Loss of function of the autism risk gene setd5 leads to neurotransmission-associated gene alterations and social impairments in zebrafish

SETD5 loss of function (LoF) mutations in humans have been recently associated to severe developmental defects, including intellectual disability (ID) and autistic spectrum disorders (ASD). The SETD5 gene encodes for a putative histone H3 methyltransferase highly expressed in the brain and falls within the critical interval deleted in the “3p25.3 microdeletion syndrome”, characterized by ID, microcephaly and congenital heart defects. The aim of this study is to characterize setd5 LoF zebrafish models generated by morpholino injection and CRISPR/Cas9 gene editing technique. Our results show that setd5 is maternally inherited displaying a widespread expression at early development stages, while at later stages its expression is more localized with an evident antero-posterior gradient, specifically including the developing central nervous system (CNS). Moreover, setd5 expression is localized in specific brain areas of adult zebrafish. In addition, antisense morpholino oligonucleotides-mediated targeting of setd5 in zebrafish embryos determined microcephaly, cardiac edema and reduced locomotory response. Moreover, compared to embryos injected with a control morpholino, setd5 morphant brains, despite their reduced size, show an increase of phospho-H3-positive mitotic cells and an increase of apoptotic rate in developing brain areas. To analyse the effects of stable setd5 gene knockout on zebrafish larvae and adults, we generated setd5 mutant zebrafish lines taking advantage of the CRISPR/Cas9 strategy. We observed that setd5 knockout causes microcephaly, a significant reduction of body length and locomotor activity in both larvae and adults. Moreover, in setd5 mutants both embryo heads and adult brains display reduced expression of synaptic proteins and specific enzymes involved in neurotransmission. In addition, setd5 mutant adults are characterized by a reduced social interaction when compared to wild type siblings. Such autism-like altered behavioural traits triggered by setd5 mutations are ameliorated by Risperidone, an antipsychotic drug commonly used to treat behavioural traits in ASD patients. The validation of the zebrafish setd5 mutants as reliable models for ASD/ID could play an important role in the study of these developmental defects. Finally, the characterization of the molecular pathways altered by setd5 LoF may support the screening for targeted compounds able to rescue the developmental and behavioural defects observed in these zebrafish mutants, to identify novel promising compounds for ameliorating behavioural alterations in human individuals affected by ASD/ID due to SETD5 haploinsufficiency.