Neurodevelopmental models of transcription factor 4 deficiency converge on a common ion channel as a potential therapeutic target for Pitt Hopkins syndrome

Abstract

The clinically pleiotropic gene, Transcription Factor 4 (TCF4), is a broadly expressed basic helix-loop-helix (bHLH) transcription factor linked to multiple neurodevelopmental disorders, including schizophrenia, 18q deletion syndrome, and Pitt Hopkins syndrome (PTHS). In vivo suppression of Tcf4 by shRNA or mutation by CRISPR/Cas9 in the developing rat prefrontal cortex resulted in attenuated action potential output. To explain this intrinsic excitability deficit, we demonstrated that haploinsufficiency of TCF4 lead to the ectopic expression of two ion channels, Scn10a and Kcnq1. These targets of TCF4 regulation were identified through molecular profiling experiments that used translating ribosome affinity purification to enrich mRNA from genetically manipulated neurons.Using a mouse model of PTHS (Tcf4+/tr), we observed a similar intrinsic excitability deficit, however the underlying mechanism appeared slightly different than our rat model - as Scn10a expression was similarly increased but Kcnq1 expression was decreased. Here, we show that the truncated TCF4 protein expressed in our PTHS mouse model binds to wild-type TCF4 protein, and we suggest the difference in Kcnq1 expression levels between these two rodent models appears to be explained by a dominant-negative function of the truncated TCF4 protein. Despite the differences in the underlying molecular mechanisms, we observed common underlying intrinsic excitability deficits that are consistent with ectopic expression of Scn10a. The converging molecular function of TCF4 across two independent rodent models indicates SCN10a is a potential therapeutic target for Pitt-Hopkins syndrome.

Diagnosis and management in Pitt‐Hopkins syndrome: First international consensus statement

“…Individuals with PTHS have mild to severe motor learning impairments and often engage in stereotypic and repetitive movements such as hand clapping and flapping, repeated hand to mouth movements, head shaking, head banging, body rocking, washing, finger crossing, and rubbing toes together . Motor milestones and self‐care skills are delayed (see Section 10).…”

“…Loss‐of‐function mutations in Tcf4 lead to the near‐complete lack of language acquisition. Language acquisition is virtually absent in most PTHS individuals . Up to 55% of individuals express only single words before 10 years of age, and indeed many have little to no expressive language throughout their lifespan ( R34 ).…”

“…Deficits in social and communication interaction skills combined with patterns of repetitive behavior, as well as impairments in adaptive skills are prominent in individuals with PTHS . Few studies have implemented direct in‐person assessments of autism characteristics, although various studies have reported on language development, insistence on sameness and on behavioral patterns of repetitive behaviors and stereotypies such as hand clapping and flapping, head banging, body rocking, or finger mannerisms …”

Molecular Mechanisms of Transcription Factor 4 in Pitt-Hopkins Syndrome

“…Originally generated as a model of constitutive germline deletion of one Tcf4 allele, our recent work has shown that a truncated TCF4 protein is in fact expressed in these mice[18]. Furthermore, we have shown this truncated protein is capable of binding to full length TCF4, confirming other previous reports that this transgenic PTHS mouse model may produce TCF4 haploinsufficiency through a dominant-negative binding mechanism[4, 18]. Although this model does not produce a complete allelic germline deletion, it still represents a powerful model for understanding PTHS biology, as the disruption of normal protein dimerization and DNA binding is a key element of the point mutations or other TCF4 gene disruptions that result in PTHS[4].…”

“…Mutations have typically been reported in the bHLH domain where the possibility for disruption of normal E-protein binding function is high[1, 4, 12, 15–17]. Not all mutations result in complete loss of TCF4 protein function, and evidence for dominant-negative effects of mutant protein has been observed[4, 18]. Reported PTHS-linked mutations that are located outside of this bHLH region have not been shown to lead to functional deficiencies in TCF4 protein, and how they result in disease remains an open question [4].…”