized tumors mainly due to overproduction of angiogenic factors. The underlying genetic defect was identified as mutations in the VHL tumor suppressor gene (3). The biological role of VHL is prominently linked to its E3 3 ubiquitin ligase activity toward a subset of cellular proteins, thus promoting their ubiquitination and degradation (4, 5). Prominent among its cellular targets are the ␣ subunits (1␣, 2␣, and 3␣) of the key transcription factor, hypoxia-inducible factor (HIF), involved in the cellular oxygen-sensing mechanism (6 -8).Tumorigenic mutations that affect E3 ligase function of VHL result in constitutive stabilization of HIF-␣, leading to transcriptional activation of several target genes of HIF (9), including those encoding critical angiogenic factors such as vascular endothelial growth factor and enzymes involved in glucose metabolism (10). Renal cell carcinomas (RCCs) harboring VHL mutations are often metastatic, and reexpression of wild-type VHL suppresses the metastatic behavior in RCC-derived cell lines (11,12), although the mechanisms remain unclear. VHL mutant cells exhibit increased scattering upon hepatocyte growth factor treatment (13). Enhanced response to SDF-1 (stromal cell-derived factor-1), due to overexpression of chemokine receptor CXCR4, was also recently identified as a possible mechanism by which VHL tumors might disseminate to distant organs (14). On the other hand, accumul...