The expression of autism susceptibility genes in the earliest stages of human cerebral cortex development

Abstract

Autism susceptibility gene (ASG) mutations are suspected to perturb developmental pathways essential for the correct formation and organisation of the brain. Many ASG products, including neurexins (NRXNs), neuroligins (NLGNs) and SHANKs, have established functions in mature synapses. However, a previous microarray study suggested that ASGs are expressed in the human cortex from as early as 8 PCW, when synapses are sparse. RNA sequencing of human cortical samples revealed that a number of ASGs that code for cell adhesion molecules, enzymes that synthesise neurotransmitters, neurotransmitter receptors, molecules involved in neurotransmitter transport, synaptic transmission, cell-cell signalling and neuronal differentiation are differentially expressed both spatially and temporally between 9 and 12 PCWs. NRXNs, NLGNs and SHANKs, were studied in further detail using RNA sequencing, qPCR and IHC. Many of the NRXN and NLGN genes showed an increase in expression between 8 and 12 PCW and were significantly increased in either the anterior or temporal cortex. The SHANK genes did not increase in expression with age and there was high variation between the RNA sequencing data and the qPCR data. Immunohistochemistry revealed that NRXN proteins have distinct distribution patterns within the cortex at these ages. NRXNs 1 and 3 were predominantly found in the cortical plate (CP) whilst NRXN2 immunopositivity was strongly co-localised with the presynaptic vesicle protein, SYP and the marker of outgrowing axons, GAP43, largely found in the pre-subplate, marginal zone and intermediate zone. Regulators of NRXN splicing, were expressed primarily in the CP whereas TOP2B, a regulator of NRXN transcription, was found throughout the cortex. The distribution of NRXN proteins suggests that, in addition to their established function in synaptogenesis, they may have diverse roles in development including axon guidance and intercellular communication between proliferating cells and/or migrating neurons.