Genes and related mitochondrial dysfunction syndromes.
Multiple mitochondrial dysfunctions syndrome (MMDS) is a group of autosomal recessive mitochondrial disorders that is associated with deficiencies related to nuclear genes: ISCA2, ISCA1, NFU1, IBA57, and BOLA3. The syndromes are relatively new and recently discovered. Individuals with MMDS have reduced function of energy production stages in mitochondria. The dysfunctions are mostly related to iron-sulfur (Fe-S) clustering system (ISC) and its biogenesis. The signs and symptoms of the patients may begin early in life, and can be quite severe leading to death more or less during infancy. Affected individuals have various symptoms including brain dysfunction (encephalopathy), hypotonia, seizures, delayed developmental milestones, and cognition and psychomotor impairments. These individuals often have difficulty growing and gaining weight at the expected rate. Diagnosis of the disease can be challenging as in the case with most of the mitochondrial disorders. However, since the genetic causes of the MMDS are known, a laboratory test focusing on the causative genes will be helpful to determine the pathogenic mutations. This in turn would facilitate reducing the number of the diseases through carrier testing and genetic counseling and utilization of preimplantation genetic diagnosis in populations, especially those that display high rate of consanguinity, which are prone to have such autosomal recessive disorders.
- iron-sulfur (Fe-S) cluster (ISC)
- multiple mitochondrial dysfunction syndromes
Mitochondria are double membrane-bound cellular organelles surrounded by outer and inner membranes [1, 2]. The organelle is considered cell’s powerhouse generating adenosine triphosphate (ATP) during cellular respiration; hence, facilitating energy conversion in eukaryotes. Uniquely, each mitochondrion has its own DNA and encodes mitochondrial genes; hence, contributing the cell’s proteome independently. The inheritance of the mitochondrial genome differs from nuclear genome since the donor of mitochondrial DNA (mtDNA) is the egg rather than sperm whose mitochondria are marked for obliteration upon entering the egg . Hence, the organelle’s DNA is inherited through females known as “maternal inheritance.” Since these organelles generate energy, most biochemical reactions in the eukaryotic cells occur in the mitochondria. These reactions include pyruvate oxidation, citric acid cycle, electron transport, and oxidative phosphorylation (OXPHOS) all needed for energy production. Mitochondria also have an important role in calcium signaling, regulation of cellular metabolism, heme synthesis, steroid synthesis, apoptosis, and the biosynthesis of iron-sulfur (IS) clusters (ISC). The high number of human diseases caused by the malfunction of the mitochondrial proteins—encoded by nuclear or mtDNA—drew attention to the importance of this organelle.
Mitochondria are genetically controlled by both nuclear DNA and the mitochondrial genome [1, 4]. A wide range of molecular defects have been identified in the human mitochondrial genome [4, 5, 6, 7, 8, 9]. Diseases due to mutations in the mitochondrial genome are clinically, genetically, and biochemically diverse [1, 2, 4, 6, 10]. Similarly, deficiencies in mitochondrial genes encoded by nuclear genome can also lead various mitochondrial disorders and a wide range of cellular perturbations such as undue reactive oxygen species and distracted apoptosis, aberrant calcium homeostasis, and deficient energy production. This in turn leads failure to meet the requirements of numerous organs, especially those with high energy needs. Hence, various pathological conditions appears due to impaired mitochondrial function in human body involving different cell types, tissues, and organs including heart and brain. Such multi-organ manifestations are all mitochondria related and these diseases varies from epilepsy to cardiac myopathies.
3. Mitochondria and genetics of mitochondria-related diseases
The mitochondrial genome is a multicopy, double-stranded circular DNA molecule, which is 16.6 kb in human . This genome encodes 13 essential proteins for the OXPHOS system and 24 components of the RNA machinery: 2 rRNAs and 22 tRNAs . It is intronless and the only noncoding region is the displacement region (D-Loop), a region of 1.1 kb. It contains both the replication origins and the transcriptional promoters. Although mitochondria are genetically controlled by both mitochondrial and nuclear genomes, mtDNA is only maternally inherited . Mitochondrial genetics differ greatly from Mendelian genetics in size, number of encoded genes, number of DNA molecules per cell, lack of introns, gene density, replication, transcription, recombination, and mode of inheritance. The 13 proteins include 7 subunits of NADH Dehydrogenase (complex I: ND1, ND2, ND3, ND4, ND4L, ND5 and ND6), Cytochrome b (subunit of complex III), 3 subunits of Cytochrome c oxidase or complex IV (COI, COII and COIII), and 2 subunits of F0F1 ATPase (ATPase 6 and ATPase 8). They are all encoded by mtDNA and synthesized in the organelle. While, complex II (Succinate Dehydrogenase) and the remaining subunits of complexes I, III, IV, and V are entirely encoded by the nuclear genome. These nuclear-encoded proteins are synthesized on cytosolic ribosomes and subsequently transported into the mitochondria.
4. Fe-S clusters (ISCs)
ISCs are evolutionarily ancient cofactors consisting of Fe (iron) and S (sulfur) associated to the cysteine sulfurs of proteins. The clusters are found in variety of organisms including archaea, protists, prokaryotes, and eukaryotes. In a eukaryotic cell, they can be found in the mitochondria, cytosol, and nucleus where they perform diverse functions . ISCs play a critical role in many fundamental molecular processes and have roles in electron transfer, structural stabilization, gene regulation, enzymatic catalysis, metabolic regulation, and sensing environmental signals . Almost 30 proteins in the mitochondria and the cytosol are involved in synthesizing and assembling these clusters. ISC have two most common forms [2Fe-2S] and [4Fe-4S] clusters. ISC-related proteins of the electron transport chain in the mitochondrion are mainly located in the inner membrane. Moreover, some of these proteins are also found in the mitochondrial matrix in the organelles. For the cluster assembly, two machineries are required, the mitochondrial ISC assembly machinery and the cytosolic IS protein assembly machinery .
Eukaryotic IS proteins are located in mitochondria, cytosol, and nucleus, where they perform diverse functions in cellular metabolism and regulation. The mitochondrial ISC assembly machinery matures all organellar IS proteins, and additionally contributes to the biogenesis of cytosolic and nuclear IS proteins by producing an unknown sulfur-containing compound (X-S) that is exported to the cytosol and used by the cytosolic IS protein assembly machinery. Hence, mitochondria are directly responsible for the essential functions (e.g., of nuclear IS proteins involved in DNA metabolism and genome maintenance).
Mitochondria forms iron-sulfur clusters of significant proteins such as DNA polymerase and DNA helicases, and, therefore, plays a significant role in survival. There are 17 different proteins forming iron-sulfur cluster machinery that places the clusters into the Apo proteins. The mechanism of formation of iron-sulfur clusters can be divided into three steps. First, it is synthesized on a scaffold protein. Second, it is bound to transfer protein after dislocation from scaffold protein. Third, the transfer protein, the cluster and the specific ISC targeting factor place the cluster into the Apo protein. The changes in the first two steps inhibit the maturation of extra mitochondrial Fe/S proteins and disturb the iron homeostasis . Assembly of Fe-S cluster also takes place by NIF, SUF, and CIA machineries. Cysteine desulfurase is an enzyme that unites Fe-S assembly machineries. It is encoded by NFS 1 which functions to deliver sulfur to ISCU . ISCU is an iron-sulfur cluster assembly enzyme; encodes component of iron-sulfur scaffold protein. The changes in this gene result in severe myopathy and lactic acidosis (“ISCU Fe-S Cluster Assembly Enzyme [
Yeast frataxin, Isu1, and Nfs1 (cysteine desulfurase) take part in
5. Genetic factors of mitochondrial dysfunction syndromes
As the names imply, multiple mitochondrial dysfunction syndromes are disease conditions affecting mitochondria and usually lead to reduced function of more than one stages of energy production in the organelle . The genetic factors causing these disorders are associated with the biogenesis of cellular ISC and currently these are the following genes:
|Gene||Cytoband||NCBI||Genomic location||MMDS-related phenotype||MIM PT#||IM||MIM LN|
|Multiple mitochondrial dysfunctions syndrome 1||AR|
|Multiple mitochondrial dysfunctions syndrome 2 with hyperglycinemia||AR|
|Multiple mitochondrial dysfunctions syndrome 3||AR|
|Multiple mitochondrial dysfunctions syndrome 4||AR|
|Multiple mitochondrial dysfunctions syndrome 5||AR|
Iron-sulfur cluster assembly 1 (ISCA1) is one of the mitochondrial proteins required for the biogenesis and assembly of ISC . This protein functions in the late stages of the ISC biogenesis and act as an iron binding molecule that may serve as a chaperone for biogenesis of Fe-S clusters . It is believed that the molecule plays its pivotal role through its interaction with IOP1 (iron-only hydrogenase-like protein)/NARFL (nuclear prelamin A recognition factor-like). Knockdown of
According to GenAtlas [23, 24], the gene has four exons and produces 14 kDa protein with 129 amino acids, which is known as mitochondrial Fe-S cluster assembly 1 homolog or otherwise HESB like domain containing 2. The gene is mapped to chromosome 9q21.33, and sits on genomic coordinates: 88.879.463–88.897.490. It is 2012 base pair long, generates four transcripts (splice variants) and highly expressed heart, esophagus, bladder, uterus, and cervix. Moreover, ISCA1 is a member of consensus coding sequence (CCDS:35056.1) which are manually checked protein annotations on the reference mouse and human genomes that ensures consistent representation of the tracks of NCBI, Ensembl, and UCSC Genome Browsers. The gene has several synonyms such as hIsca, HBLD2, and ISA1, and localizes to mitochondria as well as cytoplasm.
Effect of depletion of ISC-related proteins on the maturation of cytosolic 4Fe-4S proteins showed that some mitochondrial Fe/S proteins such as mitochondrial aconitase, SDH, several proteins of complex I, and Rieske Fe/S protein were decreased with the deficiency of ISCA1. On the other hand, cellular heme content and mitochondrial 2Fe-2S ferrochelatase were unaffected by the depletion. This implies that ISCA1 is crucial in the maturation of mitochondrial 4Fe-4S proteins . In another study,
ISCA2 stands for iron-sulfur cluster assembly 2 protein and the gene encodes for A-type iron-sulfur cluster protein. Fe-S clusters are inorganic cofactors, mostly found in metalloproteins. The gene is located on chromosome 14 and expressed from the plus strand. According to Ensembl, this gene generates 4 different transcripts and has 96 orthologues. ISCA2 is a regulatory protein found in mitochondria as well as extra mitochondrial sites such as cytosol and nucleus. The protein takes part in assembly of Fe-S clusters in mitochondria which further take part in oxidation reduction (especially in complex 1 and 2), substrate activation, iron/sulfur storage, regulation of gene expression, and enzyme activity. Alternative name for ISCA2 is “HESB-like-domain-containing protein 1” for humans. First human mutation of
NFU1 is one of the human mitochondrial components that is involved in the assembly of the Fe-S protein cluster. It helps in the transfer of [4Fe-4S] clusters to specific protein targets and facilitates their maturation .
IBA57 is a member of the Fe-S cluster assembly group. It is known as putative transferase CAF17 and Fe-S cluster assembly factor homolog.
BOLA3 is another essential protein in the Fe-S clusters production and involves in the normal maturation of lipoate-containing 2-oxoacid dehydrogenases. Another critical role of the molecule is to facilitate the assembly of the respiratory chain complexes.
Table 2 consists of previously published mutations in some Fe-S cluster genes.
|Gene||Mutation type||Mutation||Disease and phenotype||References|
|Missense||c.200T>A; p.Ile67Asn||Multiple mitochondrial dysfunctions syndrome||Haack et al. |
|Missense||c.287A>G p.His96Arg||Lethal infantile mitochondrial disorder||Kohda et al. |
|Nonsense||c.136C>T; p.Arg46*||Nonketotic hyperglycinemia,||Baker et al. |
|Microdeletion||c.225_229delGAGAA; p. Lys75*||Lethal infantile mitochondrial disorder||Kohda et al. |
|Microduplication||c.123dupA||Combined respiratory chain and 2-oxoacid dehydrogenase deficiency||Cameron et al. |
|Missense||c.313C>T; p.Arg105Trp||Leukodystrophy with acute psychomotor regression||Torraco et al. |
|Missense||c.316A>G; p.Thr106Ala||Leukodystrophy with acute psychomotor regression||Torraco et al. |
|Missense||c.436C>T; p.Arg146Thr||Leukodystrophy, fatal infantile||Debray et al. |
|Missense||c.586T>G; p.Trp196Gly||Leukodystrophy, developmental delay, feeding problems and recurrent vomiting||Torraco et al. |
|Missense||c.686C>T; p.Pro229Leu||Leukodystrophy, developmental delay, feeding problems and recurrent vomiting||Torraco et al. |
|Missense||c.706C>T; p.Pro236Ser||Leukodystrophy with acute psychomotor regression and feeding difficulties||Torraco et al. |
|Missense||c.757G>C; p.Val253Leu||Leukodystrophy with acute psychomotor regression||Torraco et al. |
|Missense||c.941A>C; p.Gln314Pro||Myopathy and encephalopathy||Ajit Bolar et al. |
|Splice||IVS2 ds A-G-2; c.678A>G||Spastic paraplegia||Lossos et al. |
|Small insertion||c.87_88ins11||Leukodystrophy with acute psychomotor regression||Torraco et al. |
|Missense||c.62G>C; p.Arg21Pro||NFU1 deficiency||Ahting et al. |
|Missense||c.544C>T; p.Arg182Trp||NFU1 deficiency||Ahting et al. |
|Missense||c.565G>A; p.Gly189Arg||Leukoencephalopathy with cysts and hyperglycinaemia||Nizon et al. [43, 44, 45]|
|Missense||c.568G>A; p.Gly190Arg||NFU1 deficiency||Ahting et al. |
|Missense||c.572A>T; p.Asp191Val||Multiple mitochondrial dysfunctions syndrome||Bai et al. |
|Missense||c.622G>T; p.Gly208Cys||Fatal infantile encephalopathy and/or pulmonary hypertension||Navarro-Sastre et al. |
|Missense||c.629G>T; p.Cys210Phe||Leukoencephalopathy||Invernizzi et al. |
|Splice||c.302+3A>G; p.Val56Glyfs*||NFU1 deficiency||Ahting et al. |
|Splice||c.545G>A; Skipping exon 6||Deficiency of the 2-oxoacid dehydrogenases accompanied by respiratory chain defects||Cameron et al. |
|Splice||c.545+5G>A; Skipping exon 6||Fatal infantile encephalopathy, pulmonary hypertension||Navarro-Sastre et al. |
|Microdeletion||c.90delC; p. Tyr30*||Multiple mitochondrial dysfunctions syndrome||Bai et al. |
|Microdeletion||c.146delC; p.Pro49LeufsX8||Spastic paraplegia||Tonduti et al. |
|Large deletion||55.6 kb region covering exons: 4–8||NFU1 deficiency||Ahting et al. |
|Missense||c.259G>A; p. Glu87Lys||Multiple mitochondrial dysfunctions syndrome||Shukla et al. |
|Missense||c.229G>A; p.Glu77Ser||Multiple mitochondrial dysfunctions syndrome||Al-Hassnan et al.  and others [27, 28]|
|Deletion and Missense||c.295delT and c.334A>G; p.Phe99Leufs*18 and p.Ser112Gly||Multiple mitochondrial dysfunctions syndrome||Toldo et al. |
Iron-sulfur clusters are indispensable inorganic cofactors for biological function and involve in numerous cellular processes such as respiration and DNA repair. The cluster’s assembly is complex and requires sophisticated protein machinery for its maturation and insertion into apoproteins. Since mitochondria is the main site for ISC biogenesis in human, any defect disturbing the biogenesis leads to a pathological outcome mostly appears as an mitochondrial entity in human. Currently, genetic alterations in several genes involving in ISC assembly and maturation have been linked to autosomal recessive mitochondrial human diseases known as multiple mitochondrial dysfunction syndromes. It is expected that more genes and alterations will appear in the literature related to ISC pathways. Moreover, there is still need to fully elucidate the phenotypic consequences of these genetic alterations and alteration of ISC pathways during the ISC related pathogenesis in human.
This study was supported by King Abdulaziz City for Science and Technology grant 11-BIO2221-20 (NK) and King Salman Center for Disability Research grant: 2180 004 (NK).