Glossary

Motor and vocal tics: A simple motor ticis a sudden, brief, involuntary, repetitive, nonpurposeful movement of a single muscle group such as an eye blink, face twitch, shoulder shrug, arm or leg jerk etc. Complex tics include forced touching, pulling clothes, a whole body jump or an abnormal walk. Vocal tics are involuntary sounds produced by moving air through the nose, mouth, or throat, or vocalizations. These are also called phonic tics and examples include throat clearing, grunting and coughing. Tourette syndrome affects ~1% of the school aged population and ~10% of these require lifelong therapy.

Echolalia: The automatic repetition of vocalizations made by another person

Stereotypies: These are repetitive and ritualistic movement or posture such as body rocking, or swaying movements, or crossing and uncrossing of legs etc.

Obsessive-compulsive disorder (OCD): A disorder characterized by intrusive, persistent thoughts (obsessions) and/or repetitive, intentional behaviours (compulsions) that result in significant distress or dysfunction. It affects 1 to 3% of the general population.

Attention Deficit Hyperactivity Disorder (ADHD): A disorder characterized byinattention and/or hyperactivity and impulsivity affecting around 5% of school aged children and causing impairment in social and academic performance; the symptoms may persist into adult life

Autism Spectrum Disorder (ASD): A developmental disorder characterized by abnormalities in social interactions and communication, as well as restricted interests and repetitive behaviours.

Synapse: Synapse formation is the key step in the development of neural networks.Synapses are specialized intercellular junctions in which cell adhesion molecules connect the presynaptic machinery of neurons for neurotransmitter release to the postsynaptic machinery for receptor signalling.

Striatum: A subcortical structure of the brain which is part of the basal ganglia system and is divided into caudate nucleus and putamen by a white matter tract called the internal capsule.

Cortices: The outer layer of the cerebral cortex composed of gray matter.

Nested Genes: A nested gene is defined as any gene located wholly within another gene. Nested genes are usually located within an intron of the host gene. Nested genes are relatively common within the genome and are usually coded on the complementary strand and transcribed in an antisense direction relative to the host gene [57]. Interestingly, nested genes often display high levels of tissue-specific expression [27]. Two hypotheses have been proposed for interactive expression of nested gene pairs. The functional co-regulation hypothesis predicts a positive correlation between levels of expression in different tissues (eg. BMCC1 and PCA3 [26] and the transcriptional collision/interference hypothesis predicts a negative correlation (eg. LRRN3 and IMMP2L) [59, 61]. Overlapping genes are four times more likely to be co-expressed than expected by random probability, however, little is known regarding the mechanism of co-regulation [57] or whether co-regulation and transcriptional interference operate simultaneously thereby constraining gene expression within the normal range. Transcriptional interference between the gene pairs has been investigated in bacteria and might take place by direct competition for the transcription apparatus and/or by formation of doublestrandedRNAs. Nesting may also serve to insulate the nested gene from cis or trans acting variations in the greater genome.

Leucine-rich repeats (LRRs) are common protein-protein interaction domains found in proteins with diverse structure and function. The LRRs are typically 20-29 amino acids in length and contain a conserved consensus sequence of LxxLxLxxN/CxL (where x can be any amino acid and L can be replaced by V, I or F). There are several subgroups of LRR proteins, differentiated by the consensus sequence and the inclusion of different combinations of supplementary domains (Fig. 2). One important subgrouping is based on the presence of a transmembrane domain. Different families within this subgroup of transmembrane LRR proteins include the AMIGO, NGL, LINGO, LRIG, FLRT, PAL, SALM, SLITRK, LRRN, LRRTM and LRTM gene families [64-67]. Multiple LRR motifs located in the N-extracellular region of these transmembrane proteins (Fig. 2)often assemble into a functional domain called a LRR domain,which typically has a horseshoe shape and is amenable for protein–proteininteractions[67-70, 75-76].Many LRR transmembrane proteins are brain enriched and/or highly expressed in the nervous system and have been implicated in nervous system development and different neural diseases including hereditary lateral temporal epilepsy and Parkinson disease [64].

The LRRTMs (leucine rich repeat transmembrane neuronal family) [65]represent a highly conserved four-member family which, with the exception of LRRTM4, are nested in the introns of different α-catenin genes. Catenin family members are adhesion proteins that can form a complex with cadherins, which themselves have been implicated in intellectual disability, autism and ASD risk. LRRTM mRNAs are mainly expressed in the nervous system, each with a distinct and highly regulated pattern. LRRTMs are synaptic organizing molecules and synaptogenic inducers in neurons, initiating excitatory presynaptic differentiation and mediating post-synaptic specializations. LRRTM1and LRRTM4are both located on 2p12 (Table 2). LRRTM1is located within intron 7 of CTNNA2 (α2-catenin) and is highly expressed within the brain and salivary gland [65]. LRRTM1 is associated with both human handedness (relative hand skill) and schizophrenia and is thought to be involved in brain development, neuronal connectivity, intracellular trafficking in axons and synaptogenesis. There is also evidence for association between LRRTM1 and abnormal asymmetrical brain structure in language-associated areas. LRRTM3, located on 10q22.1, is positioned within intron 7 of CTNNA3 (αT-catenin). This gene has a more restricted expression profile compared to LRRTM1, with expression in the brain, particularly the cerebellum. In addition, LRRTM3 is functionally and positionally linked to late-onset Alzheimer's disease [65].

The LRRN (leucine rich repeat neuronal) family of four are all brain-enriched type I transmembrane proteins. Although they seem to function as adhesion molecules or binding receptors in regulatory mechanisms, their biological activities and specific central nervous system (CNS) functions in humans are still unclear. LRRN1, located on 3p26.2, is nested within intron 8 of the long form of SUMF1 (sulphatase modifying factor 1). Lrrn1is dynamically expressed in the somites and the neural plate during development in the mouse, and mostly in the brain and kidney of the adult. LRRN1 is also noteworthy due to its location within a region (3p26.1-26.2) duplicated in children with paternally inherited and in a candidate region for recessive non-syndromic mental retardation. Chromosome 3p26.1-26.2 was also identified as a linkage region of interest in the TSAICG cohort of sib pairs (Table 4). LRRN3, located at 7q31.1, is nested within intron 3 of IMMP2L (inner mitochondrial membrane peptidase-like). Lrrn3 exhibits regulated expression in the developing ganglia and motor neurons of the neural system, and is upregulated during neuronal cortical injury [64].

The LRTM (leucine-rich repeats and transmembrane domains) family is a highly conserved two-member family which are both nested in introns of different CACNA2D (calcium channel, voltage-dependent, alpha 2/delta subunit) genes. LRTM1, nested within intron 23 of CACNA2D3 on chromosome 3p14 is expressed within the brain. LRTM2 is nested within intron 23 of CACNA2D3 on chromosome 12p13.

Fig. 2 Schematic showing the domain architecture of the various neuronal LRR transmembrane protein families discussed in this review

Nesting of LRR genes: The LRRN1, LRRN3, LRRTM1, LRRTM3 and LRTM1genes all share the curious structural relationship of being nested in a n antisense orientation within the introns of other genes. Among the ~313 LRR coding genes in the human genome the transmembrane subgrouping appears most relevant to neuronal development and synaptogenesis[64-67, 77]. This transmembrane subgrouping of LRR protein genes is further classified into various gene families on the basis of the sequence and structure of their LRR domains and other supplementary domains [64-67]. Of these LRR transmembrane genes we are aware of only seven that are nested within other genes. These seven nested genes are clustered within three gene families, namely the LRRTMs, LRRNs and LRTMs[64-67] all of which have been associated with TS often through multiple means of enquiry (Tables 1-3). There is also evidence for the convergent evolution of 'LRR gene nesting'. For example, between the three LRR coding gene families this nesting occurs within unrelated classes of harbouring genes and also within the LRRN gene family. This suggests that this nesting may serve an important regulatory function involved in neurodevelopment as it pertains to the pathogenesis of TS. Given the vulnerability of behavioural and neuropsychiatric disorders like TS to changes in gene expression – as evidenced by the high incidence and pathogenesis of gene copy number variations - one possibility is that the nested/overlapping status of these LRR genes within other genes may serve to constrain their transcription within the normal range (see Glossary for Nested genes). Conversely, when the host gene is disrupted and its transcriptional influence over the nested gene ceases, as described here in TS, such putative constraints on the nested gene would be removed. Such nesting may also provide a form of insulation from more distant variations in the surrounding genome like CNVs and breakpoints that might otherwise alter nested gene expression even from very long distances [49].