The ontogenesis of asymmetry in humans - a genetic and epigenetic perspective Inaugural – Dissertation zur Erlangung des Grades eines Doktors der Naturwissenschaften in der Fakultät für Psychologie der RUHR-UNIVERSITÄT BOCHUM vorgelegt von: Judith Schmitz, M.Sc. Psychologie Bochum, Mai 2018 I Gedruckt mit Genehmigung der Fakultät für Psychologie der RUHR-UNIVERSITÄT BOCHUM Referent: PD Dr. Sebastian Ocklenburg Korreferent: Prof. Dr. Robert Kumsta Termin der mündlichen Prüfung: 25.07.2018 II Cover illustration: The figure was used with permission of Prof. Dr. Be- ate Brand-Saberi and Dr. Nenad Maricic, Department of Anatomy and Molecu- lar embryology, Ruhr University Bochum. III Table of Contents Chapter 1 General introduction 1 1.1. Hemispheric asymmetries – the basics 2 1.1.1. Handedness 3 1.1.2. Language lateralization 4 1.2. The development of asymmetry 6 1.2.1. The emergence of visceral asymmetries 7 1.2.2. The emergence of structural hemispheric asymmetries 7 1.2.3. The emergence of motor asymmetries 8 1.2.4. The emergence of language lateralization 9 1.3. Genetics 10 1.3.1. The genetics of handedness 10 1.3.2. The genetics of language lateralization 13 1.3.3. The molecular link between visceral and hemispheric 16 asymmetries 1.4. Hemispheric asymmetries in gene expression 19 1.4.1. Lateralized gene expression in the fetal cortex 19 1.4.2. Lateralized gene expression in the fetal spinal cord 20 1.4.3. Relevance of lateralized gene expression for behavioral 21 asymmetry 1.5. Gene Ontology: Considering gene functions 23 1.6. The role of epigenetic regulation 25 1.6.1. Epigenetic mechanisms – the basics 26 1.6.2. Epigenetics in the development of hemispheric 27 asymmetries 1.7. Aim of this thesis 29 Chapter 2 The functional genetics of handedness and language lat- 31 eralization Chapter 3 Towards an epigenetic understanding of handedness on- 61 togenesis IV Chapter 4 DNA methylation in candidate genes for handedness 123 predicts handedness direction Chapter 5 KIAA0319 promoter DNA methylation predicts dichotic 149 listening performance in forced-attention conditions Chapter 6 General discussion 171 6.1. Summary of key findings 172 6.2. Molecular interrelations between asymmetry phenotypes 173 6.2.1. Handedness and language lateralization: FOXP2 and 173 LRRTM1 6.2.2. Visceral and hemispheric asymmetries: KIAA0319 175 6.2.3. Implications for phenotyping 177 6.2.4. Molecular pathways 179 6.3. The role of epigenetics 181 6.3.1. The role of birth stress 181 6.3.2. DNA methylation in twins 182 6.4. Transgenerational epigenetic inheritance – is that 184 possible? 6.4.1. Germ line epigenetic inheritance 184 6.4.2. Experience-dependent epigenetic inheritance 187 6.4.3. Gene-dependent epigenetic inheritance 189 6.5. Conclusion and outlook 192 6.5.1. Epigenetics in human cerebral tissue 193 6.5.2. Perspectives in animal research 195 References 196 Appendix 215 V Chapter 1 General introduction 1 Chapter 1 1.1. Hemispheric asymmetries – the basics In 1866, German zoologist Ernst Haeckel introduced promorphology – the sci- ence of an organism’s external form - and proposed symmetry as a fundamental criterion for classifying organisms (Haeckel, 1866). The clade of Bilateria (ani- mals displaying mirror-inverted body halves) including (but not restricted to) all vertebrates was created in 1888 (Hatschek, 1888). Besides asymmetry (organ- isms without any axis or plane of symmetry, e.g. the majority of sponges) and radial symmetry (organisms with one axis, but several planes of symmetry, e.g. starfish), bilateral symmetry is considered one of the three major types of body plans (Manuel, 2009). However, bilateral symmetry is frequently broken by ei- ther the position of non-paired internal organs in one body half (e.g., the left- sided stomach and the right-sided liver) or by anatomical differences between the left and right half of paired internal organs. For example, the human lungs are constituted of two lobes on the left and three lobes on the right side. Based on these observations, humans and other vertebrates have also been described as “pseudo-bilateral” (see Figure 1.1) (Levin, 2005). Figure 1.1: The major types of body plans. A) asymmetry. B) radial symmetry. C) bilat- eral symmetry. D) Due to the asymmetrical position of internal organs, humans and other vertebrates have been described as pseudobilateral (Levin, 2005). The brain is one of the most striking examples for structural and functional differences between the left and right half of the body. Hemispheric asymme- tries are found in several aspects of cognition such as memory, emotion, atten- tion, language and executive functions (Ocklenburg, Hirnstein, Beste, & Güntürkün, 2014). Importantly, hemispheric asymmetries are never absolute but relative, meaning that while one hemisphere is dominant for a given func- tion, the other hemisphere still contributes to this function. Moreover, while one 2 General introduction hemisphere is typically dominant in the majority of individuals, some individu- als display the opposite pattern. Among behavioral indicators of hemispheric asymmetries in humans, handedness is by far the most widely investigated (Cor- ballis, 2014). 1.1.1. Handedness Human handedness is not only found at the individual level, meaning that an individual prefers one hand over the other for fine motor tasks, but also on the population level. Approximately 90% of humans are right-handed, which has been stable for at least ten thousand years (Faurie & Raymond, 2004). Both indi- vidual-level and population-level asymmetries in limb preference have been found across the vertebrate lineage, however, with less strongly pronounced population-level asymmetries (Ströckens, Güntürkün, & Ocklenburg, 2013). The proportion of human left-handedness differs slightly across cultures, most likely due to external pressure and stigma against left-hand use (Raymond & Pontier, 2004). However, even in the most permissive countries, the proportion of left- handedness for throwing or hammering does not exceed 25% (Raymond & Pon- tier, 2004). Moreover, a large-scale meta-analysis found robust sex differences with a male to female odds ratio of 1.23 for left-handedness, suggesting a higher frequency among males (Papadatou-Pastou, Martin, Munafò, & Jones, 2008). An over-representation of left-handedness in psychological and neurodevelopmen- tal disorders has been confirmed by two meta-analyses for schizophrenia (Dra- govic & Hammond, 2005; Sommer, Ramsey, Kahn, Aleman, & Bouma, 2001), while it is more controversial for other disorders such as bipolar disorder (Nowakowska et al., 2008) or dyslexia (Geschwind & Behan, 1982). When being asked about their handedness, most people would respond based on their hand used for writing. However, there has been massive pressure towards right-hand use especially for writing, which could have distorting ef- fects on any kind of handedness research. Thus, research has either focused on hand preference for multiple activities or on hand performance (Ocklenburg, Beste, & Arning, 2014). Hand preference is most commonly determined using the Edinburgh handedness inventory (EHI) (Oldfield, 1971). In this question- naire, participants are asked to indicate their preferred hand for ten different activities. Based on the relation between left-hand and right-hand preferences, a 3 Chapter 1 lateralization quotient (LQ) is determined, ranging from -100 (left-hand prefer- ences only) over 0 (same amount of left- and right-hand preferences) to +100 (right-hand preferences only). While the extreme values (-100, +100) are defined as consistent left- and right-handedness, LQs between -99 and -60, or +60 and +99 are typically defined as moderate left- or right-handedness, while LQs be- tween -60 and +60 are indicative of ambidexterity (Isaacs, Barr, Nelson, & Devin- sky, 2006). The most common task used for the determination of hand perfor- mance is the Pegboard task (Annett, 1985). Participants are instructed to move pegs from one row of holes to another with either the left or the right hand. A quantitative measure of fine motor skill is obtained by relating the times re- quired to complete left- and right-hand trials resulting in the so-called PegQ measure. In contrast to hand preference, which usually has a J-shaped distribu- tion with a small number of left-handers, very few ambidextrals and a pro- nounced majority of right-handers, PegQ is normally distributed, making it ide- ally suited for genetic association studies (Paracchini, Diaz, & Stein, 2016). An- other commonly used task is the Tapley & Bryden test. In this paper and pencil test, participants are instructed to place dots in as many small circles as possible within a given time frame (Tapley & Bryden, 1985). 1.1.2. Language lateralization Since Pierre Paul Broca famously stated that nous parlons avec l’hémisphère gauche in 1868, left-hemispheric dominance for language has been one of the most ro- bust findings in laterality research. In language-impaired patients’ brains, Broca identified left-hemispheric lesions in a frontal area that should soon be named Broca’s area. Today, we know that while language is predominantely processed in the left hemisphere, there are also individuals with right-hemispheric or bi- lateral organization of language (Ocklenburg & Güntürkün, 2018). Historically, the hemisphere dominant for language has been assessed using the Wada test in epilepsy patients before surgery. However, as this test involves the injection of sedative into the ipsilateral carotid artery, several non-invasive