Genetics of Autism Spectrum


Head: Sabine M. Klauck

Former members: Kim Beyer, Andreas Chiocchetti, Bärbel Felder, Daniela Holzwarth, Sabine Karolus, Maren Kettner, Tatjana Kraus, Geeta Pakalapati, Annemarie Poustka†, Kinga Przibilla, Nicole Pustelnik, Stefanie Rauskolb, Tatjana Salvi, Inaam Shatila, Anja Spieler, Cornelia Wirth

Autism spectrum disorders (ASD) are severe neurodevelopmental disorders characterized by marked social deficits, deviant language and a restricted range of stereotyped repetitive behaviours usually occurring within the first three years of life (Klauck, 2006). The prevalence for ASD is 0.6-1% with a ratio of affected male to female of 4:1. Molecular genetic analyses point towards a multifactorial genetic predisposition involving risk factors for ASD like copy number variation (CNV) and single nucleotide polymorphisms, but also single rare mutations (Chiocchetti and Klauck, 2011; Klauck et al., 2011).

The goal of our project over more than two decades was to identify disease genes for ASD with three main approaches. Firstly, collaborative research focused on genome-wide linkage and association studies, and lately CNV studies, as members of the "International Molecular Genetic Study of Autism Consortium” (IMGSAC), the “Autism Genome Project (AGP) Consortium” and contributors for “TASC - The Autism Simplex Collection” (Buxbaum et al., 2014). Secondly, family-based association studies of the division’s large patient sample, collected in collaboration with F. Poustka and C.M. Freitag of the Department of Child and Adolescent Psychiatry, Psychosomatics and Psychotherapy, Goethe University Frankfurt, examined polymorphic markers in candidate genes and at defined gene loci. In addition, several candidate genes from the regions identified in the genome-wide linkage studies were screened systematically for functional variants in autistic probands. Finally, we examined protein signatures in a patient specific cell line model for autism.

In the first approach initial genome screens identified susceptibility regions most prominently on chromosomes 7q31-q35 (IMGSAC, 1998), 7q22 and 2q, 16p and 17q (IMGSAC, 2001; IMGSAC, 2001), as well as sex-limited effects of susceptibility loci on 7q, 15q and 16p, and parent-of-origin effects on 7q and 9p (Lamb et al., 2005). Consecutively, the AGP Consortium conducted genome-wide SNP (single nucleotide polymorphism) screens and revealed a new susceptibility locus on chromosome 11p12-p13, and moreover genetic components of the synaptic glutamatergic signaling system including the gene neurexin 1 (NRXN1) as promising candidates contributing to ASD (The Autism Genome Project Consortium, 2007). A European study performed by IMGSAC found linkage again on chromosome 2q and association in candidate genes RELN, GRIK2, MKL2 and SND1 (Holt et al., 2010), as well as in leucine-rich repeat genes (Sousa et al., 2010). Focusing more on the analysis of chromosomal aberrations using dense genotyping arrays by the AGP consortium, lead to the discovery that patients with ASD carry a higher global burden of rare, genic CNVs, especially for loci previously implicated in either ASD and/or intellectual disability (Pinto et al., 2010). A follow-up study of AGP integrating 2,446 ASD-affected families confirmed this previous finding of an excess of genic deletions and duplications in affected versus control groups, while showing that the genes affected by de novo CNVs and/or loss-of-function single-nucleotide variants converged on networks related to neuronal signaling and development, synapse function, and chromatin regulation (Pinto et al., 2014).

The second approach included the analysis of candidate genes using family-based association studies and/or mutational analyses. The genes WNT2 (Beyer, K. 2001 PhD thesis), RELN (Bonora et al., 2003); FOXP2 (Newbury et al., 2002), PEG/MEST, COPG2, CPA1 and CPA5 (Bonora et al., 2002), CUTL1, SRPK2, SYPL, LAMB1, NRCAM and PTPRZ1 (Bonora et al., 2005) - all located within the candidate region on chromosome 7q - were tested, but rare variants detected were not of major relevance for ASD in the patient sample under study. Likewise, further candidate genes from other genomic regions [HOPA, Xq13, (Beyer et al., 2002); MECP2, Xq28, (Beyer et al., 2002); MACROD2, 20p12.1 (Curran et al., 2011)] could be ruled out as main factors for the etiology of ASD. Instead, rare cases with a microdeletion on chromosome 15q13.3 (Pagnamenta et al., 2009), a terminal deletion of 2q37.3 (Felder et al. 2009), and deletions in the genes CNTNAP5 and DOCK4 (Pagnamenta et al. 2010), have been identified in collaborative efforts. Genetic and functional analyses of SHANK2 mutations suggest a multiple hit model of ASD (Leblond et al., 2012). Most recently, common variants in genes of the postsynaptic FMRP signaling pathway (Waltes et al., 2014) and in the gene CNTNAP2 (Chiocchetti et al., 2014) have been found as risk factors for ASD.

We have identified two missense mutations in the ribosomal protein gene RPL10 located in Xq28 in three independent families with autism (Klauck et al., 2006, Chiocchetti et al., 2011), a multicenter study with colleagues of the Universities of Frankfurt (F. Poustka) and Salzburg (H. Breitenbach-Koller). Based on the findings that the amino-acid substitutions (L206M, H213Q) confer hypomorphism with respect to the regulation of the translation process and high expression of RPL10 in hippocampus, we suggest a novel modulating disease mechanism for autism. A change in translational function may impact on those cognitive functions that are mediated through the limbic system.

Finally, we performed protein profiling by two-dimensional gel electrophoresis with protein extracts from patient-derived lymphoblastoid cell lines (LCLs), which we established in our group. Comparing LCLs from carriers of the ASD-specific mutation RPL10[H213Q] and from unrelated ASD patients without the mutation, we discovered protein signatures of oxidative stress response pointing towards a common molecular pathomechanism in ASD, characterized in our study by dysregulation of redox balance in redox-sensitive elements of energy-, protein- and redox-metabolism (Chiocchetti et al., 2014). At pathway levels, this redox-sensitive protein signature has also been identified in a yeast rpl10 deficiency and an oxidative stress model in our study. Importantly, this can be triggered by the known ASD-RPL10[H213Q] mutation or by yet unknown mutations of the unrelated ASD cohort that act upstream of RPL10 in differential expression of redox-sensitive proteins.

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