Biallelic variants in LIG3 cause a novel mitochondrial neurogastrointestinal encephalomyopathy
Authors: Bonora Elena , Chakrabarty Sanjiban , Kellaris Georgios , Tsutsumi Makiko , Bianco Francesca , Bergamini Christian , Ullah Farid , Isidori Federica , Liparulo Irene , Diquigiovanni Chiara , Masin Luca , Rizzardi Nicola , Cratere Mariapia Giuditta , Boschetti Elisa , Papa Valentina , Maresca Alessandra , Cenacchi Giovanna , Casadio Rita , Martelli Pierluigi , Matera Ivana , Ceccherini Isabella , Fato Romana , Raiola Giuseppe , Arrigo Serena , Signa Sara , Sementa Angela Rita , Severino Mariasavina , Striano Pasquale , Fiorillo Chiara , Goto Tsuyoshi , Uchino Shumpei , Oyazato Yoshinobu , Nakamura Hisayoshi , Mishra Sushil K , Yeh Yu-Sheng , Kato Takema , Nozu Kandai , Tanboon Jantima , Morioka Ichiro , Nishino Ichizo , Toda Tatsushi , Goto Yu-ichi , Ohtake Akira , Kosaki Kenjiro , Yamaguchi Yoshiki , Nonaka Ikuya , Iijima Kazumoto , Mimaki Masakazu , Kurahashi Hiroki , Raams Anja , MacInnes Alyson , Alders Mariel , Engelen Marc , Linthorst Gabor , de Koning Tom , den Dunnen Wilfred , Dijkstra Gerard , van Spaendonck Karin , van Gent Dik C , Aronica Eleonora M , Picco Paolo , Carelli Valerio , Seri Marco , Katsanis Nicholas , Duijkers Floor A M , Taniguchi-Ikeda Mariko , De Giorgio Roberto 
Published: February 2021

Abstract
Abnormal gut motility is a feature of several mitochondrial encephalomyopathies, and mutations in genes such as TYMP and POLG, have been linked to these rare diseases. The human genome encodes three DNA ligases, of which only one, ligase III (LIG3), has a mitochondrial splice variant and is crucial for mitochondrial health. We investigated the effect of reduced LIG3 activity and resulting mitochondrial dysfunction in seven patients from three independent families, who showed the common occurrence of gut dysmotility and neurological manifestations reminiscent of mitochondrial neurogastrointestinal encephalomyopathy. DNA from these patients was subjected to whole exome sequencing. In all patients, compound heterozygous variants in a new disease gene, LIG3, were identified. All variants were predicted to have a damaging effect on the protein. The LIG3 gene encodes the only mitochondrial DNA (mtDNA) ligase and therefore plays a pivotal role in mtDNA repair and replication. In vitro assays in patient-derived cells showed a decrease in LIG3 protein levels and ligase activity. We demonstrated that the LIG3 gene defects affect mtDNA maintenance, leading to mtDNA depletion without the accumulation of multiple deletions as observed in other mitochondrial disorders. This mitochondrial dysfunction is likely to cause the phenotypes observed in these patients. The most prominent and consistent clinical signs were severe gut dysmotility and neurological abnormalities, including leukoencephalopathy, epilepsy, migraine, stroke-like episodes, and neurogenic bladder. A decrease in the number of myenteric neurons, and increased fibrosis and elastin levels were the most prominent changes in the gut. Cytochrome c oxidase (COX) deficient fibres in skeletal muscle were also observed. Disruption of lig3 in zebrafish reproduced the brain alterations and impaired gut transit in vivo. In conclusion, we identified variants in the LIG3 gene that result in a mitochondrial disease characterized by predominant gut dysmotility, encephalopathy, and neuromuscular abnormalities.