Mitochondrial Small Ribosomal Subunit Research Articles

The phenotypic spectrum of PTCD3 deficiency

AbstractThe PTCD3 gene product (protein PTCD3 or MRPS39) forms the entry channel of the mitochondrial small ribosomal subunit and binds to single‐stranded mRNA. Here, we expand on the clinical manifestations of PTCD3 pathogenic variants by describing an early‐onset patient with Leigh‐like syndrome and two patients with milder form of disease, with combined oxidative phosphorylation deficiency. A 34‐year‐old male and his 33‐year‐old sister both have horizontal nystagmus, pronounced rough tremor, truncal ataxia, dysmetria, spasticity and hyperreflexia. The basal respiration rate decreased significantly for the male patient and his mother (p < 0.0001) compared to the controls. The whole genome sequencing analysis revealed two heterozygous variants in the PTCD3: c.1182T>A, p.(Tyr394Ter) and c.805C>T, p.(His269Tyr). Tyr394Ter variant ablates the C‐terminal half of the protein, including a significant portion of the central fold. In silico modelling for the variant His269Tyr shows that the inclusion of the slightly larger tyrosine sidechain is well tolerated, with no significant change in either the position or the movement of the surrounding area. The third case is a 9‐year‐old boy, who has a global developmental delay, central hypotonia, hyperreflexia and abnormal MRI. PTCD3 pathogenic variant c.538+4A>G was identified by whole exome sequencing. To test the variant's effect on splicing, an RT‐PCR experiment was performed, which revealed skipping of an out‐of‐frame exon 7.

Schizosaccharomyces pombe Sls1 is primarily required for cox1 mRNA translation.

Mitochondrial DNA (mtDNA) encodes essential subunits of the oxidative phosphorylation (OXPHOS) complexes;thus, the expression of mtDNA-encoded genes is essential for the synthesis of adenosine triphosphate. However, factors involved in mitochondrial translation have not been fully characterized. In this study, we characterized Schizosaccharomyces pombe Sls1, which has sequence similarity to Saccharomyces cerevisiae Sls1 that is required for the translation of all mtDNA-encodedmessenger RNAs (mRNAs). Deletion of S. pombe sls1 severely impaired the growth of the cells on a rich medium containing the nonfermentable carbon source glycerol, which requires mitochondrial respiration. We found that the translation of mtDNA-encoded Cox1, the largest subunit of the cytochrome c oxidase complex, was severely impaired in Δsls1 cells. Deletion of S. pombe sls1 also resulted in a barely detectable steady-state level of mature cox1 mRNA. RNA immunoprecipitation showed that S. pombe Sls1 interacts with cox1 mRNA. Sucrose gradient sedimentation analysis revealed that S. pombe Sls1 is associated with the small subunit of mitochondrial ribosomes. Our results suggest that unlike S. cerevisiae Sls1, S. pombe Sls1 is primarily required for the accumulation and translation of cox1 mRNA.

Mitochondrial ribosomal small subunit (MRPS) MRPS23 protein-protein interaction reveals phosphorylation by CDK11-p58 affecting cell proliferation and knockdown of MRPS23 sensitizes breast cancer cells to CDK1 inhibitors.

Post-translational modification of some mitoribosomal proteins has been found to regulate their functions. MRPS23 has been reported to be overexpressed in various cancers and has been predicted to be involved in increased cell proliferation. Furthermore, MRPS23 is a driver of luminal subtype breast cancer. However, its exact role and function in cancer remains unknown. METHODS ANDRESULTS: Our previous study identified protein-protein interactions involving MRPS23 and CDK11A. In this study, we confirmed the interaction of MRPS23 with the p110 and p58 isoforms of CDK11A. Phosphoprotein enrichment studies and in vitro kinase assay using CDK11A/cyclin D3 followed by MALDI-ToF/ToF analysis confirmed the phosphorylation of MRPS23 at N-terminal serine 11 residue. Breast cancer cells expressing the MRPS23 (S11G) mutant showed increased cell proliferation, increased expression of PI3-AKT pathway proteins [p-AKT (Ser47), p-AKT (Thr308), p-PDK (Ser241) and p-GSK-3β (Ser9)] and increased antiapoptotic pathway protein expression [Bcl-2, Bcl-xL, p-Bcl2 (Ser70) and MCL-1] when compared with the MRPS23 (S11A) mutant-overexpressing cells. This finding indicated the role of MRPS23 phosphorylation in the proliferation and survival of breast cancer cells. The correlation of inconsistent MRPS23 phosphoserine 11 protein expression with CDK11A in the breast cancer cells suggested phosphorylation by other kinases. In vitro kinase assay showed that CDK1 kinase also phosphorylated MRPS23 and that inhibition using CDK1 inhibitors lowered phospho-MRPS23 (Ser11) levels. Additionally, modulating the expression of MRPS23 altered the sensitivity of the cells to CDK1 inhibitors. In conclusion, phosphorylation of MRPS23 by mitotic kinases might potentially be involved in the proliferation of breast cancer cells. Furthermore, MRPS23 can be targeted for sensitizing the breast cancer cells to CDK1 inhibitors.

The mitochondrial genome sequence analysis of Ophidascaris baylisi from the Burmese python (Python molurus bivittatus).

Ophidascaris species are parasitic roundworms that inhabit the python gut, resulting in severe granulomatous lesions or even death. However, the classification and nomenclature of these roundworms are still controversial. Our study aims to identify a snake roundworm from the Burmese python (Python molurus bivittatus) and analyze the mitochondrial genome. We identified this roundworm as Ophidascaris baylisi based on the morphology and cytochrome c oxidase subunit I (cox1) sequence. Ophidascaris baylisi complete mitochondrial genome was 14,784bp in length, consisting of two non-coding regions and 36 mitochondrial genes (12 protein-coding genes, 22 tRNA genes, and two rRNA genes). The protein-coding genes used TTG, ATG, ATT, or TTA as start codons and TAG, TAA, or T as stop codons. All tRNA genes showed a TV-loop structure, except trnS1AGN and trnS2UCN revealed a D-loop structure. The mitochondrial large ribosomal subunit 16S (rrnL) and small ribosomal subunit 12S (rrnS) were 956bp and 700bp long, respectively. Phylogenetic analysis based on O. baylisi mitochondrial protein-coding genes demonstrated that O. baylisi clustered with the family Ascarididae members and was most closely related to Ophidascaris wangi. These results may enhance the nematode mitochondrial genome database and provide valuable molecular markers for further research on the taxonomy, phylogeny, and genetic relationships of Ophidascaris nematodes.

Expression and analysis of the SAM-dependent RNA methyltransferase Rsm22 from Saccharomyces cerevisiae.

The Saccharomyces cerevisiae Rsm22 protein (Sc-Rsm22), encoded by the nuclear RSM22 (systematic name YKL155c) gene, is a distant homologue of Rsm22 from Trypanosoma brucei (Tb-Rsm22) and METTL17 from mouse (Mm-METTL17). All three proteins have been shown to be associated with mitochondrial gene expression, and Sc-Rsm22 has been documented to be essential for mitochondrial respiration. The Sc-Rsm22 protein comprises a polypeptide of molecular weight 72.2 kDa that is predicted to harbor an N-terminal mitochondrial targeting sequence. The precise physiological function of Rsm22-family proteins is unknown, and no structural information has been available for Sc-Rsm22 to date. In this study, Sc-Rsm22 was expressed and purified in monomeric and dimeric forms, their folding was confirmed by circular-dichroism analyses and their low-resolution structures were determined using a small-angle X-ray scattering (SAXS) approach. The solution structure of the monomeric form of Sc-Rsm22 revealed an elongated three-domain arrangement, which differs from the shape of Tb-Rsm22 in its complex with the mitochondrial small ribosomal subunit in T. brucei (PDB entry 6sg9). A bioinformatic analysis revealed that the core domain in the middle (Leu117-Asp462 in Sc-Rsm22) resembles the corresponding region in Tb-Rsm22, including a Rossmann-like methyltransferase fold followed by a zinc-finger-like structure. The latter structure is not present in this position in other methyltransferases and is therefore a unique structural motif for this family. The first half of the C-terminal domain is likely to form an OB-fold, which is typically found in RNA-binding proteins and is also seen in the Tb-Rsm22 structure. In contrast, the N-terminal domain of Sc-Rsm22 is predicted to be fully α-helical and shares no sequence similarity with other family members. Functional studies demonstrated that the monomeric variant of Sc-Rsm22 methylates mitochondrial tRNAs in vitro. These data suggest that Sc-Rsm22 is a new and unique member of the RNA methyltransferases that is important for mitochondrial protein synthesis.

Mitochondrial ribosomal small subunit proteins (MRPS) MRPS6 and MRPS23 show dysregulation in breast cancer affecting tumorigenic cellular processes.

Human Mitoribosomal Small Subunit unit (MRPS) family of genes appears to have role in cancer. Gene expression analysis of select MRPS genes (n=9) in 15 cancer cell lines showed altered expression in cancer cells. Protein levels of MRPS6, MRPS23 showed significant overexpression in breast cancer cells and tissues. Interestingly, their overexpression did not correlate with mitochondrial ribosome translated COX2 protein levels in breast cancer. Subcellular fractionation analysis showed a distinct presence of MRPS23 in the nuclear fraction. GST/MRP6 and GST/MRPS23 pulldown assays identified 32 novel protein-protein interactions (PPIs) and MRPS23-RIPK3 interaction was validated. Co-expression module identification tool (CEMi) analysis of breast cancer gene expression and MRPS6 and MRPS23 interactions revealed hub interactions in gene expression modules having functional roles in cancer-associated cellular processes. Based on PPI network analysis a novel interaction MRPS23-p53 was validated. Knockdown of MRPS6 and MRPS23 decreased proliferation, expression of select mesenchymal markers, oncogenes, and increased expression of tumor suppressor genes. Taken together present study has revealed that MRPS6 and MRPS23 genes have pro-tumorigenic functions in breast cancer.

Expression and affinity purification of recombinant mammalian mitochondrial ribosomal small subunit (MRPS) proteins and protein-protein interaction analysis indicate putative role in tumourigenic cellular processes.

Mitochondrial ribosomal small subunit (MRPS) group of proteins is structural constituents of the small subunit of mitoribosomes involved in translation. Recent studies indicate role in tumourigenic process, however, unlike cytosolic ribosomal proteins, knowledge on the role of MRPS proteins in alternate cellular processes is very limited. Mapping protein-protein interactions (PPIs) onto known cellular processes can be a valuable tool to identify novel protein functions. In this study, to identify PPIs of MRPS proteins, we have constructed 31 glutathione-S-transferase (GST)/MRPS fusion clones. GST/MRPS fusion proteins were confirmed by MALDI-TOF analysis. GST pull-downs were performed using eight GST/MRPS proteins (MRPS9, MRPS10, MRPS11, MRPS18B, MRPS31, MRPS33, MRPS38 and MRPS39), GST alone as pull-down control and HEK293 cell lysate as the source for anchor proteins followed by nLC/MS/MS analysis and probable PPIs of eight MRPS proteins were identified. Three PPIs from GST pull-downs and interaction between six MRPS proteins and p53 previously reported in PPI database were validated. The PPI network analysis revealed putative role in cellular processes with implications for tumourigenesis. Gene expression screening of a cancer cell line panel indicated overexpression of MRPS10 and MRPS31 in breast cancer. Co-expression module identification tool analysis of breast cancer gene expression and MRPS10 and MRPS31 PPIs revealed putative role for PPI with acyl-CoA dehydrogenase in fatty acid oxidation process regulated by brain-derived neurotrophic factor signalling pathway.

First Report of Vascular Wilt Caused by Fusarium proliferatum on Cauliflower in China

HomePlant DiseaseVol. 104, No. 11First Report of Vascular Wilt Caused by Fusarium proliferatum on Cauliflower in China PreviousNext DISEASE NOTES OPENOpen Access licenseFirst Report of Vascular Wilt Caused by Fusarium proliferatum on Cauliflower in ChinaLi YanLi Yan†Corresponding author: L. Yan; E-mail Address: liyan20200510@163.comHorticulture Sub-academy, Heilongjiang Academy of Agricultural Science, Harbin, Heilongjiang 150069, ChinaSearch for more papers by this authorAffiliationsAuthors and Affiliations Li Yan † Horticulture Sub-academy, Heilongjiang Academy of Agricultural Science, Harbin, Heilongjiang 150069, China Published Online:17 Sep 2020https://doi.org/10.1094/PDIS-05-20-0995-PDNAboutSectionsPDF ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinked InRedditEmailWechat Cauliflower (Brassica oleracea var. botrytis L.), which is belongs to the Brassicaceae, is an economically important vegetable crop in China and worldwide. Statistically, China is the largest cauliflower producer in the world (FAOSTAT 2018). During September 2017 to June 2018, severe outbreaks of root rot were observed on cauliflower in several commercial fields (approximately 3.5 ha), with disease incidence of approximately 25 to 50% in the Acheng District (127°06′E, 45°53′N) of Harbin City, Heilongjiang Province, China. Symptoms included leaf yellowing and wilting, plant stunting, root rot, and vascular discoloration of the stems and roots. A total of 15 symptomatic plants were uprooted and collected from seven fields. Infected tissues were surface sterilized by soaking in 1% sodium hypochlorite for 1 min, followed by three rinses in sterile distilled water, and then placed on potato dextrose agar (PDA) and incubated at 25°C in the dark for 5 days. In total, 21 cultures were obtained and purified by single-spore subcultures on PDA and carnation leaf agar (CLA; Leslie and Summerell 2006) for morphological identification. The colonies on PDA were originally white, later becoming light to dark purple in color with abundant aerial hyphae. Macroconidia were abundant on CLA, hyaline, slightly curved, three- to five-septate, with a distinct basal foot cell, measuring 20.6 to 46.8 × 2.4 to 4.6 μm. Microconidia were in short chains on monophialides and polyphialides, oval or ellipsoid, with a flat base, zero- to one-septate, measuring 4.7 to 15.1 × 2.1 to 4.4 µm. No chlamydospores were observed. The morphological features were consistent with Fusarium proliferatum (Matsushima) Nirenberg (Leslie and Summerell 2006). Pathogenicity tests were performed on the four-true-leaf stage of healthy cauliflower ‘Jingxue no. 88’ seedlings by dipping the roots into a conidial suspension (1 × 106 conidia/ml) for 10 min, which were produced on CLA. Control plants were dipped in sterile distilled water. Seven days later, inoculated plants developed typical leaf yellowing, vascular discoloration, and wilting symptoms identical to those observed in the fields, whereas the control plants remained healthy. F. proliferatum was reisolated from symptomatic stems, confirming Koch’s postulates. Genomic DNA was extracted from fungal mycelium using the Plant Genomic DNA Kit (Tiangen, China). The nuclear ribosomal internal transcribed spacer region (ITS), mitochondrial ribosomal small subunit (mtSSU), and translation elongation factor 1-alpha (TEF-1α) regions were amplified using primers ITS1/ITS4, NSM1/NSM2 (Li et al. 1994), and EF1/EF2 (Geiser et al. 2004), respectively. The resulting 537-bp ITS, 711-bp mtSSU, and 699-bp TEF-1α sequences of isolate HYC1410080401 were deposited in GenBank (MT378328, MT378332, and MT379885, respectively). BLAST analysis showed 100% identities to the ITS and TEF-1α sequences of F. proliferatum (MN809262.1 and MF445092.1) and 99.86% similarity with the mtSSU sequence of F. proliferatum (DQ831947.1). To our knowledge, this is the first report of F. proliferatum causing cauliflower vascular wilt worldwide (Farr and Rossman 2020). The pathogen has a worldwide distribution and an unusually broad host range, which can be of great significance, and control strategies should be followed.The author(s) declare no conflict of interest.

First Report on Plum Pocket Caused by Taphrina deformans in South Korea

Plum pocket caused by the dimorphic ascomycetous fungi, Taphrina spp., results in unsightly malformations and crop loss. In 2016, Japanese plums (Prunus salicina Lindl.) with plum pocket symptoms were found in Gimcheon. Three isolates were collected from symptomatic P. salicina fruits and identified as Taphrina deformans based on morphological characteristics and molecular sequence analysis of including internal transcribed space (ITS) and the mitochondrial small ribosomal subunit (SSU) regions of the three isolates. Pathogenicity test on plum fruits confirmed that, the present T. deformans isolates are causal agent of plum pocket. To the best of our knowledge, this is the first report of plum pocket caused by T. deformans in South Korea.

RCC1L (WBSCR16) isoforms coordinate mitochondrial ribosome assembly through their interaction with GTPases.

Mitochondrial translation defects can be due to mutations affecting mitochondrial- or nuclear-encoded components. The number of known nuclear genes involved in mitochondrial translation has significantly increased in the past years. RCC1L (WBSCR16), a putative GDP/GTP exchange factor, has recently been described to interact with the mitochondrial large ribosomal subunit. In humans, three different RCC1L isoforms have been identified that originate from alternative splicing but share the same N-terminus, RCC1LV1, RCC1LV2 and RCC1LV3. All three isoforms were exclusively localized to mitochondria, interacted with its inner membrane and could associate with homopolymeric oligos to different extent. Mitochondrial immunoprecipitation experiments showed that RCC1LV1 and RCC1LV3 associated with the mitochondrial large and small ribosomal subunit, respectively, while no significant association was observed for RCC1LV2. Overexpression and silencing of RCC1LV1 or RCC1LV3 led to mitoribosome biogenesis defects that resulted in decreased translation. Indeed, significant changes in steady-state levels and distribution on isokinetic sucrose gradients were detected not only for mitoribosome proteins but also for GTPases, (GTPBP10, ERAL1 and C4orf14), and pseudouridylation proteins, (TRUB2, RPUSD3 and RPUSD4). All in all, our data suggest that RCC1L is essential for mitochondrial function and that the coordination of at least two isoforms is essential for proper ribosomal assembly.

METTL15 interacts with the assembly intermediate of murine mitochondrial small ribosomal subunit to form m4C840 12S rRNA residue.

Mammalian mitochondrial ribosomes contain a set of modified nucleotides, which is distinct from that of the cytosolic ribosomes. Nucleotide m4C840 of the murine mitochondrial 12S rRNA is equivalent to the dimethylated m4Cm1402 residue of Escherichia coli 16S rRNA. Here we demonstrate that mouse METTL15 protein is responsible for the formation of m4C residue of the 12S rRNA. Inactivation of Mettl15 gene in murine cell line perturbs the composition of mitochondrial protein biosynthesis machinery. Identification of METTL15 interaction partners revealed that the likely substrate for this RNA methyltransferase is an assembly intermediate of the mitochondrial small ribosomal subunit containing an assembly factor RBFA.

METTL15 introduces N4-methylcytidine into human mitochondrial 12S rRNA and is required for mitoribosome biogenesis.

Post-transcriptional RNA modifications, the epitranscriptome, play important roles in modulating the functions of RNA species. Modifications of rRNA are key for ribosome production and function. Identification and characterization of enzymes involved in epitranscriptome shaping is instrumental for the elucidation of the functional roles of specific RNA modifications. Ten modified sites have been thus far identified in the mammalian mitochondrial rRNA. Enzymes responsible for two of these modifications have not been characterized. Here, we identify METTL15, show that it is the main N4-methylcytidine (m4C) methyltransferase in human cells and demonstrate that it is responsible for the methylation of position C839 in mitochondrial 12S rRNA. We show that the lack of METTL15 results in a reduction of the mitochondrial de novo protein synthesis and decreased steady-state levels of protein components of the oxidative phosphorylation system. Without functional METTL15, the assembly of the mitochondrial ribosome is decreased, with the late assembly components being unable to be incorporated efficiently into the small subunit. We speculate that m4C839 is involved in the stabilization of 12S rRNA folding, therefore facilitating the assembly of the mitochondrial small ribosomal subunits. Taken together our data show that METTL15 is a novel protein necessary for efficient translation in human mitochondria.

Aim23p Interacts with the Yeast Mitochondrial Ribosomal Small Subunit.

Protein synthesis in mitochondria is generally organized in a bacterial-like manner but, at the same time, possesses several unique traits. Translation initiation in mitochondria is regulated by two protein factors, mtIF2 and mtIF3. Previously we demonstrated that Saccharomyces cerevisiae Aim23 protein is an ortholog of IF3 in budding yeast. However, the data on the interactions between Aim23p and other proteins are limited. Here, we demonstrated that Aim23p interacts with the yeast mitochondrial ribosomal small subunit both in vivo and in vitro using co-immunoprecipitation and density gradient sedimentation.

Insight into higher-level phylogeny of Neuropterida: Evidence from secondary structures of mitochondrial rRNA genes and mitogenomic data.

It is well known that the rRNA structure information is important to assist phylogenetic analysis through identifying homologous positions to improve alignment accuracy. In addition, the secondary structure of some conserved motifs is highly stable among distantly related taxa, which can provide potentially informative characters for estimating phylogeny. In this paper, we applied the high-throughput pooled sequencing approach to the determination of neuropteran mitogenomes. Four complete mitogenome sequences were obtained: Micromus angulatus (Hemerobiidae), Chrysoperla nipponensis (Chrysopidae), Rapisma sp. (Ithonidae), and Thaumatosmylus sp. (Osmylidae). This allowed us to sample more complete mitochondrial RNA gene sequences. Secondary structure diagrams for the complete mitochondrial small and large ribosomal subunit RNA genes of eleven neuropterid species were predicted. Comparative analysis of the secondary structures indicated a closer relationship of Megaloptera and Neuroptera. This result was congruent with the resulting phylogeny inferred from sequence alignments of all 37 mitochondrial genes, namely the hypothesis of (Raphidioptera + (Megaloptera + Neuroptera)).

A homozygous missense mutation in ERAL1, encoding a mitochondrial rRNA chaperone, causes Perrault syndrome.

Perrault syndrome (PS) is a rare recessive disorder characterized by ovarian dysgenesis and sensorineural deafness. It is clinically and genetically heterogeneous, and previously mutations have been described in different genes, mostly related to mitochondrial proteostasis. We diagnosed three unrelated females with PS and set out to identify the underlying genetic cause using exome sequencing. We excluded mutations in the known PS genes, but identified a single homozygous mutation in the ERAL1 gene (c.707A > T; p.Asn236Ile). Since ERAL1 protein binds to the mitochondrial 12S rRNA and is involved in the assembly of the small mitochondrial ribosomal subunit, the identified variant represented a likely candidate. In silico analysis of a 3D model for ERAL1 suggested that the mutated residue hinders protein-substrate interactions, potentially affecting its function. On a molecular basis, PS skin fibroblasts had reduced ERAL1 protein levels. Complexome profiling of the cells showed an overall decrease in the levels of assembled small ribosomal subunit, indicating that the ERAL1 variant affects mitochondrial ribosome assembly. Moreover, levels of the 12S rRNA were reduced in the patients, and were rescued by lentiviral expression of wild type ERAL1. At the physiological level, mitochondrial respiration was markedly decreased in PS fibroblasts, confirming disturbed mitochondrial function. Finally, knockdown of the C. elegans ERAL1 homologue E02H1.2 almost completely blocked egg production in worms, mimicking the compromised fertility in PS-affected women. Our cross-species data in patient cells and worms support the hypothesis that mutations in ERAL1 can cause PS and are associated with changes in mitochondrial metabolism.

RSM22, mtYsxC and PNKD-like proteins are required for mitochondrial translation in Trypanosoma brucei

Mitochondrial ribosomes evolved from prokaryotic ribosomes, with which they therefore share more common features than with their counterparts in the cytosol. Yet, mitochondrial ribosomes are highly diverse in structure and composition, having undergone considerable changes, including reduction of their RNA component and varying degree of acquisition of novel proteins in various phylogenetic lineages. Here, we present functional analysis of three putative mitochondrial ribosome-associated proteins (RSM22, mtYsxC and PNKD-like) in Trypanosoma brucei, originally identified by database mining. While in other systems the homologs of RSM22 are known as components of mitochondrial ribosomes, YsxC was linked with ribosomes only in bacteria. The PNKD-like protein shows similarity to a human protein, the defects of which cause PNKD (paroxysmal non-kinesigenic dyskinesia). Here we show that all three proteins are important for the growth of T. brucei. They play an important function in mitochondrial translation, as their ablation by RNAi rapidly and severely affected the de novo synthesis of mitochondrial proteins. Moreover, following the RNAi-mediated depletion of RSM22, structure of the small subunit of mitochondrial ribosome becomes severely compromised, suggesting a role of RSM22 in ribosomal assembly and/or stability.

Mutation in mitochondrial ribosomal protein S7 (MRPS7) causes congenital sensorineural deafness, progressive hepatic and renal failure and lactic acidemia.

Functional defects of the mitochondrial translation machinery, as a result of mutations in nuclear-encoded genes, have been associated with combined oxidative phosphorylation (OXPHOS) deficiencies. We report siblings with congenital sensorineural deafness and lactic acidemia in association with combined respiratory chain (RC) deficiencies of complexes I, III and IV observed in fibroblasts and liver. One of the siblings had a more severe phenotype showing progressive hepatic and renal failure. Whole-exome sequencing revealed a homozygous mutation in the gene encoding mitochondrial ribosomal protein S7 (MRPS7), a c.550A>G transition that encodes a substitution of valine for a highly conserved methionine (p.Met184Val) in both affected siblings. MRPS7 is a 12S ribosomal RNA-binding subunit of the small mitochondrial ribosomal subunit, and is required for the assembly of the small ribosomal subunit. Pulse labeling of mitochondrial protein synthesis products revealed impaired mitochondrial protein synthesis in patient fibroblasts. Exogenous expression of wild-type MRPS7 in patient fibroblasts rescued complexes I and IV activities, demonstrating the deleterious effect of the mutation on RC function. Moreover, reduced 12S rRNA transcript levels observed in the patient's fibroblasts were also restored to normal levels by exogenous expression of wild-type MRPS7. Our data demonstrate the pathogenicity of the identified MRPS7 mutation as a novel cause of mitochondrial RC dysfunction, congenital sensorineural deafness and progressive hepatic and renal failure.

The reassignment of three ‘lost’ Taphrina species (Taphrina bullata, Taphrina insititiae and Taphrina rhizophora) supported by the divergence of nuclear and mitochondrial DNA

The ascomycetous genus Taphrina Fries originally contained more than 90 phytopathogenic microscopic dimorphic ascomycetes causing specific infections in different vascular plants. Although species have mainly been identified historically according to their host and morphological and physiological traits, they can be studied and preserved in the form of yeasts arising from germinating ascospores. Due to low DNA sequence divergence and the lack of available strains, the number of accepted Taphrina species has currently been reduced to 28. The aim of this study is the description of three previously accepted species. Taphrina bullata (type strain CCY 58-4-1 = CBS 12783), Taphrina insititiae (type strain CCY 58-5-1 = CBS 12782) and Taphrina rhizophora (type strain CCY 58-6-1 = CBS 12781), which have been omitted from a recent key. The host range, the divergence of the 26S rRNA gene sequence, internal transcribed spacers (ITS) and the mitochondrial small ribosomal subunit (rns) sequence strongly support their reassignment as species.

Identification of New Fungal Pathogens of Coast Live Oak in California.

A decline of coast live oak (Quercus agrifolia) has been observed throughout southern California. In this study, the identity and pathogenicity of non-Botryosphaeriaceae fungal species consistently recovered from necrotic tissues of branch and bleeding trunk canker samples from these locations were assessed. Species were identified morphologically and by comparison of the complete sequence of the internal transcribed spacer (ITS) of the ribosomal DNA to sequences available in GenBank. Phylogenetic analyses were then conducted using ITS and partial sequences of the β-tubulin and mitochondrial small ribosomal subunit genes for unknown species. Fungi recovered and identified included Fusarium solani, Phaeoacremonium mortoniae, Diatrypella verrucaeformis, and a fungus described herein as Cryptosporiopsis querciphila sp. nov. One-year-old coast live oak seedlings were wound inoculated under controlled conditions to test pathogenicity of the fungal species. Each fungal species was successfully reisolated from necrotic tissue at 70 and 100% for P. mortoniae and all other species, and xylem necrosis was significantly different between all treatments and controls (P < 0.0001 at α = 0.05). Isolates of F. solani were the most aggressive tested. These species represent new records of fungal pathogens of coast live oak in California. Results from the pathogenicity test suggest that these fungi play a role in the decline of southern California coast live oak trees.

Human C4orf14 interacts with the mitochondrial nucleoid and is involved in the biogenesis of the small mitochondrial ribosomal subunit

The bacterial homologue of C4orf14, YqeH, has been linked to assembly of the small ribosomal subunit. Here, recombinant C4orf14 isolated from human cells, co-purified with the small, 28S subunit of the mitochondrial ribosome and the endogenous protein co-fractionated with the 28S subunit in sucrose gradients. Gene silencing of C4orf14 specifically affected components of the small subunit, leading to decreased protein synthesis in the organelle. The GTPase of C4orf14 was critical to its interaction with the 28S subunit, as was GTP. Therefore, we propose that C4orf14, with bound GTP, binds to components of the 28S subunit facilitating its assembly, and GTP hydrolysis acts as the release mechanism. C4orf14 was also found to be associated with human mitochondrial nucleoids, and C4orf14 gene silencing caused mitochondrial DNA depletion. In vitro C4orf14 is capable of binding to DNA. The association of C4orf14 with mitochondrial translation factors and the mitochondrial nucleoid suggests that the 28S subunit is assembled at the mitochondrial nucleoid, enabling the direct transfer of messenger RNA from the nucleoid to the ribosome in the organelle.