The genetics of situs inversus totalis without primary ciliary dyskinesia
Merel C. Postema1, Amaia Carrion-Castillo1, Simon E. Fisher1,2, Guy Vingerhoets3, Clyde Francks1,2
1 Department of Language & Genetics, Max Planck Institute for Psycholinguistics, Nijmegen, Netherlands 2 Donders Institute for Brain, Cognition and Behaviour, Radboud University, Nijmegen, Netherlands 3 Department of Experimental Psychology, Ghent University, Belgium North Sea Laterality Meeting 2018 Meeting Laterality Sea North
22 Aug 2018 | Merel Postema Situs inversus totalis
Definition Situs inversus totalis is classically reported as an autosomal recessive condition that is featured by a complete mirror- reversal of the internal organs. Symptoms • Mirror-patterning of the internal organs
Prevalence 2 • 1:6,000 – 8,000 people have SIT • 20-25% of people have SIT with PCD (= Kartagener) North Sea Laterality Meeting 2018 Meeting Laterality Sea North
2 Deng et al. Expert Rev Mol Med (2015) Primary Ciliary Dyskinesia
Definition Primary ciliary dyskinesia (PCD) is an autosomal recessive disorder that is featured by a deficit in the motile cilia
Symptoms 1 • Chronic bronchitis • Inflamed or infected sinuses • Male infertility
Prevalence 2 • 1:16,000 – 20,000 people have PCD
North Sea Laterality Meeting 2018 Meeting Laterality Sea North • 50% of people with PCD also demonstrate situs inversus totalis (SIT)
Figure copied from: plengdut.com
1 Reiter & Leroux, Nat Rev Mol Cell Biol (2017); 2 Deng et al. Expert Rev Mol Med (2015) Cilia and embryonic asymmetry
Nodal flow is central in LR symmetry breaking:
. North Sea Laterality Meeting 2018 Meeting Laterality Sea North
Figure copied from: plengdut.com
et al. et al. Fliegauf 2007, Hamada 2002 North Sea Laterality Meeting 2018 that have unrelated SIT to PCD reversebrainto the participantstypical in direction torqueof the particular in brain structural and functional asymmetriesassociated language,with seemsbut “We that visceralcompleteconclude reversal effect no lateralizationhas the on of .” 1 language dominance? braintorque and laterality, visceral of forsymmetry breaking Different mechanisms PCD unlinkedtoSIT with appearedHandedness non in (N=5/9 handedness levels of left showed increased SIT withoutPCD McManus, McManus, - 1 PCD SIT) Proc - Biol Sci (2004) Aims of the study
What mechanisms are involved in visceral laterality? • Identify novel (non-ciliary) genes/pathways involved in left-right asymmetry patterning • Examine the genetic relationship between brain and body asymmetry North Sea Laterality Meeting 2018 Meeting Laterality Sea North Study participants
Mean age = 33 y, Mean education = 13 y Sex: M = 8, F = 7 Handedness: R =11, L = 4
LH = 1 N = 6 with PCD (= Kartagener) North Sea Laterality Meeting 2018 Meeting Laterality Sea North N = 15 N = 15 LH = 5 N = 9 without PCD Vingerhoets et al., Brain Struct Funct (2018) Whole-genome sequencing Illumina next-generation sequencing North Sea Laterality Meeting 2018 Meeting Laterality Sea North WGS data analysis GATK Best Practices Workflow
Mean number of variants per genome = 5.18E6 North Sea Laterality Meeting 2018 Meeting Laterality Sea North WGS data analysis
Mean number of variants per genome = 5.18E6
Gene-set enrichment analysis: • Test whether mutated genes are functionally related across subjects, according to gene classification schemes • P-value is corrected for multiple testing of gene sets
Identify most likely causal gene per subject: North Sea Laterality Meeting 2018 Meeting Laterality Sea North • Genes already known to be mutated in PCD and/or SIT, heterotaxia Mean of 8 affected genes per subject (min=5, max=15) Results gene-set enrichment analysis SIT cases with PCD (N = 6)
model P-value Enrichment term N PCD cases recessive 4.32E-05 outer dynein arm assembly 5 recessive 6.27E-05 microtubule-based movement 6 recessive 0.00307 ciliary plasm 6 recessive 7.00E-05 dynein complex 5 recessive 0.0331 microtubule motor activity 4 recessive 0.021 dynein light chain binding 4 recessive 0.0101 Situs inversus totalis 6 recessive 0.000121 Ciliary dyskinesia 6 recessive 0.0218 Abnormal respiratory epithelium morphology 3 North Sea Laterality Meeting 2018 Meeting Laterality Sea North Results gene-set enrichment analysis All controls (N = 15)
model P-value Enrichment term N controls Recessive - No significant terms - North Sea Laterality Meeting 2018 Meeting Laterality Sea North Results gene-set enrichment analysis SIT cases without PCD (N = 9)
N non-PCD model P-value Enrichment term cases Recessive - No significant terms -
SIT cases without PCD & left handed (N=5)
model P-value Enrichment term N non-PCD cases Recessive - No significant terms - North Sea Laterality Meeting 2018 Meeting Laterality Sea North Most likely monogenic causes in cases Recessive model
Subject Phenotype Sex Age LI1 Edin Hand CHD Gene Type** ClinVar SI06 LRRC6 Kartagener Male 46 -0.82 1 R 0 Homozygous Kartagener SI08 DNAH11 Kartagener Female 23 -0.99 0.9 R 0 Homozygous PCD SI11 DNAAF1 Kartagener Female 32 -0.95 0.6 R 0 Comphet Kartagener SI13 CCDC114 Kartagener Male 48 -0.45 0.9 L* 0 Homozygous Kartagener SI15 DNAH5 Kartagener Female 31 -0.79 0.7 R 0 Comphet Kartagener SI17 DNAH5 Kartagener Male 39 -0.75 0.5 R 0 Comphet Kartagener SI02 PKD1L1 non-PCD SIT Male 50 -0.93 0.9 R 0 homozygous SA | SIT SI12 DNAH5 non-PCD SIT Female 40 0.88 -1 R 0 Comphet Kartagener SI16 CCDC151 non-PCD SIT Male 21 0.08 0.9 L 0 Homozygous Kartagener SI03 non-PCD SIT Female 26 -0.78 -0.8 L 0 Unsolved SI04 non-PCD SIT Male 23 -0.81 -1 L 1 Unsolved ** SI05 non-PCD SIT Male 27 -0.81 0.9 R 0 Unsolved SI07 non-PCD SIT Female 35 0.61 0.9 R 1 Unsolved SI09 non-PCD SIT Female 36 -0.33 0.7 L* 0 Unsolved SI14 non-PCD SIT Male 18 -0.44 -0.8 L 1 Unsolved North Sea Laterality Meeting 2018 Meeting Laterality Sea North
1 Vingerhoets et al., Brain Struct Funct (2018) Results gene-set enrichment analysis Unsolved cases (N=6)
N non-PCD model P-value Enrichment term cases Recessive - No significant terms -
N non-PCD model P-value Enrichment term cases Dominant - No significant terms - North Sea Laterality Meeting 2018 Meeting Laterality Sea North Summary [1/2]
• Some SIT-only cases had recessive mutations in known PCD genes • Six SIT-only cases had no obvious monogenic basis for their trait (recessive model) • No significant functional links between mutated genes across the unsolved cases • None of the unsolved cases has obvious candidate mutations linked to laterality phenotypes
• Most LH cases were unsolved, so we cannot link SIT to North Sea Laterality Meeting 2018 Meeting Laterality Sea North handedness through genetics after all. Summary [2/2]
• Perhaps recessive mutations in known PCD-genes (in non-PCD SIT cases) have lower penetrance for PCD, but are still causative for SIT • Some ciliary mutations may disrupt laterality without causing PCD-like symptoms • Environmental factors instead of monogenic basis • Few genetic epidemiology studies in SIT • More complex genetic model than monogenic MUTATIONAL LOAD TEST • Monogenic model is still possible, but would have to involve genetic heterogeneity across a set of genes which are not currently linked in terms of their known biology Non-coding variation (other than splice sites)
North Sea Laterality Meeting 2018 Meeting Laterality Sea North • • Structural variation STRUCTURAL VARIANT ANALYSIS (CNVATOR, LUMPY) Acknowledgements
• Amaia Carrion-Castillo • Simon E. Fisher • Guy Vingerhoets • Clyde Francks North Sea Laterality Meeting 2018 Meeting Laterality Sea North