Functional Analysis of PTCHD1 Reveals Interactions with Synaptic Machinery and Involvement in the Hedgehog Pathway

Thursday, May 15, 2014
Atrium Ballroom (Marriott Marquis Atlanta)
K. Mittal1, K. Sritharan2, B. Degagne2 and J. B. Vincent3, (1)250, College Street, Centre for Addiction and Mental Health, Toronto, ON, Canada, (2)Neurogenetics, Centre for Addiction and Mental Health, Toronto, ON, Canada, (3)Centre for Addiction & Mental Health, Toronto, ON, Canada

Autism Spectrum Disorder (ASD) includes a range of complex neurodevelopment disorders. Approximately 75% have lifelong disability requiring substantial social and educational support.


This study is focused on investigating the complex functional aspects of a recently identified gene -- PTCHD1, and how its disruption leads to Autism Spectrum Disorder and/or Intellectual Disability. The insight gained will help in more precise phenotypic characterization of the individuals with disrupted PTCHD1.


Firstly, we sought to identify PTCHD1 splice variants that might have higher or more specific expression in brain, and may thus be more relevant to the neurobiology in individuals with autism and/or intellectual disability who have PTCHD1 mutations. We performed mRNA expression analysis using mRNA from multiple tissues, including brain regions. Western blots were performed to confirm these findings.

To establish the involvement of PTCHD1 in the Hedgehog (Hh) signaling  pathway, expression analysis was carried out with Hh pathway genes. We also probed for sub-cellular localization of PTCHD1 in cilia. Immunoprecipitation studies and expression studies were also performed to identify potential synaptic interacting partners or downstream targets.


We identified a new transcript skipping exon 2 which is predicted to encode a 542 amino acid protein in comparison to the 888 amino acid protein encoded by the PTCHD1 long isoform. It has just 4 transmembrane domains, and encodes a 62 kDa protein as compared to 12-transmembrane domains in the long isoform and a 101 kDa protein. We also found the presence of an additional exon upstream of exon 1. The quantitative expression analysis demonstrates that expression of these PTCHD1 transcripts is highest in human cerebellum.

The quantitative expression analysis with over expression of PTCHD1 revealed increased levels of  neuroligin and neurexin mRNAs. Immunoprecipitation studies indicate interactions including between PTCHD1 and Postsynaptic Density protein 95 (PSD-95) proteins. We also confirmed localization of PTCHD1 in cilia- which is where Hh receptors PTCH1 and 2 function, as well as evidence of PTCHD1 dimerization.


The new PTCHD1 transcripts seem to have an important role to play in the brain and in its sub regions, thus, these transcripts and encoded isoforms may be more relevant to autism and intellectual disability. The interacting partners would help us elucidate the etiology underlying the disease phenotype.

The quantitative expression analysis with over expression of PTCHD1 suggests a possible regulatory effect of PTCHD1 on genes that encode synaptic proteins. PSD-95 determines the size and strength of synapses in the postsynaptic density of neuronal excitatory synapses, and this putative interaction with the PTCHD1 protein could be important to elucidate disease etiology.

The Hh pathway plays an important role in embryonic development and adult stem cell functioning. Protein components of primary cilia are required for Hh signaling. We hypothesize that PTCHD1 localization to primary cilia could inhibit the Hh pathway in the developing brain, also that PTCHD1, as a chemosensor for extracellular SHh, may influence generation or functioning of synapses.