Site-specific HNK-1 epitope on alternatively spliced fibronectin type-III repeats in tenascin-C promotes neurite outgrowth of hippocampal neurons through contactin-1.

The human natural killer-1 (HNK-1) carbohydrate epitope, composed of a unique sulfated trisaccharide (HSO3-3GlcAß1-3Galß1-4GlcNAc-R), is highly expressed during brain development and regulates higher brain function. However, it remains unclear which glycoprotein carries the HNK-1 epitope in the embryonic brain and the functional role it plays. Here, we showed that one of the major HNK-1 carrier proteins in the embryonic brain is tenascin-C (TNC), an extracellular matrix protein that regulates neurite outgrowth by interacting with the GPI-anchored protein contactin-1 (CNTN). Because the alternatively spliced fibronectin type-III (FNIII) repeats in TNC give rise to many isoforms and affect neuronal function, we evaluated neurite outgrowth of primary hippocampal neurons on purified recombinant FNIII repeats with or without the HNK-1 epitope as a substrate. We found that the presence of the HNK-1 epitope on the C domain of TNC promoted neurite outgrowth, and that this signal was mediated by CNTN, which is an HNK-1-expressing neuronal receptor. The neurite-promoting activity of the HNK-1 epitope on TNC required neuronal HNK-1 expression, which was defective in neurons lacking the glucuronyltransferases GlcAT-P and GlcAT-S. These results suggest that the HNK-1 epitope is a key modifier of TNC and CNTN in the regulation of embryonic brain development.

PLP1 and CNTN1 gene variation modulates the microstructure of human white matter in the corpus callosum.

The corpus callosum is the brain's largest commissural fiber tract and is crucial for interhemispheric integration of neural information. Despite the high relevance of the corpus callosum for several cognitive systems, the molecular determinants of callosal microstructure are largely unknown. Recently, it was shown that genetic variations in the myelin-related proteolipid 1 gene PLP1 and the axon guidance related contactin 1 gene CNTN1 were associated with differences in interhemispheric integration at the behavioral level. Here, we used an innovative new diffusion neuroimaging technique called neurite orientation dispersion and density imaging (NODDI) to quantify axonal morphology in subsections of the corpus callosum and link them to genetic variation in PLP1 and CNTN1. In a cohort of 263 healthy human adults, we found that polymorphisms in both PLP1 and CNTN1 were significantly associated with callosal microstructure. Importantly, we found a double dissociation between gene function and neuroimaging variables. Our results suggest that genetic variation in the myelin-related gene PLP1 impacts white matter microstructure in the corpus callosum, possibly by affecting myelin structure. In contrast, genetic variation in the axon guidance related gene CNTN1 impacts axon density in the corpus callosum. These findings suggest that PLP1 and CNTN1 gene variations modulate specific aspects of callosal microstructure that are in line with their gene function.

Role of F3/contactin expression profile in synaptic plasticity and memory in aged mice.

We have recently shown that overexpression of the F3/contactin adhesive glycoprotein (also known as Contactin-1) promotes neurogenesis in adult hippocampus, which correlates with improved synaptic plasticity and memory. Because F3/contactin levels physiologically decrease with age, here, we aim at investigating whether its overexpression might counteract the cognitive decline in aged animals. For this we use 20- to 24-month-old TAG/F3 transgenic mice in which F3/contactin overexpression is driven by regulatory sequences from the gene encoding the transient axonal glycoprotein TAG-1 throughout development. We show that aged TAG/F3 mice display improved hippocampal long-term potentiation and memory compared with wild-type littermates. The same mice undergo a decrease of neuronal apoptosis at the hippocampal level, which correlated to a decrease of active caspase-3; by contrast, procaspase-3 and Bax as well as the anti-apoptotic and plasticity-related pathway BDNF/CREB/Bcl-2 were rather increased. Interestingly, amyloid-precursor protein processing was shifted toward sAPPα generation, with a decrease of sAPPß and amyloid-beta levels. Our data confirm that F3/contactin plays a role in hippocampal synaptic plasticity and memory also in aged mice, suggesting that it acts on molecular pathways related to apoptosis and amyloid-beta production.

Neurobehavioral studies, in transgenic F3/CONTACTIN (C57BL/6J × CBA) mice, on cognitive and anxiety aspects during late-adolescential period.

OBJECTIVE: Besides than in the control of developmental events, axonal adhesive glycoproteins may be also involved in functions requiring fine organization and connectivity of the nervous tissue. We previously demonstrated morphological alterations and functional cerebellar deficits in transgenic mice (TAG/F3 mice) ectopically expressing the F3/Contactin axonal glycoprotein under the control of a selected regulatory region from the Transient Axonal Glycoprotein (TAG-1) gene. In the present study, the hippocampal function was explored by evaluating the ability of TAG/F3 mice to encode spatial and non-spatial relationships between discrete stimuli and to analyze an anxiety-related behavior. MATERIALS AND METHODS: To the first end, mice were placed in an "open-Field" containing five objects and, after three sessions of habituation (S2-S4), their reactivity to objects displacement (S5-S4) and object substitution (S7-S6) was examined.To the second end, mice were placed in the "elevated zero maze", a standard test to explore the anxiety-related behavior, in order to study, in transgenic mice, the effects of F3 misexpression on emotional reactivity by measuring the avoidance of the unsheltered open sectors. RESULTS: Statistical evaluations of reactivity to object novelty, TAG-F3 mice showed a lower DO exploration with respect to wild-type mice and, regarding DOs, TAG/F3 mice interacted less than wild-type mice, showing an impaired spatial change response. Furthermore, the number of HDIPS in transgenic TAG/F3 mice resulted significantly lower with respect to the controls (wild type). CONCLUSIONS: These results indicate that the coordinated expression of axonal adhesive glycoproteins may be relevant for the functional maturation of the hippocampus.

Contactin-1 regulates myelination and nodal/paranodal domain organization in the central nervous system.

Myelin, a multilayered membrane sheath formed by oligodendrocytes around axons in the CNS, enables rapid nerve impulse conduction and sustains neuronal health. The signals exchanged between axons and oligodendrocytes in myelin remain to be fully elucidated. Here we provide genetic evidence for multiple and critical functions of Contactin-1 in central myelin. We document dynamic Contactin-1 expression on oligodendrocytes in vivo, and progressive accumulation at nodes of Ranvier and paranodes during postnatal mouse development. Nodal and paranodal expression stabilized in mature myelin, but overall membranous expression diminished. Contactin-1-deficiency disrupted paranodal junction formation as evidenced by loss of Caspr, mislocalized potassium Kv1.2 channels, and abnormal myelin terminal loops. Reduced numbers and impaired maturation of sodium channel clusters accompanied this phenotype. Histological, electron microscopic, and biochemical analyses uncovered significant hypomyelination in Contactin-1-deficient central nerves, with up to 60% myelin loss. Oligodendrocytes were present in normal numbers, albeit a minor population of neuronal/glial antigen 2-positive (NG2(+)) progenitors lagged in maturation by postnatal day 18, when the mouse null mutation was lethal. Major contributing factors to hypomyelination were defects in the generation and organization of myelin membranes, as judged by electron microscopy and quantitative analysis of oligodendrocyte processes labeled by GFP transgenically expressed from the proteolipid protein promoter. These data reveal that Contactin-1 regulates both myelin formation and organization of nodal and paranodal domains in the CNS. These multiple roles distinguish central Contactin-1 functions from its specific role at paranodes in the periphery, and emphasize mechanistic differences in central and peripheral myelination.

F3/Contactin acts as a modulator of neurogenesis during cerebral cortex development.

The expression of the cell recognition molecule F3/Contactin (CNTN1) is generally associated with the functions of post-mitotic neurons. In the embryonic cortex, however, we find it expressed by proliferating ventricular zone (VZ) precursors. In contrast to previous findings in the developing cerebellum, F3/Contactin transgenic overexpression in the early cortical VZ promotes proliferation and expands the precursor pool at the expense of neurogenesis. At later stages, when F3/Contactin levels subside, however, neurogenesis resumes, suggesting that F3/Contactin expression in the VZ is inversely related to neurogenesis and plays a role in a feedback control mechanism, regulating the orderly progression of cortical development. The modified F3/Contactin profile therefore results in delayed corticogenesis, as judged by downregulation in upper and lower layer marker expression and by BrdU birth dating, indicating that, in this transgenic model, increased F3/Contactin levels counteract neuronal precursor commitment. These effects also occur in primary cultures and are reproduced by addition of an F3/Fc fusion protein to wild type cultures. Together, these data indicate a completely novel function for F3/Contactin. Parallel changes in the generation of the Notch Intracellular Domain and in the expression of the Hes-1 transcription factor indicate that activation of the Notch pathway plays a role in this phenotype, consistent with previous in vitro reports that F3/Contactin is a Notch1 ligand.