An interesting article written by Monique Balemans titled “Phenotyping the EHMT1 Mouse – a model for Kleefstra syndrome.

Kleefstra syndrome (KS) is a form of syndromic intellectual disability (ID), with moderate to severe Intellectual Disability, general developmental delay, childhood hypertonia and craniofacial abnormalities as core features. In addition, patients can display heart defects, renal/urologic defects, genital defects in males, severe respiratory infections, epilepsy, sleep disturbances, absence of speech, autistic-like features in childhood, and in few cases extreme apathy or catatonic-like features after puberty.

The syndrome is caused by deletion or mutation of one of the two copies of the Euchromatin histone methyltransferase 1 (EHMT1) gene. In every cell of our body, the DNA is wrapped around histone proteins. These histones have so-called tails, that protrude from the complex and are susceptible for modification. The EHMT1 protein is able to mono- and dimethylate lysine 9 of the histone H3 N-terminal tail. Following this modification, the DNA-histone complex is compacted and the genetic information at that specific site is (temporarily) not available any more. This process is called gene silencing. Thus, haploinsufficiency of the EHMT1 gene will probably result in aberrant gene expression.

The mechanisms underlying the many features observed in KS patients are yet unknown. These need to be elucidated in order to develop potential therapeutic strategies. Studies in humans have limited possibilities for investigations that aim to explain basic mechanisms of brain disorders. Thus, animal models are invaluable for improving our understanding of the disease pathology. Mouse Ehmt1 is highly similar to the human EHMT1 gene, with a 97% homology in amino acid sequence. Therefore we chose to phenotype Ehmt1 heterozygous knockout (Ehmt1+/-) mice as a mammalian model for KS. Using these mice, we wanted to obtain more insight into Ehmt1 protein function, with a focus on its role in neuronal functioning and behaviour.

Our initial studies pointed at altered exploration/anxiety behaviour in the Ehmt1+/- mice. Therefore we investigated whether the autistic-like features observed in KS patients were mimicked in these mice, by performing a battery of behavioural tests. The Ehmt1+/- mice showed reduced activity and exploration, and increased anxiety compared with normal (wildtype) mice. They also demonstrated diminished social play when encountering a mouse from a different litter, and a delayed or absent response to social novelty when exposed to a stranger mouse. This indicated that the hyperactivity and autistic-like features of KS patients are recapitulated in the Ehmt1+/- mice.

Patients with KS show diminished cognitive functioning, presenting as moderate to severe ID. In agreement with this, Ehmt1+/- mice displayed deficits in several learning and memory tests. These deficits were associated with a reduced complexity of neurons in a brain area called the hippocampus. Further analysis of these neurons revealed a reduced number of contact points between the neurons, and alterations in functioning of these cells. These data showed indications for an ID-like phenotype in the Ehmt1+/- mice.

In addition to ID, KS patients show three more core features i.e. general developmental delay, hypertonia, and craniofacial abnormalities. Our studies revealed that young Ehmt1+/- mice also showed a delayed postnatal development and features of hypertonia. Furthermore, we found that Ehmt1+/- mice showed brachycephalic crania, a shorter nose, and hypertelorism, reminiscent of the craniofacial dysmorphisms seen in KS. This aberrant bone formation was further nvestigated by performing a gene expression analysis. Interestingly, the mRNA levels of several bone tissue related genes were higher in the Ehmt1+/- mice, which may have contributed to the craniofacial abnormalities observed in these mice.
Since Ehmt1+/- mice display both developmental and behavioural deficits, the Ehmt1 protein probably plays an important role in brain development and neuronal functioning. We investigated brain morphology and layering of wildtype and Ehmt1+/- mice, but did not observe gross morphological differences. A specific staining for the Ehmt1 protein in the brain showed that it is present in all parts of the brain in adult and aged mice. Interestingly, we observed that Ehmt1 is highly expressed in two brain regions that harbor newborn neurons and/or neuronal stem cells, indicating that it may play a role in these cells.

In conclusion, the Ehmt1+/- mice recapitulate many features of KS patients, including ID, general developmental delay, childhood hypotonia, craniofacial dysmorphisms, hypoactivity, and autistic-like features. These mice therefore represent a valuable mammalian model for this disorder. Furthermore, studies with brain and bone tissue of the Ehmt1+/- mice suggest that Ehmt1 controls the expression of many different genes. Hopefully, future studies with the Ehmt1+/- mice will advance toward understanding disease pathology and eventually the development of therapeutic strategies for patients with KS.

Monique Balemans,
August 2013