three-dimensional-quantification-dendritic-spines-from-pyramidal

Three-dimensional Quantification of Dendritic Spines from Pyramidal Neurons Derived from Human Induced Pluripotent Stem Cells

Dendritic spines are small protrusions that correspond to the post-synaptic compartments of excitatory synapses in the central nervous system. They are ...

Institut Pasteur

Université Paris Diderot, Sorbonne Paris Cité

The neural cell-adhesion molecules contactin 4, contactin 5 and contactin 6 are involved in brain development, and disruptions in contactin genes may confer increased risk for autism spectrum disorders (ASD). We describe a co-culture of rat cortical neurons and HEK293 cells overexpressing and delivering the secreted forms of rat contactin 4-6. We quantified their effects on the length and branching of neurites. Contactin 4-6 effects were different depending on the contactin member and duration of co-culture. At 4 days in culture, contactin 4 and -6 increased the length of neurites, while contactin 5 increased the number of roots. Up to 8 days in culture, contactin 6 progressively increased the length of neurites while contactin 5 was more efficient on neurite branching. We studied the molecular sites of interaction between human contactin 4, -5 or -6 and the human Protein Tyrosine Phosphatase Receptor Gamma (PTPRG), a contactin partner, by modeling their 3D structures. As compared to contactin 4, we observed differences in the Ig2 and Ig3 domains of contactin 5 and -6 with the appearance of an omega loop that could adopt three distinct conformations. However, interactive residues between human contactin 4-6 and PTPRG were strictly conserved. We did not observe any differences in PTPRG binding on contactin 5 and -6 either. Our data suggest that the differential contactin effects on neurite outgrowth do not result from distinct interactions with PTPRG. A better understanding of the contactin cellular properties should help elucidate their roles in ASD.

Contactin 4, -5 and -6 differentially regulate neuritogenesis while they display identical PTPRG binding sites.

Les mol&eacute;cules d&#39;adh&eacute;sion-cellulaire neuronal contactin 4, contactin 5 et 6 de contactin sont impliqu&eacute;s dans le d&eacute;veloppement du cerveau, et les perturbations dans la contactin de g&egrave;nes peuvent conf&eacute;rer &agrave; un risque accru de troubles du spectre autistique (TSA). Nous d&eacute;crivons une co-culture de neurones corticaux de rat et les cellules HEK293 surexprimant et livrer les formes s&eacute;cr&eacute;t&eacute;es de rat contactin 4-6. Nous avons quantifi&eacute; leurs effets sur la longueur et la ramification des neurites. Contactin 4-6 effets ont &eacute;t&eacute; diff&eacute;rents selon la dur&eacute;e de co-culture et le membre contactin. &Agrave; 4 jours de culture, contactin 4 et -6 a augment&eacute; la longueur des neurites, tandis que 5 contactin a augment&eacute; le nombre de racines. Jusqu&#39;&agrave; 8 jours de culture, contactin 6 progressivement augment&eacute; la longueur des neurites tout en contactin 5 &eacute;tait plus efficace sur la ramification des neurites. Nous avons &eacute;tudi&eacute; les sites mol&eacute;culaires de l&#39;interaction humaine 4 contactin, -5 ou -6 et l&#39;humain prot&eacute;ine Tyrosine Phosphatase Receptor Gamma (PTPRG), partenaire de contactin, par leurs structures 3D de mod&eacute;lisation. Par rapport &agrave; contactin 4, nous avons observ&eacute; des diff&eacute;rences dans les domaines Ig2 et Ig3 de contactin 5 et -6 avec l&#39;apparition d&#39;une boucle om&eacute;ga qui pourrait adopter trois conformations distinctes. Toutefois, les r&eacute;sidus interactives entre humains contactin 4-6 et PTPRG &eacute;taient strictement conserv&eacute;s. On n&#39;a pas observ&eacute; de diff&eacute;rences quant au lie PTPRG contactin 5 et -6 non plus. Nos r&eacute;sultats sugg&egrave;rent que les effets diff&eacute;rentiels de contactin sur la croissance des neurites ne r&eacute;sultent pas d&#39;interactions distinctes avec PTPRG. Une meilleure compr&eacute;hension des propri&eacute;t&eacute;s cellulaires contactin devrait aider &agrave; &eacute;lucider leur r&ocirc;le dans l&#39;ASD.

Contactin 4, -5 et -6 diff&eacute;rentiellement r&eacute;glementent neuritogenesis alors qu&#39;ils affichent des sites identiques de liaison PTPRG.

Le molecole di adesione cellulare neurale anche 4, anche 5 e anche 6 sono coinvolti nello sviluppo del cervello, e interruzioni nel anche geni possono conferire aumento del rischio di disturbi dello spettro autistico (ASD). Descriviamo un co-culture dei neuroni corticali di topo e cellule HEK293 con sovraesprimono relativa e trasportando le forme secrete di ratto anche 4-6. Abbiamo quantificato gli effetti sulla lunghezza e la ramificazione dei neuriti. Anche 4-6 effetti erano diverse a seconda del membro contactin e la durata del co-culture. A 4 giorni in coltura, contactin 4 e -6 aumentato la lunghezza dei neuriti, mentre 5 contactin aumentato il numero di radici. Fino a 8 giorni in coltura, anche 6 progressivamente aumentato la lunghezza dei neuriti mentre anche 5 era pi&ugrave; efficiente sulla diramazione dei neuriti. Abbiamo studiato i siti molecolari dell&#39;interazione tra umano contactin 4, -5 o -6 e l&#39;umano proteina tirosina fosfatasi del recettore Gamma (PTPRG), un partner contactin, modellando le strutture 3D. Rispetto al anche 4, abbiamo osservato le differenze nei domini Ig2 e Ig3 contactin 5 e -6 con l&#39;aspetto di un ciclo di omega che potrebbe adottare tre distinte conformazioni. Tuttavia, residui interattivi tra 4-6 contactin umano e PTPRG erano rigorosamente conservati. Non ha fatto osservare eventuali differenze nell&#39;associazione PTPRG contactin 5 e -6 o. I nostri dati suggeriscono che gli effetti contactin differenziali su neuriti escrescenza non derivano da interazioni distinte con PTPRG. Una migliore comprensione delle propriet&agrave; cellulari contactin dovrebbe aiutare a chiarire i loro ruoli in ASD.

Laura Gouder

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