Review ArticleNeuropilins in neoplasms: Expression, regulation, and function☆
Introduction
Neuropilin (NRP) is a 130- to 140-kDa transmembrane glycoprotein. Fujisawa and colleges first identified NRP in 1987 as the antigen to a monoclonal antibody named A5 that specifically bound to the superficial neuropile in the Xenopus laevis optic tectum [1]. Subsequently, NRP was found to be a receptor for the class 3 semaphorins (SEMA3), a family of chemorepulsive guidance molecules capable of collapsing axonal growth cones and repelling axons of ganglia. NRP2 was discovered in 1997 [2], [3]. The two neuropilin genes, NRP1 and NRP2, share 45% homology and map to different chromosomes [4]. In humans, NRP1 is located on chromosome 10, and NRP2 is located on chromosome 2 [4]. Both genes are composed of 17 exons, but NRP2 is expressed as several alternatively spliced transcripts [4]. Both NRPs have similar overall structure but differ significantly in their expression patterns, regulation, and ligand binding specificities. The extracellular regions of NRPs are composed of separate subdomains: the a1a2 domain is involved in SEMA3-binding, the b1b2 domain is involved in both SEMA3- and vascular endothelial growth factor (VEGF)-binding (discussed in more detail below), and the c domain is involved in dimerization [1], [5]. The cytoplasmic domain of either NRP is relatively short, consisting of only 40 amino acids, and contains no known signaling motif. NRP1 and NRP2 sequences end in the same three C-terminal amino acids, SEA. This region has been shown to bind to GIPC (GAIP interacting protein at the C terminus) [6]. GIPC, which is also called NIP (neuropilin interacting protein), is a 40 kDa protein containing a PDZ domain that interacts with RGS-GAIP, a GTPase-activating protein (GAP) for Gαi subunits. The signaling function of GIPC in response to NRP ligands has not been elucidated [6]. In addition to the transmembrane forms, neuropilins also exist as soluble isoforms containing only the extracellular ligand binding a1a2 and b1b2 subdomains but lacking the c, transmembrane, and cytoplasmic domains [4], [7]. Premature truncation within introns results in these smaller isoforms that contain C-terminal intron-derived amino acids. To date, four soluble neuropilin-1 (sNRP1) species and one sNRP2 species have been reported [8], [9]. These sNRPs can function as competitive antagonists for NRP ligands, for example, VEGF [5], [7].
NRPs are receptors for two different ligand families, the SEMA family of axonal guidance regulators [3] and the VEGF family of angiogenesis factors [4]. The semaphorins are divided into seven classes consisting of transmembrane proteins, glycosylphosphatidylinosotol (GIP)-linked proteins, as well as soluble proteins (class 3). Semaphorin 3A (SEMA3A, human protein; Sema3A, murine or chick protein; originally called collapsin-1) [10] was the first NRP1 ligand identified [3], [11]. The class 3 semaphorin family is comprised of 6 secreted proteins, SEMA3A, B, C, D, E, and F. Although the ligand binding domains of NRP1 and NRP2 are similar, there is a degree of specificity in SEMA/NRP binding and activity. SEMA3A binds NRP1; SEMA3F binds NRP2; whereas SEMA3B binds both NRP1 and NRP2 [12]. Sensory neurons such as dorsal root ganglia (DRG) express NRP1 and bind SEMA3A, resulting in functional collapse of axonal growth cones [1]. SEMA3F binds NRP2 and repels sympathetic neurons such as superior cervical ganglia (SCG) [13], [25]. The “sema”-domain of Sema3A binds to the a1a2 domain of NRP1, and the carboxy-terminal C-domain of Sema3A binds the b1b2 domain of NRP1 [14]. The Sema3A protein dimerizes via a cysteine at residue 723, between the Ig domain and basic tail. Only dimerized Sema3A protein can functionally collapse axonal growth cones [15]. The sema-domain alone is inactive since it cannot dimerize. In the neuronal guidance pathway, NRPs act as co-receptors with plexins, transmembrane receptors that transduce the semaphorin signal [16]. Recently, Plexin-A4 has been found to signal via NRP1, while Plexin-A3 signals via NRP2 [17].
In 1998, NRP1 was identified as a functional receptor for VEGF165 [18]. Exon 7 of VEGF165 binds to the b1b2 domain of NRP1. VEGF121 has no exon 7-domain and therefore cannot bind NRP1 [18], [19]. Both NRP1 and NRP2 bind VEGF165, but differ in their binding affinities to other family members. For example, NRP1 binds VEGF-B, VEGF-E, and placenta growth factor-2 (PlGF2, a splice isoform of PlGF), whereas NRP2 binds VEGF145 and VEGF-C [4]. In the angiogenesis pathway, NRP1 functions as a co-receptor with VEGFR-2 that increases the binding of VEGF165 to VEGFR-2 and that enhancesVEGF165-mediated chemotaxis of EC [18], [20]. NRP1 also regulates VEGF-mediated permeability in cultured EC [21]. Transgenic mice overexpressing sNRP1 in the skin have decreased vascular permeability [22]. VEGF and SEMA3A have overlapping binding domains on NRP1 and thus are competitive inhibitors of one another [4]. The fact that these two disparate ligand families can bind to the same receptor and yet mediate two different processes, neuronal guidance and angiogenesis, suggests common molecular mechanisms in these processes. These results have sparked new areas of investigation and raise the question of whether other neuronal guidance mediators may affect angiogenesis [23], [24].
Section snippets
Neuropilin expression patterns
Although NRPs initially received their name from their neuronal localization, it is now known that NRPs are expressed in adult human organs [18] by a variety of tissue types [7]. NRP1 and NRP2 have specific and overlapping expression patterns in several tissues. For example, both sensory and sympathetic neurons express NRP1, making them susceptible to the repulsive effects of SEMA3A, whereas NRP2 is found only on sympathetic, but not on sensory neurons [25]. As a consequence, only sympathetic
Concluding remarks
In summary, semaphorins and neuropilins play a dual role as mediators of both neuronal guidance and angiogenesis. The regulation of NRP1 and NRP2 is complex and independently mediated. NRP1 and NRP2 are expressed in different tissues in a largely non-overlapping pattern. Carcinomas typically overexpress NRP1, whereas neuronal tumors largely express NRP2. NRPs can promote angiogenesis through their VEGF ligands or inhibit angiogenesis through their SEMA ligands. The NRP expression pattern in
Acknowledgments
This work was supported by NIH grants CA37392 and CA45548 (M. Klagsbrun); the SPORE in Skin Cancer Developmental Project Award (NIH, Department of Dermatology, Brigham and Women's Hospital, Boston, MA) (D. R. Bielenberg) and Grants-in-Aid for Scientific Research from the Japanese Ministry of Education, Culture, Sports, Science, and Technology (S. Takashima). We thank Dr. Akio Shimizu for PCR primers and Dr. Jay Harper for the murine ISH probe. We thank Dr. Isaiah Fidler for providing cell
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