Kallmann De Morsier syndrome: FGF-signaling insufficiency?

Kallmann syndrome (KAL) associates hypogonadotropic hypogonadism and anosmia, i.e. a deficiency of the sense of smell. Anosmia is related to the absence or the hypoplasia of the olfactory bulbs. Hypogonadism is due to GnRH deficiency, and is likely to result from the failed embryonic migration of GnRH-synthesizing neurons. These cells normally migrate from the olfactory epithelium to the forebrain along the olfactory nerve pathway. Kallmann syndrome is genetically heterogeneous. The gene responsible for the X-chromosome linked form of the disease, KAL-1, has been identified in 1991. KAL1 encodes a ~95 kDa glycoprotein of unknown function, which is present locally in various extracellular matrices during the period of organogenesis. The recent finding that FGFR1 mutations are involved in an autosomal dominant form of Kallmann syndrome (KAL-2), combined to the analysis of mutant mouse embryos that no longer express Fgfr1 in the telencephalon, suggests that the disease results from a deficiency in FGF-signaling at the earliest stage of olfactory bulb morphogenesis. We propose that the role of the KAL1 gene product, the extracellular matrix protein anosmin-1, is to enhance FGF-signaling, and suggest that the gender difference in anosmin-1 dosage (because KAL1 partially escapes X-inactivation) explains the higher prevalence of the disease in males.     

A novel nonsense mutation of the KAL gene in two brothers with Kallmann syndrome

Kallmann syndrome (KS), defined by the association of hypogonadotropic hypogonadism and anosmia or hyposmia, can be caused by mutations in the KAL gene on Xp 22.3. This gene encodes an extracellular matrix glycoprotein called anosmin-1, which belongs to the class of cell adhesion molecules. In the absence of a functional KAL protein, migration of both olfactory and gonadotropin-releasing hormone neurons is arrested. A defective anosmin-1 molecule may also play a role in the development of synkinesia and renal agenesis, which are exclusively seen in the X-linked form of KS. We describe the clinical presentation and molecular diagnosis of the defect in two brothers with KS. An X-linked mode of transmission was assumed on the basis of synkinesia and the presence of oligomenorrhoea in the mother. A novel nonsense mutation was found in exon 13 of the KAL gene, encoding the region of the fourth fibronectin type III repeat of anosmin-1, which results in an apparently nonfunctional truncated protein.     

Transforming growth factor-β regulates the expression of anosmin (KAL-1) in human retinal pigment epithelial cells

In a microarray analysis of human retinal pigment epithelial cells (HRPE) treated with TGF-β, in addition to the alteration of a number of known Extracellular matrix (ECM)-related genes regulated by TGF-β, we found a significant increase in the expression of Kallmann Syndrome (KAL)-1 gene, that codes for the protein anosmin-1. Enhanced expression of KAL-1 by TGF-β was validated by real-time PCR analysis. In in vitro experiments, TGF-β receptor inhibitor abolished TGF-β-induced expression of KAL-1. Immunofluorescence staining showed increased presence of anosmin-1 in TGF-β treated HRPE cells, with distinct localization at the intercellular junctions. Treatment of HRPE cells with TGF-β enhanced secretion of anosmin-1 and the release of anosmin-1 was further augmented by heparin sulfate. Enhanced secretion of anosmin-1 in the presence of TGF-β and heparin was also observed in other ocular cells such as corneal epithelial and corneal fibroblast cultures. The role of anosmin-1, a protein with adhesion functions, in retinal structure, function and pathology has not been known and remains to be investigated.     

FGFR1 and anosmin-1 underlying genetically distinct forms of Kallmann syndrome are co-expressed and interact in olfactory bulbs

Kallmann syndrome is a genetically heterogeneous developmental disease characterised by a partial or complete lack of olfactory bulb development. Two genes underlying this disease have so far been identified: the KAL-1 gene, which encodes anosmin-1, an extracellular matrix protein that promotes axonal guidance and branch formation in vitro; and KAL-2, which encodes the known FGFR1. The implication of FGFR1 and anosmin-1 in the same developmental disease led us to test whether anosmin-1 and FGFR1 interact during the development of the olfactory system. In this paper, we showed that the two proteins co-localise in the olfactory bulb during development in rat. Using cross-immunoprecipitation assays of olfactory bulb extracts, we also demonstrated that anosmin-1 and FGFR1 are comprised within the same protein complex. Moreover, we show that anosmin-1 expression in CHO transfected cells increases FGFR1 accumulation, suggesting that anosmin-1 may act as a positive extracellular regulator of FGFR1 signalling. Taken together, our findings strongly suggest that anosmin-1 is an essential component of a FGFR1 pathway that plays a key role during olfactory bulb morphogenesis.     

Kallmann syndrome 1 gene is expressed in the marsupial gonad

Kallmann syndrome is characterized by hypogonadotrophic hypogonadism and anosmia. The syndrome can be caused by mutations in several genes, but the X-linked form is caused by mutation in the Kallmann syndrome 1 (KAL1). KAL1 plays a critical role in gonadotropin-releasing hormone (GnRH) neuronal migration that is essential for the normal development of the hypothalamic-pituitary-gonadal axis. Interestingly, KAL1 appears to be missing from the rodent X, and no orthologue has been detected as yet. We investigated KAL1 during development and in adults of an Australian marsupial, the tammar wallaby, Macropus eugenii. Marsupial KAL1 maps to an autosome within a group of genes that was added as a block to the X chromosome in eutherian evolution. KAL1 expression was widespread in embryonic and adult tissues. In the adult testis, tammar KAL1 mRNA and protein were detected in the germ cells at specific stages of differentiation. In the adult testis, the protein encoded by KAL1, anosmin-1, was restricted to the round spermatids and elongated spermatids. In the adult ovary, anosmin-1 was not only detected in the oocytes but was also localized in the granulosa cells throughout folliculogenesis. This is the first examination of KAL1 mRNA and protein localization in adult mammalian gonads. The protein localization suggests that KAL1 participates in gametogenesis not only through the development of the hypothalamic-pituitary-gonadal axis by activation of GnRH neuronal migration, but also directly within the gonads themselves. Because KAL1 is autosomal in marsupials but is X-linked in eutherians, its conserved involvement in gametogenesis supports the hypothesis that reproduction-related genes were actively recruited to the eutherian X chromosome.     

Anosmin-1 involved in neuronal cell migration is hypoxia inducible and cancer regulated

Functional expression of KAL1 gene is critical in the migration of GnRH neurons from the olfactory placode to the hypothalamus in embryogenesis. This gene thus far has not been shown to play a functional role in any other physiological or pathological process either in the developed brain or in peripheral tissues. We show here that KAL1 gene expression is decreased in early stage and increased in later stages of cancers. Screening of colon, lung and ovarian cancer cDNA panels indicated significant decrease in KAL1 expression in comparison to corresponding uninvolved tissues. However, KAL1 expression increased with the progression of cancer from early (I and II) stages to later (III and IV) stages of the cancer. There was a direct correlation between the TGF-β and KAL1 expression in colon cancer cDNA. Using colon cancer cell lines, we showed that TGF-β induces KAL1 gene expression and secretion of anosmin-1 protein (KAL1 coded protein). We further report that hypoxia induces anosmin-1 expression; anosmin-1 protects cancer cells from apoptosis activated by hypoxia and increases cancer cell mobility. Using siRNA technique we found that KAL1 expression following hypoxia is hypoxia-inducible factor (HIF-1) α dependent. Our results suggest that KAL1 gene expression plays an important role in cancer metastasis and protection from apoptosis.     

Mutation analyses in pedigrees and sporadic cases of ethnic Han Chinese Kallmann syndrome patients

Kallmann syndrome, a form of idiopathic hypogonadotropic hypogonadism, is characterized by developmental abnormalities of the reproductive system and abnormal olfaction. Despite association of certain genes with idiopathic hypogonadotropic hypogonadism, the genetic inheritance and expression are complex and incompletely known. In the present study, seven Kallmann syndrome pedigrees in an ethnic Han Chinese population were screened for genetic mutations. The exons and intron–exon boundaries of 19 idiopathic hypogonadotropic hypogonadism (idiopathic hypogonadotropic hypogonadism)-related genes in seven Chinese Kallmann syndrome pedigrees were sequenced. Detected mutations were also tested in 70 sporadic Kallmann syndrome cases and 200 Chinese healthy controls. In pedigrees 1, 2, and 7, the secondary sex characteristics were poorly developed and the patients’ sense of smell was severely or completely lost. We detected a genetic mutation in five of the seven pedigrees: homozygous KAL1 p.R191ter (pedigree 1); homozygous KAL1 p.C13ter (pedigree 2; a novel mutation); heterozygous FGFR1 p.R250W (pedigree 3); and homozygous PROKR2 p.Y113H (pedigrees 4 and 5). No genetic change of the assayed genes was detected in pedigrees 6 and 7. Among the 70 sporadic cases, we detected one homozygous and one heterozygous PROKR2 p.Y113H mutation. This mutation was also detected heterozygously in 2/200 normal controls and its pathogenicity is likely questionable. The genetics and genotype–phenotype relationships in Kallmann syndrome are complicated. Classical monogenic inheritance does not explain the full range of genetic inheritance of Kallmann syndrome patients. Because of stochastic nature of genetic mutations, exome analyses of Kallmann syndrome patients may provide novel insights.     

Kallmann syndrome: mutations in the genes encoding prokineticin-2 and prokineticin receptor-2

Kallmann syndrome combines anosmia, related to defective olfactory bulb morphogenesis, and hypogonadism due to gonadotropin-releasing hormone deficiency. Loss-of-function mutations in KAL1 and FGFR1 underlie the X chromosome-linked form and an autosomal dominant form of the disease, respectively. Mutations in these genes, however, only account for approximately 20% of all Kallmann syndrome cases. In a cohort of 192 patients we took a candidate gene strategy and identified ten and four different point mutations in the genes encoding the G protein-coupled prokineticin receptor-2 (PROKR2) and one of its ligands, prokineticin-2 (PROK2), respectively. The mutations in PROK2 were detected in the heterozygous state, whereas PROKR2 mutations were found in the heterozygous, homozygous, or compound heterozygous state. In addition, one of the patients heterozygous for a PROKR2 mutation was also carrying a missense mutation in KAL1, thus indicating a possible digenic inheritance of the disease in this individual. These findings reveal that insufficient prokineticin-signaling through PROKR2 leads to abnormal development of the olfactory system and reproductive axis in man. They also shed new light on the complex genetic transmission of Kallmann syndrome.     

Localization of anosmin-1a and anosmin-1b in the inner ear and neuromasts of zebrafish

Anosmin-1, encoded by the KAL-1 gene, is the protein defective in the X-linked form of Kallmann syndrome. This human developmental disorder is characterized by defects in cell migration and axon target selection. Anosmin-1 is an extracellular matrix protein that plays a role, in vitro, in processes such as cell adhesion, neurite outgrowth, axon guidance, and axon branching. The zebrafish possesses two orthologues of the KAL-1 gene: kal1a and kal1b, which encode anosmin-1a and anosmin-1b, respectively. Previous in situ hybridization studies have shown that kal1a and kal1b mRNAs are expressed in undetermined cells of the inner ear but not in neuromast cells. Using specific antibodies against anosmin-1a and anosmin-1b, we report here that both proteins are expressed in sensory hair cells of the inner ear cristae ampullaris and the lateral line neuromasts. Accumulation of these proteins was observed mainly at the level of the hair bundle and also at the cell membrane. In neuromast hair cells, immunogold scanning electronmicroscopy demonstrated that anosmin-1a and anosmin-1b were present at the surface of the stereociliary bundle. In addition, anosmin-1a, but not anosmin-1b, was detected on the track of the ampullary nerve. This is the first report of anosmin-1 expression in sensory hair cells of the inner ear and lateral line, and along the ampullary nerve track.     

Extended and flexible domain solution structure of the extracellular matrix protein anosmin-1 by X-ray scattering, analytical ultracentrifugation and constrained modelling

Kallmann's syndrome corresponds to a loss of sense of smell and hypogonadotrophic hypogonadism. Defects in anosmin-1 result in the X-linked inherited form of Kallmann's syndrome. Anosmin-1 is an extracellular matrix protein comprised of an N-terminal, cysteine-rich (Cys-box) domain and a whey acidic protein-like (WAP) domain, followed by four fibronectin type III (FnIII) domains. The solution structures of recombinant proteins containing the first three domains (PIWF1) and all six domains (PIWF4) were determined by X-ray scattering and analytical ultracentrifugation. Guinier analyses showed that PIWF1 and PIWF4 have different radii of gyration (R(G)) values of 3.1 nm and 6.7 nm, respectively, but similar cross-sectional radii of gyration (R(XS)) values of 1.5 nm and 1.9 nm, respectively. Distance distribution functions showed that the maximum lengths of PIWF1 and PIWF4 were 11 nm and 23 nm, respectively. Analytical ultracentrifugation gave sedimentation coefficients of 2.52 S and 3.55 S for PIWF1 and PIWF4, respectively. The interpretation of the scattering data by constrained modelling requires homology models for all six domains in anosmin-1. While models were already available for the WAP and FnIII domains, searches suggested the Cys-box domain may resemble the cysteine-rich region of the insulin-like growth factor receptor. Automated constrained molecular modelling based on joining the anosmin-1 domains with structurally randomised linkers resulted in 10,000 models for anosmin-1. A trial-and-error search showed that about 0.1-1.4% of these models fitted the X-ray data. The best models showed that the three domains and six domains in PIWF1 and PIWF4, respectively, were extended. The inter-domain linkers in anosmin-1 could not all be extended at the same time, and there was evidence for inter-domain flexibility. Models with folded-back domain arrangements do not fit the data. These solution structures account for the known biological function of anosmin-1, in particular its ability to interact with its three macromolecular ligands.     

Caenorhabditis elegans BAH-1 Is a DUF23 Protein Expressed in Seam Cells and Required for Microbial Biofilm Binding to the Cuticle

The cuticle of Caenorhabditis elegans, a complex, multi-layered extracellular matrix, is a major interface between the animal and its environment. Biofilms produced by the bacterial genus Yersinia attach to the cuticle of the worm, providing an assay for surface characteristics. A C. elegans gene required for biofilm attachment, bah-1, encodes a protein containing the domain of unknown function DUF23. The DUF23 domain is found in 61 predicted proteins in C. elegans, which can be divided into three distinct phylogenetic clades. bah-1 is expressed in seam cells, which are among the hypodermal cells that synthesize the cuticle, and is regulated by a TGF-β signaling pathway.     

Novel Mechanisms of Fibroblast Growth Factor Receptor 1 Regulation by Extracellular Matrix Protein Anosmin-1

Activation of fibroblast growth factor (FGF) signaling is initiated by a multiprotein complex formation between FGF, FGF receptor (FGFR), and heparan sulfate proteoglycan on the cell membrane. Cross-talk with other factors could affect this complex assembly and modulate the biological response of cells to FGF. We have previously demonstrated that anosmin-1, a glycosylated extracellular matrix protein, interacts with the FGFR1 signaling complex and enhances its activity in an IIIc isoform-specific and HS-dependent manner. The molecular mechanism of anosmin-1 action on FGFR1 signaling, however, remains unknown. Here, we show that anosmin-1 directly binds to FGFR1 with high affinity. This interaction involves domains in the N terminus of anosmin-1 (cysteine-rich region, whey acidic protein-like domain and the first fibronectin type III domain) and the D2–D3 extracellular domains of FGFR1. In contrast, anosmin-1 binds to FGFR2IIIc with much lower affinity and displays negligible binding to FGFR3IIIc. We also show that FGFR1-bound anosmin-1, although capable of binding to FGF2 alone, cannot bind to a FGF2·heparin complex, thus preventing FGFR1·FGF2·heparin complex formation. By contrast, heparin-bound anosmin-1 binds to pre-formed FGF2·FGFR1 complex, generating an anosmin-1·FGFR1·FGF2·heparin complex. Furthermore, a functional interaction between anosmin-1 and the FGFR1 signaling complex is demonstrated by immunofluorescence co-localization and Transwell migration assays where anosmin-1 was shown to induce opposing effects during chemotaxis of human neuronal cells. Our study provides molecular and cellular evidence for a modulatory action of anosmin-1 on FGFR1 signaling, whereby binding of anosmin-1 to FGFR1 and heparin can play a dual role in assembly and activity of the ternary FGFR1·FGF2·heparin complex.FGF⁵ signaling plays an important role in a wide range of fundamental biological responses (1–3). Both FGF and FGFR bind to heparan sulfate (HS) and heparin, a highly sulfated type of HS produced in connective tissue mast cells. Heparan sulfate proteoglycans (HSPG) are the cell surface co-receptors essential for the formation of functional FGF·FGFR signaling complex (4, 5). There are four structurally related FGFRs (FGFR1–4), which consist of an extracellular ligand-binding region containing three immunoglobulin (Ig)-like domains (D1–D3), a single transmembrane domain, and a cytoplasmic domain with protein-tyrosine kinase catalytic activity. The 22 members of the FGF family bind to the interface formed by the D2/D3 domains and the linker between these domains (6, 7), whereas a conserved positively charged region in D2 serves as the HS binding site (8). An unusual stretch of seven to eight acidic residues designated as the “acid box” is present in the linker connecting D1 and D2. Alternative splicing events occur to generate various isoforms, including a truncated receptor lacking D1 and the D1–D2 linker or a full-length receptor that differs in the second half of D3, designated as IIIb and IIIc isoforms (5). Two crystal structures have been proposed to demonstrate how the FGF·FGFR·heparin complex is assembled (9, 10). Recent evidence suggests that both may be biologically relevant (11, 12).The diversity of FGF signaling pathways and consequent biological functions require that activation of FGFR should be tightly regulated. Such regulation can occur either at the level of the extracellular receptor-ligand complex assembly or via intracellular modulation of downstream effectors (13). Extracellular regulation mainly involves the interaction between each component of the FGF·FGFR·HS signaling complex. For example, FGF8 is shown to bind mostly to the FGFR IIIc isoforms, whereas FGF7 acts as the preferential ligand for the FGFR2 IIIb isoform (13, 14). Sequence specificity, length, and sulfation patterns of HS are also important regulators of the FGF·FGFR interaction (15, 16).Cell surface proteins other than FGFs and HSPGs participate in FGFR signaling regulation. FLRT3 (a member of the fibronectin-leucine-rich transmembrane protein family) promotes FGF signaling and interacts with FGFR1 and FGFR4 via its extracellular fibronectin type III (FnIII) domain (17). Sef (similar expression to fgf genes) functions as an antagonist of FGF signaling in zebrafish. The two FnIII regions of Sef are essential for its function and interaction with FGFR1 and FGFR2 (18). Neuronal cell adhesion molecule (NCAM), N-cadherin, and L1 have also been identified as functionally relevant in FGFR-mediated neurite outgrowth (19–22). The FnIII domains of NCAM bind to the D2 and D3 domains of FGFR1 (19) and FGFR2 (23) to induce ligand-independent receptor phosphorylation.Anosmin-1, an extracellular matrix-associated glycosylated protein, appears to be a novel member of the extracellular FGFR signaling modulators (24, 25). Loss-of-function mutations of anosmin-1 and FGFR1 are associated with Kallmann syndrome (KS), underlying X-linked, and autosomal dominant/recessive inheritance mode, respectively (26–28). KS is a human developmental genetic disorder characterized by loss of sense of smell (anosmia) caused by abnormal olfactory bulb development and delayed, even arrested puberty caused by disrupted migration of the gonadotropin-releasing hormone (GnRH)-secreting neuron. We previously reported that anosmin-1 acts as an FGFR1IIIc isoform-specific co-ligand, which enhances signaling activity. In human embryonic GnRH olfactory neuroblast FNC-B4 cells, anosmin-1 induced neurite outgrowth and cytoskeletal rearrangements through FGFR1-dependent mechanisms involving p42/44 and p38 mitogen-activated protein kinases and Cdc42/Rac1 activation (25). A functional interaction is also demonstrable between anosmin-1 and FGFR1 in optic nerve oligodendrocyte precursor development (24). Structurally, anosmin-1 comprises an N-terminal cysteine-rich domain (CR) and a whey acidic protein-like (WAP) domain, followed by four tandem FnIII repeats and a C-terminal histidine rich region (Fig. 1a). Current evidence suggests that anosmin-1 functions by affecting FGF2-induced activation of FGFR1 signaling rather than by directly stimulating the receptor. However, the precise molecular mechanism of this interaction remains unclear.Open in a separate windowFIGURE 1.Generation of recombinant anosmin-1, anosmin-1 mutants, FGFR1D1D3, and FGFR1D2D3 proteins. a, the schematic structures of recombinant proteins of anosmin-1 and FGFR1. Each domain in the wild type (PIWF4), point mutants (mPIWF4^N267K, mPIWF4^E514K, and mPIWF4^F517L), and truncated (PIWF1, PIWF2, and PIF4) anosmin-1 protein analogues are represented by a shaded rectangle. V5 and 6His epitopes at the C terminus are represented by a clear rectangle. Each immunoglobulin-like domain in the full ectodomain (FGFR1D1D3) and truncated form (FGFR1D2D3) of FGFR1 is represented by a half circle. The acid box (AB) is represented by a filled rectangle. H, histidine-rich region. b, 0.5–1 μg of purified recombinant proteins are loaded in each lane and visualized by colloidal blue staining. Molecular mass markers in kilodaltons are shown on the left.We now report for the first time that anosmin-1 directly binds to FGFR1 using surface plasmon resonance (SPR), chemical cross-linking, and immunofluorescence co-localization studies in living cells. This interaction occurs between the N-terminal CR, WAP, and the first FnIII domain of anosmin-1 and D2 and D3 ectodomains of FGFR1. Moreover, SPR studies using sequential injections and Transwell migration assays in immortalized FNC-B4-hTERT cells suggest that anosmin-1 can have opposing effects in the formation and activation of the FGF2·FGFR1·heparin complex depending on the order of their binding interactions with anosmin-1.     

The secreted Alzheimer-related amyloid precursor protein fragment has an essential role in C. elegans

Mutations in the gene encoding the amyloid precursor protein (APP) or the enzymes that process APP are correlated with familial Alzheimer disease. Alzheimer disease is also associated with insulin resistance (type 2 diabetes). In our recently published study,¹ we obtained genetic evidence that the extracellular fragment of APL-1, the C. elegans ortholog of human APP, may act as a signaling molecule to modulate insulin and nuclear hormone pathways in C. elegans development. In addition, independent of insulin and nuclear hormone signaling, high levels of the extracellular fragment of APL-1 (sAPL-1) leads to a temperature-sensitive embryonic lethality, which is dependent on activity of a predicted receptor protein tyrosine phosphatase (MOA-1/R155.2). Furthermore, this embryonic lethality is enhanced by knockdown of a predicted prion-like protein (pqn-29). The precise molecular mechanisms underlying these processes remain to be determined. Here, we present hypothetical models as to how sAPL-1 signaling influences metabolic and developmental pathways. Together, with previous findings in mammals that the extracellular domain of mammalian APP (sAPP) binds to a death-receptor,² our findings support the model that sAPP signaling affects critical biological processes.     

The purine nucleoside phosphorylase pnp-1 regulates epithelial cell resistance to infection in C. elegans

Intestinal epithelial cells are subject to attack by a diverse array of microbes, including intracellular as well as extracellular pathogens. While defense in epithelial cells can be triggered by pattern recognition receptor-mediated detection of microbe-associated molecular patterns, there is much to be learned about how they sense infection via perturbations of host physiology, which often occur during infection. A recently described host defense response in the nematode C. elegans called the Intracellular Pathogen Response (IPR) can be triggered by infection with diverse natural intracellular pathogens, as well as by perturbations to protein homeostasis. From a forward genetic screen, we identified the C. elegans ortholog of purine nucleoside phosphorylase pnp-1 as a negative regulator of IPR gene expression, as well as a negative regulator of genes induced by extracellular pathogens. Accordingly, pnp-1 mutants have resistance to both intracellular and extracellular pathogens. Metabolomics analysis indicates that C. elegans pnp-1 likely has enzymatic activity similar to its human ortholog, serving to convert purine nucleosides into free bases. Classic genetic studies have shown how mutations in human purine nucleoside phosphorylase cause immunodeficiency due to T-cell dysfunction. Here we show that C. elegans pnp-1 acts in intestinal epithelial cells to regulate defense. Altogether, these results indicate that perturbations in purine metabolism are likely monitored as a cue to promote defense against epithelial infection in the nematode C. elegans.     

Anosmin-1, defective in the X-linked form of Kallmann syndrome,promotes axonal branch formation from olfactory bulb output neurons

The physiological role of anosmin-1, defective in the X chromosome-linked form of Kallmann syndrome, is not yet known. Here, we show that anti-anosmin-1 antibodies block the formation of the collateral branches of rat olfactory bulb output neurons (mitral and tufted cells) in organotypic cultures. Moreover, anosmin-1 greatly enhances axonal branching of these dissociated neurons in culture. In addition, coculture experiments with either piriform cortex or anosmin-1-producing CHO cells demonstrate that anosmin-1 is a chemoattractant for the axons of these neurons, suggesting that this protein, which is expressed in the piriform cortex, attracts their collateral branches in vivo. We conclude that anosmin-1 has a dual branch-promoting and guidance activity, which plays an essential role in the patterning of mitral and tufted cell axon collaterals to the olfactory cortex.     

Functions of the extracellular matrix in development: Lessons from Caenorhabditis elegans

Cell-extracellular matrix interactions are crucial for the development of an organism from the earliest stages of embryogenesis. The main constituents of the extracellular matrix are collagens, laminins, proteoglycans and glycosaminoglycans that form a network of interactions. The extracellular matrix and its associated molecules provide developmental cues and structural support from the outside of cells during development. The complex nature of the extracellular matrix and its ability for continuous remodeling poses challenges when investigating extracellular matrix-based signaling during development. One way to address these challenges is to employ invertebrate models such as Caenorhabditis elegans, which are easy to genetically manipulate and have an invariant developmental program. C. elegans also expresses fewer extracellular matrix protein isoforms and exhibits reduced redundancy compared to mammalian models, thus providing a simpler platform for exploring development. This review summarizes our current understanding of how the extracellular matrix controls the development of neurons, muscles and the germline in C. elegans.