The epilepsy phenotypic spectrum associated with a recurrent CUX2 variant

Abstract

Objective: Cut homeodomain transcription factor CUX2 plays an important role in dendrite branching, spine development, and synapse formation in layer II to III neurons of the cerebral cortex. We identify a recurrent de novo CUX2 p.Glu590Lys as a novel genetic cause for developmental and epileptic encephalopathy (DEE).

Methods: The de novo p.Glu590Lys variant was identified by whole-exome sequencing (n = 5) or targeted gene panel (n = 4). We performed electroclinical and imaging phenotyping on all patients.

Results: The cohort comprised 7 males and 2 females. Mean age at study was 13 years (0.5-21.0). Median age at seizure onset was 6 months (2 months to 9 years). Seizure types at onset were myoclonic, atypical absence with myoclonic components, and focal seizures. Epileptiform activity on electroencephalogram was seen in 8 cases: generalized polyspike-wave (6) or multifocal discharges (2). Seizures were drug resistant in 7 or controlled with valproate (2). Six patients had a DEE: myoclonic DEE (3), Lennox-Gastaut syndrome (2), and West syndrome (1). Two had a static encephalopathy and genetic generalized epilepsy, including absence epilepsy in 1. One infant had multifocal epilepsy. Eight had severe cognitive impairment, with autistic features in 6. The p.Glu590Lys variant affects a highly conserved glutamine residue in the CUT domain predicted to interfere with CUX2 binding to DNA targets during neuronal development.

Interpretation: Patients with CUX2 p.Glu590Lys display a distinctive phenotypic spectrum, which is predominantly generalized epilepsy, with infantile-onset myoclonic DEE at the severe end and generalized epilepsy with severe static developmental encephalopathy at the milder end of the spectrum. Ann Neurol 2018;83:926-934.

Figure 1. The recurrent Glu590Lys pathogenic missense variant and CUX2 structure

A. The CUX2 protein consists of three DNA-binding CUT domains (pink) and a single homeodomain (blue). Vertical lines below the protein show the location of missense variants that are reported more than twice in the Gnomad dataset. The CUX2 p.Glu590Lys pathogenic variant lies in the first CUT domain, these CUT domains are largely devoid of missense variants in the general population (data from gnomAD). B. The glutamine residue at amino acid position 590 (blue arrow) is fully conserved among cut-like Homeobox proteins across 19 species using Clustal alignment. GERP score GERP++=4.770, phyloP100way_vertebrate= 9.998, phastCons100way_vertebrate =1.000). C. High conservation of a negatively charge amino acid in third helix of all human CUT domains (IPR003350), as well as in related Cro/C1-type helix-turn-helix domain (IPR001387) (blue arrow). Clustal alignment of CUT domains of human ONECUT, SATB and CUX proteins, along with the H-L-H domain of bacteriophage lambda RPC1.

Figure 2. In silico predictions of the consequences of the p.Glu590Lys

A. Glu590 in CUT1 domain lays in the 3^rd helix of the CUT domain. Sequence Chain View of 1X2L Solution structure of the first CUT domain of human homeobox protein CUX2 (Cut-like 2). Alpha helix is not obviously modified by the Glu to Lys mutation. pdb-viewer superposition of 1X2L and the structure generated by MODELLER mutate_model.py script. B. Similar position in third helix of E54 in 1X2L structure (CUT domain of human homeobox protein CUX2) and D41 in 2D5V structure (rat HNF-6alpha DNA-binding domain in complex with the TTR promoter), as reported on PDB (http://www.ebi.ac.uk/pdbe/)) C. Three 3D views of 2D5V structure, from http://www.rcsb.org/pdb, showing the position of D41 within the major groove of DNA.