Mechanism of KMT5B haploinsufficiency in neurodevelopment in humans and mice.

Pathogenic variants in KMT5B, a lysine methyltransferase, are associated with global developmental delay, macrocephaly, autism, and congenital anomalies (OMIM# 617788). Given the relatively recent discovery of this disorder, it has not been fully characterized. Deep phenotyping of the largest (n = 43) patient cohort to date identified that hypotonia and congenital heart defects are prominent features that were previously not associated with this syndrome. Both missense variants and putative loss-of-function variants resulted in slow growth in patient-derived cell lines. KMT5B homozygous knockout mice were smaller in size than their wild-type littermates but did not have significantly smaller brains, suggesting relative macrocephaly, also noted as a prominent clinical feature. RNA sequencing of patient lymphoblasts and Kmt5b haploinsufficient mouse brains identified differentially expressed pathways associated with nervous system development and function including axon guidance signaling. Overall, we identified additional pathogenic variants and clinical features in KMT5B-related neurodevelopmental disorder and provide insights into the molecular mechanisms of the disorder using multiple model systems.

KMT5B is required for early motor development.

Disruptive variants in lysine methyl transferase 5B (KMT5B/SUV4-20H1) have been identified as likely-pathogenic among humans with neurodevelopmental phenotypes including motor deficits (i.e., hypotonia and motor delay). However, the role that this enzyme plays in early motor development is largely unknown. Using a Kmt5b gene trap mouse model, we assessed neuromuscular strength, skeletal muscle weight (i.e., muscle mass), neuromuscular junction (NMJ) structure, and myofiber type, size, and distribution. Tests were performed over developmental time (postnatal days 17 and 44) to represent postnatal versus adult structures in slow- and fast-twitch muscle types. Prior to the onset of puberty, slow-twitch muscle weight was significantly reduced in heterozygous compared to wild-type males but not females. At the young adult stage, we identified decreased neuromuscular strength, decreased skeletal muscle weights (both slow- and fast-twitch), increased NMJ fragmentation (in slow-twitch muscle), and smaller myofibers in both sexes. We conclude that Kmt5b haploinsufficiency results in a skeletal muscle developmental deficit causing reduced muscle mass and body weight.

Differential effects by sex with Kmt5b loss.

Lysine methyl transferase 5B (KMT5B) has been recently highlighted as a risk gene in genetic studies of neurodevelopmental disorders (NDDs), specifically, autism spectrum disorder (ASD) and intellectual disability (ID); yet, its role in the brain is not known. The goal of this work was to neurodevelopmentally characterize the effect(s) of KMT5B haploinsufficiency using a mouse model. A Kmt5b gene-trap mouse line was obtained from the Knockout Mouse Project. Wild type (WT) and heterozygous (HET) mice were subjected to a comprehensive neurodevelopmental test battery to assess reflexes, motor behavior, learning/memory, social behavior, repetitive movement, and common ASD comorbidities (obsessive compulsion, depression, and anxiety). Given the strong sex bias observed in the ASD patient population, we tested both a male and female cohort of animals and compared differences between genotypes and sexes. HET mice were significantly smaller than WT littermates starting at postnatal day 10 through young adulthood which was correlated with smaller brain size (i.e., microcephaly). This was more severe in males than females. HET male neonates also had delayed eye opening and significantly weaker reflexes than WT littermates. In young adults, significant differences between genotypes relative to anxiety, depression, fear, and extinction learning were observed. Interestingly, several sexually dimorphic differences were noted including increased repetitive grooming behavior in HET females and an increased latency to hot plate response in HET females versus a decreased latency in HET males. LAY SUMMARY: Lysine methyl transferase 5B (KMT5B) has been recently highlighted as a risk gene in neurodevelopmental disorders (NDDs), specifically, autism spectrum disorder (ASD) and intellectual disability (ID); yet its role in the brain is not known. Our study indicates that mice lacking one genomic copy of Kmt5b show deficits in neonatal reflexes, sociability, repetitive stress-induced grooming, changes in thermal pain sensing, decreased depression and anxiety, increased fear, slower extinction learning, and lower body weight, length, and brain size. Furthermore, several outcomes differed by sex, perhaps mirroring the sex bias in ASD.