Chd8 haploinsufficiency impairs early brain development and protein homeostasis later in life

Abstract

Background Chromodomain helicase DNA-binding protein 8 (Chd8) is a high-confidence risk gene for autism spectrum disorder (ASD). However, how Chd8 haploinsufficiency impairs gene expression in the brain and impacts behavior at different stages of life is unknown. Methods We generated a mutant mouse line with an ASD-linked loss-of-function mutation in Chd8 (V986*; stop codon mutation). We examined the behavior of Chd8 mutant mice along with transcriptional changes in the cerebral cortex as a function of age, with a focus on one embryonic (E14.5) and three postnatal ages (1, 6, and 12 months). Results Chd8V986*/+ mutant mice displayed macrocephaly, reduced rearing responses and reduced center time in the open field, and enhanced social novelty preference. Behavioral phenotypes were more evident in Chd8V986*/+ mutant mice at 1 year of age. Pup survival was reduced in wild-type x Chd8V986*/+ crosses when the mutant parent was female. Transcriptomic analyses indicated that pathways associated with synaptic and neuronal projections and sodium channel activity were reduced in the cortex of embryonic Chd8V986*/+ mice and then equalized relative to wild-type mice in the postnatal period. At 12 months of age, expression of genes associated with endoplasmic reticulum (ER) stress, chaperone-mediated protein folding, and the unfolded protein response (UPR) were reduced in Chd8V986*/+ mice, whereas genes associated with the c-MET signaling pathway were increased in expression. Limitations It is unclear whether the transcriptional changes observed with age in Chd8V986*/+ mice reflect a direct effect of CHD8-regulated gene expression, or if CHD8 indirectly affects the expression of UPR/ER stress genes in adult mice as a consequence of neurodevelopmental abnormalities. Conclusions Collectively, these data suggest that UPR/ER stress pathways are reduced in the cerebral cortex of aged Chd8V986*/+ mice. Our study uncovers neurodevelopmental and age-related phenotypes in Chd8V986*/+ mice and highlights the importance of controlling for age when studying Chd8 haploinsufficient mice.

Controlling litter effects to enhance rigor and reproducibility with rodent models of neurodevelopmental disorders

“…In both cases, an experimental manipulation is being evaluated that has the potential to influence brain development in the pre- and/or early postnatal period. As a case in point, our data with wild-type and Chd8 V986*/+ mice, which model a high-confidence autism gene mutation [ 17 ], indicate that within litter variation is lower than between litters (Fig. 1 ).…”

“…Mice were maintained on a C57BL/6J background and raised in a facility with a 12:12 light:dark cycle with ad libitum access to food (Teklad 2020X, Envigo, Huntingdon, UK) and water. Male mice heterozygous for a high confidence CHD8 mutation ( Chd8 V986*/+ ), generated as previously described [ 17 ], were time mated with wild-type females. Matings were set up in the evening before the start of the dark cycle, using one male mouse and one female mouse per breeding cage.…”

“…Females were separated and single-housed upon confirmation of a vaginal plug the next day, considered as gestation day 0.5 (E0.5). Genotyping was performed as previously described [ 17 ]. Dams were sacrificed on E15.5, and embryos were collected by dissection in PBS.…”

“…These shared features within a litter are precisely what contribute to differences between litters. Other plausible contributing factors to litter effects that are seldom considered include differences in maternal behaviors, particularly if the mother harbors a mutant allele that affects maternal behaviors [ 17 , 24 ], and variation within the in utero environment. These factors cannot be accounted for statistically if the litter is not controlled as a variable.…”

“…When using rodents to assess genetic contributors to NDDs, it is crucial to test mutant dams for early signs of maternal behavior deficits that have the potential to confound results [ 17 ]. Litter size in addition to maternal behaviors, including licking/grooming, small or absent milk spots in pups, and failure to retrieve pups should be considered [ 24 , 36 ].…”