CNTNAP2 intracellular domain (CICD) generated by γ-secretase cleavage improves autism-related behaviors

Abstract

As the most prevalent neurodevelopmental disorders in children, autism spectrum disorders (ASD) are characterized by deficits in language development, social interaction, and repetitive behaviors or inflexible interests. Contactin associated protein like 2 (CNTNAP2), encoding a single transmembrane protein (CNTNAP2) with 1331 amino acid residues, is a widely validated ASD-susceptible gene. Cntnap2-deficient mice also show core autism-relevant behaviors, including the social deficits and repetitive behavior. However, the cellular mechanisms underlying dysfunction CNTNAP2 and ASD remain elusive. In this study, we found a motif within the transmembrane domain of CNTNAP2 was highly homologous to the γ-secretase cleavage site of amyloid-β precursor protein (APP), suggesting that CNTNAP2 may undergo proteolytic cleavage. Further biochemical analysis indicated that CNTNAP2 is cleaved by γ-secretase to produce the CNTNAP2 intracellular domain (CICD). Virally delivery of CICD to the medial prefrontal cortex (mPFC) in Cntnap2-deficient (Cntnap2−/−) mice normalized the deficit in the ASD-related behaviors, including social deficit and repetitive behaviors. Furthermore, CICD promoted the nuclear translocation of calcium/calmodulin-dependent serine protein kinase (CASK) to regulate the transcription of genes, such as Prader Willi syndrome gene Necdin. Whereas Necdin deficiency led to reduced social interaction in mice, virally expression of Necdin in the mPFC normalized the deficit in social preference of Cntnap2−/− mice. Our results thus reveal a critical function of CICD and highlight a role of the CNTNAP2-CASK-Necdin signaling pathway in ASD.

Investigation of a Novel Mouse Model of Prader-Willi Syndrome with Invalidation ofNecdinandMagel2

“…Our teams and others also reported impaired hypothalamic regulation in Magel2 KO mice with abnormal oxytocin (OT) maturation (10,14,18,19), and disrupted development and function of pro-opiomelanocortin (POMC) neurons (19,20). Necdin KO mice display variable lethality after birth (21,22) due to respiratory distress (23), growth retardation, motor deficit in infancy (24), sensory deficits (25), high scraping, cognitive alterations (22) and alterations of social and circadian behaviors (26,27). At the neuroanatomical level, Necdin KO mice display a reduction in the number of OT and gonadotropin-releasing hormone (GnRH)-producing neurons, alterations in perinatal serotonergic metabolism and development (22,28), and alterations in clock gene expression (26).…”

“…Behavioural changes are present throughout the patient's life. Anxiety, interactions with novel objects or social interactions are altered in Necdin (27) and Magel2 KO mice (14,17), with a variability in the severity of the phenotype, partly due to the environmental context. Madin KO mice showed a preference for the novel object versus a familiar one and for a congener mouse versus an empty cage.…”

Contactin-associated protein-like 2 (CNTNAP2) mutations impair the essential α-secretase cleavages, leading to autism-like phenotypes

“…We recently reported that CICD enters the nucleus and functions as a transcription factor to improve autism-related behaviors. 31 Therefore, disruptions in the α-cleavage of CNTNAP2, such as by pathogenic mutations shown in Fig. 4, may affect the downstream CNTNAP2 signaling pathway.…”

“…30 Recently, we found that CNTNAP2 is cleaved by presenilins-dependent γ-secretase to produce the CNTNAP2 intracellular domain (CICD), and expression of the CICD improves autism-like behaviors. 31 Our current study reveals that in addition to its γ-secretase cleavage, CNTNAP2 undergoes complex proteolytic processing by furin and a disintegrin and metalloproteinase (ADAM)10/17-dependent α-secretase. The main α-cleavage product C79 by ADAM10 is required for the subsequent γ-secretase cleavage to generate the CICD.…”

“…In a separate study, we discovered that CNTNAP2 is cleaved by γ-secretase to generate the CNTNAP2 intracellular domain (CICD), and CNTNAP2-CICD expression improves ASD-related behaviors. 31 L-685,458, a potent γ-secretase inhibitor (GSI), was applied to treat HEK cells co-transfected with CNTNAP2 and empty vector, ADAM10 or ADAM17. L-685,458 treatment significantly increased C79 levels in the empty vector (1.34 ± 0.06 folds, p = 0.0043), ADAM10 (1.97 ± 0.12 folds, p = 0.0013), and ADAM17 (1.78 ± 0.22 folds, p = 0.0227) overexpressing cells, indicating that the main α-cleavage product C79 is further cleaved by γ-secretase (Fig.…”