Reduction of BDNF expression in Fmr1 knockout mice worsens cognitive deficits but improves hyperactivity and sensorimotor deficits.

Fragile X syndrome (FXS) is a common cause of inherited intellectual disability and a well-characterized form of autism spectrum disorder. As brain-derived neurotrophic factor (BDNF) is implicated in the pathophysiology of FXS we examined the effects of reduced BDNF expression on the behavioral phenotype of an animal model of FXS, Fmr1 knockout (KO) mice, crossed with mice carrying a deletion of one copy of the Bdnf gene (Bdnf(+/-)). Fmr1 KO mice showed age-dependent alterations in hippocampal BDNF expression that declined after the age of 4 months compared to wild-type controls. Mild deficits in water maze learning in Bdnf(+/-) and Fmr1 KO mice were exaggerated and contextual fear learning significantly impaired in double transgenics. Reduced BDNF expression did not alter basal nociceptive responses or central hypersensitivity in Fmr1 KO mice. Paradoxically, the locomotor hyperactivity and deficits in sensorimotor learning and startle responses characteristic of Fmr1 KO mice were ameliorated by reducing BNDF, suggesting changes in simultaneously and in parallel working hippocampus-dependent and striatum-dependent systems. Furthermore, the obesity normally seen in Bdnf(+/-) mice was eliminated by the absence of fragile X mental retardation protein 1 (FMRP). Reduced BDNF decreased the survival of newborn cells in the ventral part of the hippocampus both in the presence and absence of FMRP. Since a short neurite phenotype characteristic of newborn cells lacking FMRP was not found in cells derived from double mutant mice, changes in neuronal maturation likely contributed to the behavioral phenotype. Our results show that the absence of FMRP modifies the diverse effects of BDNF on the FXS phenotype.

Chromosomal location, exon structure, and vascular expression patterns of the human PDGFC and PDGFD genes.

BACKGROUND: Platelet-derived growth factor (PDGF), which is a major mitogen for vascular smooth muscle cells and has been implicated in the pathogenesis of arteriosclerosis, is composed of dimers of PDGF-A and PDGF-B polypeptide chains, encoded by different genes. Here, we have analyzed the chromosomal localization, structure, and expression of 2 newly identified human genes of the PDGF family, called PDGFC and PDGFD. METHODS AND RESULTS: We used fluorescence in situ hybridization to locate PDGFC and PDGFD in chromosomes 4q32 and 11q22.3 to 23.2, respectively. Exon structures of PDGFC and PDGFD were determined by sequencing from genomic DNA clones. The coding region of PDGFC consists of 6 and PDGFD of 7 exons, of which the last 2 encode the C-terminal PDGF cystine knot growth factor homology domain. An N-terminal CUB domain is encoded by exons 2 and 3 of both genes, and a region of proteolytic cleavage involved in releasing and activating the growth factor domain is located in exon 4 in PDGFC and exon 5 in PDGFD. PDGF-C was expressed predominantly in smooth muscle cells and PDGF-D in fibroblastic adventitial cells, and both genes were active in cultured endothelial cells and in a variety of tumor cell lines. Both PDGF-C and PDGF-D also stimulated human coronary artery smooth muscle cells. CONCLUSIONS: PDGFC and PDGFD have similar genomic structures, which resemble those of the PDGFA and PDGFB genes. Their expression in the arterial wall and cultured vascular cells suggests that they can transduce proliferation/migration signals to pericytes and smooth muscle cells.

PDGF-D is a specific, protease-activated ligand for the PDGF beta-receptor.

The term 'platelet-derived growth factor' (PDGF) refers to a family of disulphide-bonded dimeric isoforms that are important for growth, survival and function in several types of connective tissue cell. So far, three different PDGF chains have been identified - the classical PDGF-A and PDGF-B and the recently identified PDGF-C. PDGF isoforms (PDGF-AA, AB, BB and CC) exert their cellular effects by differential binding to two receptor tyrosine kinases. The PDGF alpha-receptor (PDGFR-alpha) binds to all three PDGF chains, whereas the beta-receptor (PDGFR-beta) binds only to PDGF-B. Gene-targeting studies using mice have shown that the genes for PDGF-A and PDGF-B, as well as the two PDGFR genes, are essential for normal development. Furthermore, overexpression of PDGFs is linked to different pathological conditions, including malignancies, atherosclerosis and fibroproliferative diseases. Here we have identify and characterize a fourth member of the PDGF family, PDGF-D. PDGF-D has a two-domain structure similar to PDGF-C and is secreted as a disulphide-linked homodimer, PDGF-DD. Upon limited proteolysis, PDGF-DD is activated and becomes a specific agonistic ligand for PDGFR-beta. PDGF-DD is the first known PDGFR-beta-specific ligand, and its unique receptor specificity indicates that it may be important for development and pathophysiology in several organs.

VEGFR3 gene structure, regulatory region, and sequence polymorphisms.

Vascular endothelial growth factor receptor 3 (VEGFR-3) is required for cardiovascular development during embryogenesis. In adults, this receptor is expressed in lymphatic endothelial cells, and mutant VEGFR3 alleles have been implicated in human hereditary lymphedema. To better understand the basis of its specific endothelial lineage-restricted expression, we have characterized the VEGFR3 gene and its regulatory 5' flanking region. The human gene contains 31 exons, of which exons 30a and 30b are alternatively spliced. The VEGFR3 proximal promoter is TATA-less and contains stretches of sequences homologous with the mouse Vegfr3 promoter region. In transfection experiments of cultured cells, the Vegfr3 promoter was shown to control endothelial cell-specific transcription of downstream reporter genes. This result was further confirmed in vivo; in a subset of transgenic mouse embryos, a 1.6 kb Vegfr3 promoter fragment directed weak lymphatic endothelial expression of the LacZ marker gene. This suggests that endothelial cell-specific elements occur in the proximal promoter, although further enhancer elements are probably located elsewhere. The sequence, organization, and variation in the VEGFR3 gene and its regulatory region provide important tools for the molecular genetic analysis of the lymphatic system and its disorders.

Expression and glycosylation studies of human FGF receptor 4.

Fibroblast growth factor receptor subtype 4 (FGFR4) has been shown to have special activation properties and just one splicing form, unlike the other FGFRs. FGFR4 overexpression is correlated with breast cancer and therefore FGFR4 is a target for drug design. Our aim is to overexpress high amounts of homogeneous FGFR4 extracellular domain (FGFR4(ed)) for structural studies. We show that baculovirus-insect cell-expressed FGFR4(ed) is glycosylated on three (N88, N234, and N266) of the six possible N-glycosylation sites but is not O-glycosylated. The deglycosylated triple mutant was expressed and had binding properties similar to those of glycosylated FGFR4(ed), but was still heterogeneous. Large amounts of FGFR4(ed) have been produced into inclusion bodies in Escherichia coli and refolded at least partly correctly but the refolded E. coli-produced FGFR4(ed) still aggregates.

PDGF-C is a new protease-activated ligand for the PDGF alpha-receptor.

Platelet-derived growth factors (PDGFs) are important in many types of mesenchymal cell. Here we identify a new PDGF, PDGF-C, which binds to and activates the PDGF alpha-receptor. PDGF-C is activated by proteolysis and induces proliferation of fibroblasts when overexpressed in transgenic mice. In situ hybridization analysis in the murine embryonic kidney shows preferential expression of PDGF-C messenger RNA in the metanephric mesenchyme during epithelial conversion. Analysis of kidneys lacking the PDGF alpha-receptor shows selective loss of mesenchymal cells adjacent to sites of expression of PDGF-C mRNA; this is not found in kidneys from animals lacking PDGF-A or both PDGF-A and PDGF-B, indicating that PDGF-C may have a unique function.