Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
Case Reports
. 2020 Mar:51:68-78.
doi: 10.1016/j.mito.2020.01.004. Epub 2020 Jan 7.

LONP1 de novo dominant mutation causes mitochondrial encephalopathy with loss of LONP1 chaperone activity and excessive LONP1 proteolytic activity

Arnaud Besse  1 , Daniel Brezavar  1 , Jennifer Hanson  1 , Austin Larson  2 , Penelope E Bonnen  3
Affiliations
Case Reports

LONP1 de novo dominant mutation causes mitochondrial encephalopathy with loss of LONP1 chaperone activity and excessive LONP1 proteolytic activity

Arnaud Besse et al. Mitochondrion. 2020 Mar.

Abstract

LONP1 is an ATP-dependent protease and chaperone that plays multiple vital roles in mitochondria. LONP1 is essential for mitochondrial homeostasis due to its role in maintenance of the mitochondrial genome and its central role in regulating mitochondrial processes such as oxidative phosphorylation, mitophagy, and heme biosynthesis. Bi-allelic LONP1 mutations have been reported to cause a constellation of clinical presentations. We report a patient heterozygous for a de novo mutation in LONP1: c.901C>T,p.R301W presenting as a neonate with seizures, encephalopathy, pachygyria and microcephaly. Assays of respiratory chain activity in muscle showed complex II-III function at 8% of control. Functional studies in patient fibroblasts showed a signature of dysfunction that included significant decreases in known proteolytic targets of LONP1 (TFAM, PINK1, phospho-PDH E1α) as well as loss of mitochondrial ribosome subunits MRPL44 and MRPL11 with concomitant decreased activity and level of protein subunits of oxidative phosphorylation complexes I and IV. These results indicate excessive LONP1 proteolytic activity and a loss of LONP1 chaperone activity. Further, we demonstrate that the LONP1 N-terminal domain is involved in hexamer stability of LONP1 and that the ability to make conformational changes is necessary for LONP1 to regulate proper functioning of both its proteolytic and chaperone activities.

Keywords: Chaperone; Encephalopathy; LONP1; Mitochondria; Oxidative phosphorylation; Protease; Seizures.

PubMed Disclaimer

Conflict of interest statement

Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Figures

Figure 1.
Figure 1.
Mitochondrial encephalopathy caused by de novo LONP1 mutation. A. CT and MRI of the brain of Subject with severe encephalopathy show pachygyria and small irregular calcifications in the periventricular white matter of the frontal lobes bilaterally. CT in the first days of life showed abnormal decrease in signal intensity and thickening of the white matter of the temporal lobes and posterior parietal lobes, and MRI of the brain at age 2 months showed progressive white matter volume loss. B. Electron transport chain activity was measured in muscle and in dermal fibroblasts. Citrate synthase and lactate dehydrogenase activities were also measured in both tissues. Dashed red line is placed at 40% of normal which is the threshold for Modified Walker clinical diagnostic criteria of being designated abnormal. C. Electron transport chain protein components were measured by Western blotting in three controls (C1, C2, C3) and Subject (LONP1) fibroblasts. D. Parents were shown to be negative for the mutation found in the Subject, LONP1 c.901C>T, p.R301W.
Figure 2.
Figure 2.
LONP1 gene and protein structure. A. The N-terminal domain of the protein is comprised of a mitochondrial localization signal (MTS) and a protein interaction domain that facilitates polypeptide binding (blue). The ATPase domain is shown in yellow and the proteolytic domain in the C terminus is shown in red. Mutations reported in patients with LONP1 related disease are noted. Black font and lines indicate mutations that were observed in patients with CODAS clinical presentation. Blue font and lines indicate mutations that were observed in patients with classical mitochondrial disease clinical presentation. Red is for the patient reported here as having a de novo dominant mutation and mitochondrial encephalopathy. PHYRE2 was used to generate the monomer three-dimensional structure of NM_004793, amino acids 124 – 959. B. Three-dimensional modeling of the hexamer of LONP1 protein sequence from NM_004793, amino acids 200 – 959. C. The difference in energy for protein folding between the reference sequence and the mutant, p.R301W. D. GERP and PhyloP both show constraint at nucleotide c.901C>T. E. Multiple species alignment of LONP1 nucleotide sequence centered on the LONP1 c.901C>T patient mutation shows conservation across species at this site.
Figure 3.
Figure 3.
mtDNA transcription, translation and mitophagy pathway targets of LONP1 are diminished in LONP1 Subject. A. Mitochondrial transcription, ribosome proteins, TFAM, MT-ND5, MT-COX1, MRPL44, MRPL11, were measured by Western blotting in three controls (C1, C2, C3) and Subject (LONP1) fibroblasts. mtDNA copy number in Subject fibroblasts (S) were the same as control fibroblasts (C) B. Mitophagy & Kreb’s cycle entry proteins PINK1 and phospho-PDH-E1α Ser293/Ser300 show deficiency in LONP1 Subject fibroblasts.
Figure 4.
Figure 4.
Wild type LONP1 does not rescue defects in LONP1 Subject cells and overexpression of LONP1 induces dysfunction. A. Electron transport chain proteins NDUFB8 which is part of complex I (C1) and COX4I1 which is part of complex IV (CIV). B. Proteins involved in mtDNA transcription, translation & maintenance TFAM, MRPL44 and MRPL11. C. Mitophagy proteins PINK1 and PDH-E1α Ser300 are not affected by overexpression of LONP1. Subject fibroblasts (S), control fibroblasts (C).
Figure 5.
Figure 5.
Oligomerization is changed in LONP1 mutants and TFAM binding appears normal. A. LONP1 oligomerization was assessed through co-transfection of HEK293T cells with expression vectors allowing the concomitant expression of tagged proteins: co-immunoprecipitation of concomitant expression of tagged proteins: 1) V5-LONP1 WT + FLAG-LONP1 mutants, and 2) V5-LONP1s + FLAG-LONP1s. B. Binding of LONP1 binding with TFAM was assessed similarly to oligomerization using co-transfection followed by co-immunoprecipitation of FLAG-TFAM + V5-LONP1 WT and mutants. Colocalization of LONP1 and TFAM was tested through co-transfection of plasmids with FLAG-tagged TFAM and V5-tagged (Green) LONP1 WT or V5-tagged LONP1 mutants. Immunoflourescence shows TFAM-FLAG expression in Red and V5-LONP1 expression in Green. Merged images are shown.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Bernier FP, Boneh A, Dennett X et al. (2002) Diagnostic criteria for respiratory chain disorders in adults and children. Neurology 59:1406–11 - PubMed
    1. Besse A, Wu P, Bruni F et al. (2015) The GABA Transaminase, ABAT, Is Essential for Mitochondrial Nucleoside Metabolism. Cell Metab 21:417–27 - PMC - PubMed
    1. Bienert S, Waterhouse A, de Beer TA et al. (2017) The SWISS-MODEL Repository-new features and functionality. Nucleic Acids Res 45:D313–D319 - PMC - PubMed
    1. Bonnen PE, Yarham JW, Besse A et al. (2013) Mutations in FBXL4 cause mitochondrial encephalopathy and a disorder of mitochondrial DNA maintenance. Am J Hum Genet 93:471–81 - PMC - PubMed
    1. Bota DA, Davies KJ (2002) Lon protease preferentially degrades oxidized mitochondrial aconitase by an ATP-stimulated mechanism. Nat Cell Biol 4:674–80 - PubMed

Publication types

Supplementary concepts

LinkOut - more resources