Human CRY1 variants associate with attention deficit/hyperactivity disorder
O. Emre Onat, M. Ece Kars, Şeref Gül, Kaya Bilguvar, Yiming Wu, Ayşe Özhan, Cihan Aydın, A. Nazlı Başak, M. Allegra Trusso, Arianna Goracci, Chiara Fallerini, Alessandra Renieri, Jean-Laurent Casanova, Yuval Itan, Cem E. Atbaşoğlu, Meram C. Saka, İ. Halil Kavaklı, Tayfun Özçelik
O. Emre Onat, M. Ece Kars, Şeref Gül, Kaya Bilguvar, Yiming Wu, Ayşe Özhan, Cihan Aydın, A. Nazlı Başak, M. Allegra Trusso, Arianna Goracci, Chiara Fallerini, Alessandra Renieri, Jean-Laurent Casanova, Yuval Itan, Cem E. Atbaşoğlu, Meram C. Saka, İ. Halil Kavaklı, Tayfun Özçelik
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Research Article Genetics

Human CRY1 variants associate with attention deficit/hyperactivity disorder

Abstract

Attention deficit/hyperactivity disorder (ADHD) is a common and heritable phenotype frequently accompanied by insomnia, anxiety, and depression. Here, using a reverse phenotyping approach, we report heterozygous coding variations in the core circadian clock gene cryptochrome 1 in 15 unrelated multigenerational families with combined ADHD and insomnia. The variants led to functional alterations in the circadian molecular rhythms, providing a mechanistic link to the behavioral symptoms. One variant, CRY1Δ11 c.1657+3A>C, is present in approximately 1% of Europeans, therefore standing out as a diagnostic and therapeutic marker. We showed by exome sequencing in an independent cohort of patients with combined ADHD and insomnia that 8 of 62 patients and 0 of 369 controls carried CRY1Δ11. Also, we identified a variant, CRY1Δ6 c.825+1G>A, that shows reduced affinity for BMAL1/CLOCK and causes an arrhythmic phenotype. Genotype-phenotype correlation analysis revealed that this variant segregated with ADHD and delayed sleep phase disorder (DSPD) in the affected family. Finally, we found in a phenome-wide association study involving 9438 unrelated adult Europeans that CRY1Δ11 was associated with major depressive disorder, insomnia, and anxiety. These results defined a distinctive group of circadian psychiatric phenotypes that we propose to designate as “circiatric” disorders.

Authors

O. Emre Onat, M. Ece Kars, Şeref Gül, Kaya Bilguvar, Yiming Wu, Ayşe Özhan, Cihan Aydın, A. Nazlı Başak, M. Allegra Trusso, Arianna Goracci, Chiara Fallerini, Alessandra Renieri, Jean-Laurent Casanova, Yuval Itan, Cem E. Atbaşoğlu, Meram C. Saka, İ. Halil Kavaklı, Tayfun Özçelik

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Figure 8

Phenotype-genotype characterization of family DSPD-36 and functional characterization of CRY1Δ6.

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Phenotype-genotype characterization of family DSPD-36 and functional cha...
(A–C) Phenotype-genotype characterization of family DSPD-36. (A) The family DSPD-36 was assessed as described in the legend to Figure 3. (B and C) c.825+1G>A causes skipping of 141-bp exon 6, leading to an in-frame deletion of 47 residues in the middle of the PHR and clock-binding domains of the CRY1 protein. M, DNA marker. (D–J) Functional characterization of CRY1Δ6. (D) Docking analysis of CRY1 and the CLOCK PAS-B domain. (E) The critical amino acid residue of CRY1 (R256) interacts with CLOCK PAS-B (W362). (F) Analysis of the effect of FL WT CRY1, CRY1Δ6, and CRY1Δ11 on CLOCK/BMAL1-driven transcription with the Per1:Luc assay. An E-box–driven luciferase reporter plasmid Per1:Luc was coexpressed in HEK293T cells along with plasmids consisting of CLOCK and BMAL1 cDNAs and decreasing amounts of WT CRY1 or CRY1Δ6. Data represent the mean induction of bioluminescence over basal levels from duplicate transfections 48 hours after transfection. Error bars represent the SD from at least 3 biological experiments. Co-IP assay with (G) CLOCK, BMAL1, (H) PER2, CLOCK, BMAL1, and human CRY1s (hCRY1). Blots in G and H are representative of at least 3 independent experiments. (I) Rescue assays were performed with human CRY1s with at least 4 biological replicates. Samples were normalized to the initial luminescence signal. The graph below indicates the period differences, with the whiskers representing the mean ± SEM values. n = 5 per group. Data were pooled from 2 independent experiments. **P = 0.002, by unpaired t test. mPer2-Luc, mouse Per2-Luc. (J) Degradation assay with human CRY1::Luc and its variants. The graph below was generated by fitting a 1-phase decay curve to the data, which indicate the half-life, with the horizontal line representing the mean. n = 3 per condition, pooled from 3 independent experiments. **P = 0.002, by unpaired t test.