CRISPR-Cas9-engineered PIGV341E mouse model mirrors human phenotype, shows hippocampal synaptic dysfunctions and suggests a major role of impaired Eph/Abl1 signaling in GPI anchor deficiencies

Pathogenic germline mutations in PIGV result in a glycosylphosphatidylinositol biosynthesis deficiency (GPIBD). Individuals with pathogenic biallelic mutations in genes of the GPI pathway show cognitive impairments, a motor delay and often epilepsies. Thus far, the pathophysiology underlying the disease remains unclear and suitable rodent models are lacking that mimic human pathophysiology. We therefore generated a mouse model using CRISPR-Cas9 to introduce the most prevalent hypomorphic missense mutation in European patients, at a site that is also conserved in mice, Pigv:c.1022C>A (p.A341E). Pigv341E mice showed deficits in motor coordination and cognitive impairment with poorer long-term spatial memory. Furthermore, Pigv341E mice showed alterations in sociability and sleep patterns reflecting the human phenotype. Immunohistochemistry showed decreased synaptophysin-immunoreactivity and electrophysiology recordings demonstrated reduced hippocampal synaptic transmission in Pigv341E mice that may explain impaired memory formation. Moreover, we performed single-cell RNA sequencing of hippocampal cells from brains of adult Pigv341E mice that exhibit changes of gene expression, most prominently in a microglia subtype and subicular neurons. A significant reduction of Abl1 transcripts in several cell clusters suggest a link to the signaling pathway of GPI-anchored ephrins. We also identified increased levels of Hdc that might affect histamine metabolism with consequences for circadian rhythm and neuronal excitability. In summary, we present here the first mouse model with a patient-specific hypomorphic mutation that mirrors the human phenotype and will allow us to explore the pathophysiology in GPIBDs in future studies. Overall design: For single-cell RNA sequencing we collected both left and right hippocampi from Pigv341E mice (n=4) and wild-type mice (n=4). Isolated hippocampus cells were pooled by genotype, resulting in a total of 2 samples from which sequencing libraries were generated using the Chromium Single-cell Gene Expression Solution v3 (10X Genomics, Inc.) and sequenced with the Illumina NovaSeq 6000. Both the pooled Pigv341E and wild-type sample were sequenced on the same lane.