Reference Gene Validation via RT-qPCR for Human iPSC-Derived Neural Stem Cells and Neural Progenitors.

Correct selection of the reference gene(s) is the most important step in gene expression analysis. The aims of this study were to identify and evaluate the panel of possible reference genes in neural stem cells (NSC), early neural progenitors (eNP) and neural progenitors (NP) obtained from human-induced pluripotent stem cells (hiPSC). The stability of expression of genes commonly used as the reference in cells during neural differentiation is variable and does not meet the criteria for reference genes.

Controlling the mitochondrial antisense - role of the SUV3-PNPase complex and its co-factor GRSF1 in mitochondrial RNA surveillance.

Transcription of the human mitochondrial genome produces a vast amount of non-coding antisense RNAs. These RNA species can form G-quadraplexes (G4), which affect their decay. We found that the mitochondrial degradosome, a complex of RNA helicase SUPV3L1 (best known as SUV3) and the ribonuclease PNPT1 (also known as PNPase), together with G4-melting protein GRSF1, is a key player in restricting antisense mtRNAs.

Bezafibrate Upregulates Mitochondrial Biogenesis and Influence Neural Differentiation of Human-Induced Pluripotent Stem Cells.

Bezafibrate (BZ) regulates mitochondrial biogenesis by activation of PPAR's receptors and enhancing the level of PGC-1α coactivator. In this report, we investigated the effect of BZ on the expression of genes (1) that are linked to different pathways involved in mitochondrial biogenesis, e.g.

Dedicated surveillance mechanism controls G-quadruplex forming non-coding RNAs in human mitochondria.

The GC skew in vertebrate mitochondrial genomes results in synthesis of RNAs that are prone to form G-quadruplexes (G4s). Such RNAs, although mostly non-coding, are transcribed at high rates and are degraded by an unknown mechanism. Here we describe a dedicated mechanism of degradation of G4-containing RNAs, which is based on cooperation between mitochondrial degradosome and quasi-RNA recognition motif (qRRM) protein GRSF1.

Human SUV3 helicase regulates growth rate of the HeLa cells and can localize in the nucleoli.

The human SUV3 helicase (SUV3, hSUV3, SUPV3L1) is a DNA/RNA unwinding enzyme belonging to the class of DexH-box helicases. It localizes predominantly in the mitochondria, where it forms an RNA-degrading complex called mitochondrial degradosome with exonuclease PNP (polynucleotide phosphorylase). Association of this complex with the polyA polymerase can modulate mitochondrial polyA tails.

[Dialogues between cell nuclei and mitochondria].

Mitochondria are not only ATP producing organelles, but they play pivotal roles in apoptosis, neurodegeneration, cancer and aging. Mammalian mitochondrial genome is a small DNA molecule of about 16.5 kb, encoding less than 20 polypeptides and a set of ribosomal RNAs and tRNAs.

Yeast and human mitochondrial helicases.

Mitochondria are semiautonomous organelles which contain their own genome. Both maintenance and expression of mitochondrial DNA require activity of RNA and DNA helicases. In Saccharomyces cerevisiae the nuclear genome encodes four DExH/D superfamily members (MSS116, SUV3, MRH4, IRC3) that act as helicases and/or RNA chaperones.

Human mitochondrial RNA decay mediated by PNPase-hSuv3 complex takes place in distinct foci.

RNA decay is usually mediated by protein complexes and can occur in specific foci such as P-bodies in the cytoplasm of eukaryotes. In human mitochondria nothing is known about the spatial organization of the RNA decay machinery, and the ribonuclease responsible for RNA degradation has not been identified. We demonstrate that silencing of human polynucleotide phosphorylase (PNPase) causes accumulation of RNA decay intermediates and increases the half-life of mitochondrial transcripts.

RNA degradation in yeast and human mitochondria.

Expression of mitochondrially encoded genes must be finely tuned according to the cell's requirements. Since yeast and human mitochondria have limited possibilities to regulate gene expression by altering the transcription initiation rate, posttranscriptional processes, including RNA degradation, are of great importance. In both organisms mitochondrial RNA degradation seems to be mostly depending on the RNA helicase Suv3.

Human Suv3 protein reveals unique features among SF2 helicases.

Suv3 is a helicase that is involved in efficient turnover and surveillance of RNA in eukaryotes. In vitro studies show that human Suv3 (hSuv3) in complex with human polynucleotide phosphorylase has RNA degradosome activity. The enzyme is mainly localized in mitochondria, but small fractions are found in cell nuclei.

The human Suv3 helicase interacts with replication protein A and flap endonuclease 1 in the nucleus.

The hSuv3 (human Suv3) helicase has been shown to be a major player in mitochondrial RNA surveillance and decay, but its physiological role might go beyond this functional niche. hSuv3 has been found to interact with BLM (Bloom's syndrome protein) and WRN (Werner's syndrome protein), members of the RecQ helicase family involved in multiple DNA metabolic processes, and in protection and stabilization of the genome. In the present study, we have addressed the possible role of hSuv3 in genome maintenance by examining its potential association with key interaction partners of the RecQ helicases.

Involvement of human ELAC2 gene product in 3' end processing of mitochondrial tRNAs.

Accurate tRNA processing is crucial for human mitochondrial genome expression, but the mechanisms of mt-tRNA cleavage and the key enzymes involved in this process are poorly characterized. At least two activities are required for proper mt-tRNA maturation: RNase P cleaving precursor molecules at the 5' end and tRNase Z at the 3' end. In human mitochondria only RNase P has been identified so far.

The human core exosome interacts with differentially localized processive RNases: hDIS3 and hDIS3L.

The eukaryotic RNA exosome is a ribonucleolytic complex involved in RNA processing and turnover. It consists of a nine-subunit catalytically inert core that serves a structural function and participates in substrate recognition. Best defined in Saccharomyces cerevisiae, enzymatic activity comes from the associated subunits Dis3p (Rrp44p) and Rrp6p.

Assays of the helicase, ATPase, and exoribonuclease activities of the yeast mitochondrial degradosome.

The mitochondrial degradosome (mtEXO) is the main enzymatic complex in RNA degradation, processing, and surveillance in Saccharomyces cerevisiae mitochondria. It consists of two nuclear-encoded subunits: the ATP-dependent RNA helicase Suv3p and the 3' to 5' exoribonuclease Dss1p. The two subunits depend on each other for their activity; the complex can therefore be considered as a model system for the cooperation of RNA helicases and exoribonucleases in RNA degradation.

RNA turnover in human mitochondria: more questions than answers?

Protein complexes responsible for RNA degradation play important role in three key aspects of RNA metabolism: they control stability of physiologically functional transcripts, remove the unnecessary RNA processing intermediates and destroy aberrantly formed RNAs. In mitochondria the post-transcriptional events seem to play a major role in regulation of gene expression, therefore RNA turnover is of particular importance. Despite many years of research, the details of this process are still a challenge.

Human mitochondrial RNA turnover caught in flagranti: involvement of hSuv3p helicase in RNA surveillance.

The mechanism of human mitochondrial RNA turnover and surveillance is still a matter of debate. We have obtained a cellular model for studying the role of hSuv3p helicase in human mitochondria. Expression of a dominant-negative mutant of the hSUV3 gene which encodes a protein with no ATPase or helicase activity results in perturbations of mtRNA metabolism and enables to study the processing and degradation intermediates which otherwise are difficult to detect because of their short half-lives.

In vivo and in vitro approaches for studying the yeast mitochondrial RNA degradosome complex.

The mitochondrial degradosome (mtEXO) of S. cerevisiae is the main exoribonuclease of yeast mitochondria. It is involved in many pathways of mitochondrial RNA metabolism, including RNA degradation, surveillance, and processing, and its activity is essential for mitochondrial gene function.

Interaction of human SUV3 RNA/DNA helicase with BLM helicase; loss of the SUV3 gene results in mouse embryonic lethality.

The SUV3 gene is present in all eukaryotes and encodes an RNA/DNA helicase which operates both in mitochondria and cell nuclei. To assess its function in mammals we generated a mouse mutant strain in which the 3' part of the SUV3 gene is disrupted. The mutated allele is a hypomorph transmitted from one generation to another at a frequency about 35% lower than expected while mice homozygous for the mutation die in utero before midgestation.

In vitro reconstitution and characterization of the yeast mitochondrial degradosome complex unravels tight functional interdependence.

The mitochondrial degradosome (mtEXO), the main RNA-degrading complex of yeast mitochondria, is composed of two subunits: an exoribonuclease encoded by the DSS1 gene and an RNA helicase encoded by the SUV3 gene. We expressed both subunits of the yeast mitochondrial degradosome in Escherichia coli, reconstituted the complex in vitro and analyzed the RNase, ATPase and helicase activities of the two subunits separately and in complex. The results reveal a very strong functional interdependence.

Prediction of the structure of the common perimitochondrial localization signal of nuclear transcripts in yeast.

Many nuclear genes encoding mitochondrial proteins require specific localization of their mRNAs to the vicinity of mitochondria for proper expression. Studies in Saccharomyces cerevisiae have shown that the cis-acting signal responsible for subcellular localization of mRNAs is localized in the 3' UTR of the transcript. In this paper we present an in silico approach for prediction of a common perimitochondrial localization signal of nuclear transcripts encoding mitochondrial proteins.

Down-regulation of human RNA/DNA helicase SUV3 induces apoptosis by a caspase- and AIF-dependent pathway.

Background Information: The nuclear gene hSUV3 (human SUV3) encodes an ATP-dependent DNA/RNA helicase. In the yeast Saccharomyces cerevisiae the orthologous Suv3 protein is localized in mitochondria, and is a subunit of the degradosome complex which regulates RNA surveillance and turnover. In contrast, the functions of human SUV3 are not known to date.

Up-regulation of human PNPase mRNA by beta-interferon has no effect on protein level in melanoma cell lines.

Human mitochondrial polynucleotide phosphorylase (hPNPase) is an exoribonuclease localized in mitochondria. The exact physiological function of this enzyme is unknown. Recent studies have revealed the existence of a relationship between induction of hPNPase mRNA and both cellular senescence and growth arrest of melanoma cells following beta-interferon treatment.

Differential stability of mitochondrial mRNA in HeLa cells.

The physiological significance and metabolism of oligoadenylated and polyadenylated human mitochondrial mRNAs are not known to date. After study of eight mitochondrial transcripts (ND1, ND2, ND3, ND5, CO1, CO2, ATP6/8 and Cyt. b) we found a direct correlation between the half-lives of mitochondrial mRNAs and their steady-state levels.

Balance between transcription and RNA degradation is vital for Saccharomyces cerevisiae mitochondria: reduced transcription rescues the phenotype of deficient RNA degradation.

The Saccharomyces cerevisiae SUV3 gene encodes the helicase component of the mitochondrial degradosome (mtEXO), the principal 3'-to-5' exoribonuclease of yeast mitochondria responsible for RNA turnover and surveillance. Inactivation of SUV3 (suv3Delta) causes multiple defects related to overaccumulation of aberrant transcripts and precursors, leading to a disruption of mitochondrial gene expression and loss of respiratory function. We isolated spontaneous suppressors that partially restore mitochondrial function in suv3Delta strains devoid of mitochondrial introns and found that they correspond to partial loss-of-function mutations in genes encoding the two subunits of the mitochondrial RNA polymerase (Rpo41p and Mtf1p) that severely reduce the transcription rate in mitochondria.

Human ATP-dependent RNA/DNA helicase hSuv3p interacts with the cofactor of survivin HBXIP.

The human SUV3gene encodes an NTP-dependent DNA/RNA DExH box helicase predominantly localized in mitochondria. Its orthologue in yeast is a component of the mitochondrial degradosome complex involved in the mtRNA decay pathway. In contrast to this, the physiological function of human SUV3 remains to be elucidated.