Platelet-derived growth factor (PDGF) is a critical regulator of mesenchymal cell migration and proliferation. The vital functions of PDGFs for angiogenesis, as well as development of kidney, brain, cardiovascular system and pulmonary alveoli during embryogenesis, have been well demonstrated by gene knock-out approaches. Clinical studies reveal that aberrant expression of PDGF and its receptor is often associated with a variety of disorders including atherosclerosis, fibroproliferative diseases of lungs, kidneys and joints, and neoplasia. PDGF contributes to cancer development and progression by both autocrine and paracrine signaling mechanisms. In this review article, important features of the PDGF isoforms and their cell surface receptor subunits are discussed, with regards to signal transduction, PDGF-isoform specific cellular responses, and involvement in angiogensis, and tumorstromal interactions.

ABSTRACT

Platelet-derived growth factors (PDGFs) regulate a diverse array of cellular processes, including cell proliferation, transformation, migration, survival, and apoptosis of mesenchymal cells, in development as well as during pathogenesis (reviewed in references 31 and 42). For over 2 decades, PDGFs were thought to exist as the homodimers PDGF AA and BB or the heterodimer PDGF AB. These PDGF dimers are processed intracellularly and secreted as active dimers that readily activate PDGF receptors (PDGFRs). Recently, two new PDGF ligands (PDGF CC and DD) were discovered that have a unique two-domain structure with an N-terminal complement subcomponent C1r/C1s, Uegf, Bmp1 (CUB) domain and a C-terminal PDGF/vascular endothelial growth factor domain. PDGF CC and DD are secreted as full-length, latent dimers, and the proteolytic removal of the CUB domain is required for the growth factor domain of PDGF CC or DD to activate the PDGF receptors (4,20,21).PDGFs exert their biological functions through the activation of dimeric receptors made up of two structurally similar protein-tyrosine kinase receptor subunits (␣␣-, ␣␤-, or ␤␤-

Increasing evidence indicates the significance of platelet-derived growth factor receptor-b (b-PDGFR) signaling in prostate cancer (PCa). Accordingly, preclinical studies suggest the potential of b-PDGFR as a therapeutic target in metastatic PCa. However, a ligand responsible for b-PDGFR activation in PCa was unknown, and recent clinical trials with imatinib mesylate showed limited success due to normal tissue toxicity. Similarly, in spite of mounting evidence indicating the significance of matriptase in PCa, little is known about its substrates or molecular actions during PCa progression. Here, we identified PDGF-D as a ligand for b-PDGFR in PCa and discovered matriptase as its regulator. Matriptase activates PDGF-D by proteolytic removal of the CUB domain in a 2-step process, creating a hemidimer, followed by growth factor domain dimer (GFD-D) generation. Matriptase can deactivate PDGF-D by further proteolytic cleavage within the GFD, revealing its biphasic regulation. Importantly, PDGF-D/matriptase colocalization is accompanied with b-PDGFR phosphorylation in human PCa tissues. This study unveiled a novel signaling axis of matriptase/PDGF-D/b-PDGFR in PCa, providing new insights into functional interplay between serine protease and growth factor signaling networks.

Summary In tobacco and other Solanaceae species, the tobacco N gene confers resistance to tobacco mosaic virus (TMV), and leads to induction of standard defense and resistance responses. Here, we report the use of N‐transgenic tomato to identify a fast‐neutron mutant, sun1‐1 (suppressor of N), that is defective in N‐mediated resistance. Induction of salicylic acid (SA) and expression of pathogenesis‐related (PR) genes, each signatures of systemic acquired resistance, are both dramatically suppressed in sun1‐1 plants after TMV treatment compared to wild‐type plants. Application of exogenous SA restores PR gene expression, indicating that SUN1 acts upstream of SA. Upon challenge with additional pathogens, we found that the sun1‐1 mutation impairs resistance mediated by certain resistance (R) genes, (Bs4, I, and Ve), but not others (Mi‐1). In addition, sun1‐1 plants exhibit enhanced susceptibility to TMV, as well as to virulent pathogens. sun1‐1 has been identified as an EDS1 homolog present on chromosome 6 of tomato. The discovery of enhanced susceptibility in the sun1‐1 (Le_eds1‐1) mutant plant, which contrasts to reports in Nicotiana benthamiana using virus‐induced gene silencing, provides evidence that the intersection of R gene‐mediated pathways with general resistance pathways is conserved in a Solanaceous species. In tomato, EDS1 is important for mediating resistance to a broad range of pathogens (viral, bacterial, and fungal pathogens), yet shows specificity in the class of R genes that it affects (TIR‐NBS‐LRR as opposed to CC‐NBS‐LRR). In addition, a requirement for EDS1 for Ve‐mediated resistance in tomato exposes that the receptor‐like R gene class may also require EDS1.

Human adaptor protein, phosphotyrosine interaction, PH domain and leucine zipper containing 1 (APPL1) and adaptor protein, phosphotyrosine interaction, PH domain and leucine zipper containing 2 (APPL2) are homologous effectors of the small guanosine triphosphatase RAB5 that interact with a diverse set of receptors and signaling proteins and are proposed to function in endosome-mediated signaling. Herein, we investigated the membrane targeting properties of the APPL1 and APPL2 Bin/ Amphiphysin/Rvs (BAR), pleckstrin homology (PH) and phosphotyrosine binding (PTB) domains. Coimmunopre-cipitation and yeast two-hybrid studies demonstrated that full-length APPL proteins formed homooligomers and heterooligomers and that the APPL minimal BAR domains were necessary and sufficient for mediating APPL-APPL interactions. When fused to a fluorescent protein and overexpressed, all three domains (minimal BAR, PH and PTB) were targeted to cell membranes. Furthermore, full-length APPL proteins bound to phosphoinositides, and the APPL isolated PH or PTB domains were sufficient for in vitro phosphoinositide binding. Live cell imaging showed that full-length APPL-yellow fluorescent protein (YFP) fusion proteins associated with cytosolic membrane structures that underwent movement, fusion and fission events. Overexpression of full-length APPL-YFP fusion proteins was sufficient to recruit endogenous RAB5 to enlarged APPL-associated membrane structures, although APPL1 was not necessary for RAB5 membrane targeting. Taken together, our findings suggest a role for APPL proteins as dynamic scaffolds that modulate RAB5-associated signaling endosomal membranes by their ability to undergo domain-mediated oligomerization, membrane targeting and phosphoinositide binding. Human APPL1 (adaptor protein, phosphotyrosine interaction , PH domain and leucine zipper containing 1)/APPL (adaptor protein containing PH domain, PTB domain and Leucine zipper motif)/DIP13a (DCC-interacting protein 13a) and APPL2/DIP13b (referred to herein as APPL1 and APPL2, respectively) are two highly homologous proteins that contain three domains: an N-terminal Bin/Amphiphysin/ Rvs (BAR) domain, a central pleckstrin homology (PH) domain and a C-terminal phosphotyrosine binding (PTB) domain. APPL1 interacts with a diverse set of receptors, including the netrin-1 receptor DCC (deleted in colorectal carcinoma) (1), the nerve growth factor (NGF) receptor TrkA (2,3), the follicle-stimulating hormone (FSH) receptor (FSHR) (4,5) and the AdipoR1 and AdipoR2 adiponectin receptors (6,7). APPL1 also associates with signaling proteins, including AKT (v-akt murine thymoma viral oncogene homolog) proteins (5,8,9), phosphatidylinositol 3-kinase (PI3K) subu-nits (8) and the OCRL (oculocerebrorenal syndrome of Lowe) and INPP5B (inositol polyphosphate-5-phosphatase, 75 kDa) phosphatidylinositol 5-phosphatases (10). Many of these interactions are mediated by the APPL1 PTB domain, suggesting that APPL1 may function as an adaptor linked to distinct signaling pathways. APPL2 was originally iden...

The PDGF family members are potent mitogens for cells of mesenchymal origin and serve as important regulators of cell migration, survival, apoptosis, and transformation. Tumor-derived PDGF ligands are thought to function in both autocrine and paracrine manners, activating receptors on tumor and surrounding stromal cells. PDGF-C and -D are secreted as latent dimers, unlike PDGF-A and -B. Cleavage of the CUB domain from the PDGF-C and -D dimers is required for their biological activity. At present, little is known about the proteolytic processing of PDGF-C, the rate-limiting step in the regulation of PDGF-C activity. Here we show that the breast carcinoma cell line, MCF7, engineered to overexpress PDGF-C, produces proteases capable of cleaving PDGF-C to its active form. Increased PDGF-C expression enhances cell proliferation, anchorage independent cell growth, and tumor cell motility by autocrine signaling. In addition, MCF7-produced PDGF-C induces fibroblast cell migration in a paracrine manner. Interestingly, PDGF-C enhances tumor cell invasion in the presence of fibroblast, suggesting a role of tumor-derived PDGF-C in tumor-stromal interactions. In the present study, we identify tissue plasminogen activator (tPA) and matriptase as major proteases for processing of PDGF-C in MCF7 cells. In in vitro studies, we also show that urokinase plasminogen activator (uPA) is able to process PDGF-C. Furthermore, by site-directed mutagenesis, we identify the cleavage site for these proteases in PDGF-C. Lastly, we provide evidence suggesting a 2-step proteolytic processing of PDGF-C involving creation of a hemidimer, followed by growth factor domain dimer (GFD-D) generation.

Tag1 is a transposable element first identified as an insertion in the CHL1 gene of Arabidopsis. The chl1::Tag1 mutant originated from a plant (ecotype Landsberg erecta) that had been transformed with the maize transposon Activator (Ac), which is distantly related to Tag1. Genomic analysis of untransformed Landsberg erecta plants demonstrated that two identical Tag1 elements are present in the Landsberg erecta genome. To determine what provides transposase function for Tag1 transposition, we examined Tag1 excision in different genetic backgrounds. First, the chl1::Tag1 mutant was backcrossed to untransformed wild-type Arabidopsis plants to remove the Ac element(s) from the genome. F2 progeny that had no Ac elements but still retained Tag1 in the CHL1 gene were identified. Tag1 still excised in these Ac-minus progeny producing CHL1 revertants; therefore, Ac is not required for Tag1 excision. Next, Tag1 was inserted between a cauliflower mosaic virus 35S promoter and a beta-glucuronidase (GUS) marker gene and transformed into tobacco. Transformants showed blue-staining sectors indicative of Tag1 excision. Transgenic tobacco containing a defective Tag1 element, which was constructed in vitro by deleting an internal 1.4-kb EcoRI fragment, did not show blue-staining sectors. We conclude that Tag1 is an autonomous element capable of independent excision. The 35S-GUS::Tag1 construct was then introduced into Arabidopsis. Blue-staining sectors were found in cotyledons, leaves, and roots, showing that Tag1 undergoes somatic excision during vegetative development in its native host.