Defensive and offensive behaviors in social interaction settings in a Kleefstra syndrome mouse model

Background

Kleefstra syndrome is a neurodevelopmental disorder, caused by either heterozygous mutations in the euchromatic histone methyltransferase 1 (EHMT1) or in the lysine methyltransferase 2C (KMT2C) gene (Kleefstra et al. 2006; Koemans et al. 2017). Most Kleefstra syndrome patients (95.7%) also have autism spectrum disorder (ASD) (Vermeulen et al. 2017). EHMT1 regulates gene expression via methylation of histones, usually as a repressor, but can active the expression of certain genes as well (Ohno et al. 2013). EHMT1 expression is abundant in the developing neural system. The complete lack of EHMT1 leads to embryonic lethality in Ehmt1 knockout mice, indicating the essential role of this enzyme in the developing central nervous system (Tachibana et al. 2005). Ehmt1^+/- mice, models of Kleefstra syndrome, display similar behavioral phenotypes to those observed in human patients e.g., increased anxiety-like behavior, developmental delay, deficits in social behavior, etc. (Balemans et al. 2010). The memory and learning problems of Ehmt1^+/- mice may be caused by synaptic dysfunction in the hippocampus (Balemans et al. 2013).

In a recent clinical study, aggressive or emotional outburst were reported by parents of patients with Kleefstra syndrome (Haseley et al. 2021). This symptom has not been assessed in Ehmt1^+/- mice so far.

In this preprint, defensive and offensive (DO) behaviors, as murine analogues for aggressive outbursts, were evaluated in Ehmt1^+/- mice.

 

Key findings

An Ehmt1^+/- or wild-type (WT) mouse (host) was placed in an open-field box for 20 minutes to explore the novel environment. The time spent sitting in the corners or exploring the cues and walls were monitored. After this 20 minute-session, a non-cage mate mouse of the same sex (Ehmt1^+/- or WT, visitor) was placed in the box, and their interaction was recorded. It is a modified version of a previously published host-visitor interaction protocol (Granon et al. 2003). Attacking and biting were defined as offensive behaviors. DO behaviors were scored according to the following scale:

0: no defense

1: the mouse stands on their hind paws and/or rattles its tail

2: the mouse engages in a fight (biting)

3: distress vocalization

Overall, 74 trials were conducted, using 10 WT and 10 Ehmt1^+/- male mice (WT host vs. WT visitor, WT host vs. Ehmt1^+/- visitor, Ehmt1^+/- vs. WT visitor, etc.).

 

Exploratory behavior:

No differences were detected among Ehmt1^+/- and WT hosts during those initial 20 minutes. Hosts of both genotypes spent more time exploring the box than sitting in the corners. During the last 10 minutes, visitor mice spent more time exploring the box than the hosts. Overall, role had a main effect in the first 20 minutes.

 

DO behaviors

DO behaviors were not detected between WT-WT mice. In the case of WT (host)- Ehmt1^+/-(visitor) pairs, 6% of the trials contained DO behaviors, while in Ehmt1^+/-(host)- WT (visitor) trials, this ratio was 23%. In WT- Ehmt1^+/- pairings, it was always the Ehmt1^+/- mouse who started the fight. Interestingly, 42% of the Ehmt1^+/-– Ehmt1^+/- trials contained DO behaviors, thus, genotype had a main effect.

 

Why I liked this preprint

The authors developed a novel method for assessing aggressive outburst in mice and described this behavior in Kleefstra syndrome model mice in detail.

 

Questions for the authors

1. As you mentioned in the Discussion section, Ehmt1^+/- showed extreme aggressive behavior after social isolation. How could you quantify this behavior and elucidate the underlying mechanism in the CNS?
2. Interestingly, treatment with the EHMT1/2 inhibitor or silencing Ehmt1/2 expression in the prefrontal cortex of Shank3-deficient ASD mice led to improvement in social behavior in three chamber-test (Wang et al. 2020). Does this finding might suggest, that EHMT1 has a more complex role in the progression of ASD and the inhibition of EHMT1 might be therapeutic for ASD patients?

 

 

References

 

Balemans M.C., Huibers M.M., Eikelenboom N.W., Kuipers A.J., van Summeren R.C., Pijpers M.M., Tachibana M., Shinkai Y., van Bokhoven H. & Van der Zee C.E. (2010) Reduced exploration, increased anxiety, and altered social behavior: Autistic-like features of euchromatin histone methyltransferase 1 heterozygous knockout mice. Behav Brain Res 208, 47-55.

Balemans M.C., Kasri N.N., Kopanitsa M.V., Afinowi N.O., Ramakers G., Peters T.A., Beynon A.J., Janssen S.M., van Summeren R.C., Eeftens J.M., Eikelenboom N., Benevento M., Tachibana M., Shinkai Y., Kleefstra T., van Bokhoven H. & Van der Zee C.E. (2013) Hippocampal dysfunction in the Euchromatin histone methyltransferase 1 heterozygous knockout mouse model for Kleefstra syndrome. Hum Mol Genet 22, 852-66.

Granon S., Faure P. & Changeux J.P. (2003) Executive and social behaviors under nicotinic receptor regulation. Proc Natl Acad Sci U S A 100, 9596-601.

Haseley A., Wallis K. & DeBrosse S. (2021) Kleefstra syndrome: Impact on parents. Disabil Health J 14, 101018.

Kleefstra T., Brunner H.G., Amiel J., Oudakker A.R., Nillesen W.M., Magee A., Genevieve D., Cormier-Daire V., van Esch H., Fryns J.P., Hamel B.C., Sistermans E.A., de Vries B.B. & van Bokhoven H. (2006) Loss-of-function mutations in euchromatin histone methyl transferase 1 (EHMT1) cause the 9q34 subtelomeric deletion syndrome. Am J Hum Genet 79, 370-7.

Koemans T.S., Kleefstra T., Chubak M.C., Stone M.H., Reijnders M.R.F., de Munnik S., Willemsen M.H., Fenckova M., Stumpel C., Bok L.A., Sifuentes Saenz M., Byerly K.A., Baughn L.B., Stegmann A.P.A., Pfundt R., Zhou H., van Bokhoven H., Schenck A. & Kramer J.M. (2017) Functional convergence of histone methyltransferases EHMT1 and KMT2C involved in intellectual disability and autism spectrum disorder. PLoS Genet 13, e1006864.

Ohno H., Shinoda K., Ohyama K., Sharp L.Z. & Kajimura S. (2013) EHMT1 controls brown adipose cell fate and thermogenesis through the PRDM16 complex. Nature 504, 163-+.

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Wang Z.J., Zhong P., Ma K., Seo J.S., Yang F., Hu Z., Zhang F., Lin L., Wang J., Liu T., Matas E., Greengard P. & Yan Z. (2020) Amelioration of autism-like social deficits by targeting histone methyltransferases EHMT1/2 in Shank3-deficient mice. Mol Psychiatry 25, 2517-33.

 

doi: https://doi.org/10.1242/prelights.31778

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