Complex I deficiency is a common cause of mitochondrial disease, resulting from mutations in genes encoding structural subunits, assembly factors or defects in mitochondrial gene expression. Advances in genetic diagnostics and sequencing have led to identification of several variants in NUBPL (nucleotide binding protein-like), encoding an assembly factor of complex I, which are potentially pathogenic. To help assign pathogenicity and learn more about the function of NUBPL, amino acid substitutions were recreated in the homologous Ind1 protein of the yeast model Yarrowia lipolytica. Leu102Pro destabilized the Ind1 protein, leading to a null-mutant phenotype. Asp103Tyr, Leu191Phe and Gly285Cys affected complex I assembly to varying degrees, whereas Gly136Asp substitution in Ind1 did not impact on complex I levels nor dNADH:ubiquinone activity. Blue-native polyacrylamide gel electrophoresis and immunolabelling of the structural subunits NUBM and NUCM revealed that all Ind1 variants accumulated a Q module intermediate of complex I. In the Ind1 Asp103Tyr variant, the matrix arm intermediate was virtually absent, indicating a dominant effect. Dysfunction of Ind1, but not absence of complex I, rendered Y. lipolytica sensitive to cold. The Ind1 Gly285Cys variant was able to support complex I assembly at 28°C, but not at 10°C. Our results indicate that Ind1 is required for progression of assembly from the Q module to the full matrix arm. Cold sensitivity could be developed as a phenotype assay to demonstrate pathogenicity of NUBPL mutations and other complex I defects.

Despite combination antiretroviral therapies making HIV a chronic rather than terminal condition for many people, the prevalence of HIV-associated neurocognitive disorders (HAND) is increasing. This is especially problematic for children living with HIV. Children diagnosed HAND rarely display the hallmark pathology of HIV encephalitis in adults, namely infected macrophages and multinucleated giant cells in the brain. This finding has also been documented in rhesus macaques infected perinatally with simian immunodeficiency virus (SIV). However, the extent and mechanisms of lack of susceptibility to encephalitis in perinatally HIV-infected children remain unclear. In the current study, we compared brains of macaques infected with pathogenic strains of SIV at different ages to determine neuropathology, correlates of neuroinflammation, and potential underlying mechanisms. Encephalitis was not found in the macaques infected within 24 h of birth despite similar high plasma viral load and high monocyte turnover. Macaques developed encephalitis only when they were infected after 4 months of age. Lower numbers of CCR5-positive cells in the brain, combined with a less leaky blood-brain barrier, may be responsible for the decreased virus infection in the brain and consequently the absence of encephalitis in newborn macaques infected with SIV.

Background

The nucleotide binding protein-like (NUBPL) gene was first reported as a cause of mitochondrial complex I deficiency (MIM 613621, 618242) in 2010. To date, only eight patients have been reported with this mitochondrial disorder. Five other patients were recently reported to have NUBPL disease but their clinical picture was different from the first eight patients. Here, we report clinical and genetic findings in five additional patients (four families).

Methods

Whole exome sequencing was used to identify patients with compound heterozygous NUBPL variants. Functional studies included RNA-Seq transcript analyses, missense variant biochemical analyses in a yeast model (Yarrowia lipolytica) and mitochondrial respiration experiments on patient fibroblasts.

Results

The previously reported c.815-27T>C branch-site mutation was found in all four families. In prior patients, c.166G>A [p.G56R] was always found in cis with c.815-27T>C, but only two of four families had both variants. The second variant found in trans with c.815-27T>C in each family was: c.311T>C [p.L104P] in three patients, c.693+1G>A in one patient and c.545T>C [p.V182A] in one patient. Complex I function in the yeast model was impacted by p.L104P but not p.V182A. Clinical features include onset of neurological symptoms at 3-18 months, global developmental delay, cerebellar dysfunction (including ataxia, dysarthria, nystagmus and tremor) and spasticity. Brain MRI showed cerebellar atrophy. Mitochondrial function studies on patient fibroblasts showed significantly reduced spare respiratory capacity.

Conclusion

We report on five new patients with NUBPL disease, adding to the number and phenotypic variability of patients diagnosed worldwide, and review prior reported patients with pathogenic NUBPL variants.