The mitochondrial inner membrane consists of two domains, inner boundary membrane and cristae membrane that are connected by crista junctions. Mitofilin/Fcj1 was reported to be involved in formation ...of crista junctions, however, different views exist on its function and possible partner proteins. We report that mitofilin plays a dual role. Mitofilin is part of a large inner membrane complex, and we identify five partner proteins as constituents of the mitochondrial inner membrane organizing system (MINOS) that is required for keeping cristae membranes connected to the inner boundary membrane. Additionally, mitofilin is coupled to the outer membrane and promotes protein import via the mitochondrial intermembrane space assembly pathway. Our findings indicate that mitofilin is a central component of MINOS and functions as a multifunctional regulator of mitochondrial architecture and protein biogenesis.
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► Mitofilin/Fcj1 is part of a large protein complex of the mitochondrial inner membrane ► This MINOS complex regulates cristae morphology ► In MINOS mutants cristae are released from the inner boundary membrane (IBM) ► Mitofilin interacts with the outer membrane and promotes protein import
The present prospective study aimed at determining the impact of cell-free tumor DNA (ct-DNA), CA125 and HE4 from blood and ascites for quantification of tumor burden in patients with advanced ...high-grade serous epithelial ovarian cancer (EOC).
Genomic DNA was extracted from tumor FFPE and ct-DNA from plasma before surgery and on subsequent post-surgical days. Extracted DNA was subjected to hybrid-capture based next generation sequencing. Blood and ascites were sampled before surgery and on subsequent post-surgical days. 20 patients (10 undergoing complete resection (TR0), 10 undergoing incomplete resection (TR>0)) were included.
The minor allele frequency (MAF) of TP53 mutations in ct-DNA of all patients with TR0 decreased significantly, compared to only one patient with TR>0. It was not possible to distinguish between patients with TR0 and patients with TR>0, using CA125 and HE4 from blood and ascites.
Based upon the present findings, ct-DNA assessment in patients with high-grade serous EOC might help to better determine disease burden compared to standard tumor markers. Further studies should prospectively evaluate whether this enhancement of accuracy can help to optimize management of patients with EOC.
Previous work indicated that lysine-specific demethylase 1 (Lsd1) can positively regulate the oxidative and thermogenic capacities of white and beige adipocytes. Here we investigate the role of Lsd1 ...in brown adipose tissue (BAT) and find that BAT-selective Lsd1 ablation induces a shift from oxidative to glycolytic metabolism. This shift is associated with downregulation of BAT-specific and upregulation of white adipose tissue (WAT)-selective gene expression. This results in the accumulation of di- and triacylglycerides and culminates in a profound whitening of BAT in aged Lsd1-deficient mice. Further studies show that Lsd1 maintains BAT properties via a dual role. It activates BAT-selective gene expression in concert with the transcription factor Nrf1 and represses WAT-selective genes through recruitment of the CoREST complex. In conclusion, our data uncover Lsd1 as a key regulator of gene expression and metabolic function in BAT.
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•Loss of Lsd1 in brown adipocytes induces a brown-to-white fat cell conversion•Lsd1 controls BAT- and WAT-selective genes via a dual mechanism•Lsd1 deletion in brown adipocytes shifts oxidative to glycolytic metabolism•Lsd1 controls thermogenesis in BAT to counteract obesity
Duteil et al. find that BAT-selective Lsd1 ablation downregulates BAT-selective and upregulates WAT-selective gene expression. Lsd1 deletion shifts oxidative to glycolytic metabolism, resulting in whitening of BAT. Lsd1 maintains BAT properties in concert with the transcription factor Nrf1 and CoREST.
Increased expression of the Microphthalmia-associated transcription factor (MITF) contributes to melanoma progression and resistance to BRAF pathway inhibition. Here we show that the lack of MITF is ...associated with more severe resistance to a range of inhibitors, while its presence is required for robust drug responses. Both in primary and acquired resistance, MITF levels inversely correlate with the expression of several activated receptor tyrosine kinases, most frequently AXL. The MITF-low/AXL-high/drug-resistance phenotype is common among mutant BRAF and NRAS melanoma cell lines. The dichotomous behaviour of MITF in drug response is corroborated in vemurafenib-resistant biopsies, including MITF-high and -low clones in a relapsed patient. Furthermore, drug cocktails containing AXL inhibitor enhance melanoma cell elimination by BRAF or ERK inhibition. Our results demonstrate that a low MITF/AXL ratio predicts early resistance to multiple targeted drugs, and warrant clinical validation of AXL inhibitors to combat resistance of BRAF and NRAS mutant MITF-low melanomas.
Posttranslational modifications of histones, such as methylation, regulate chromatin structure and gene expression. Recently, lysine-specific demethylase 1 (LSD1), the first histone demethylase, was ...identified. LSD1 interacts with the androgen receptor and promotes androgen-dependent transcription of target genes by ligand-induced demethylation of mono- and dimethylated histone H3 at Lys 9 (H3K9) only. Here, we identify the Jumonji C (JMJC) domain-containing protein JMJD2C as the first histone tridemethylase regulating androgen receptor function. JMJD2C interacts with androgen receptor in vitro and in vivo. Assembly of ligand-bound androgen receptor and JMJD2C on androgen receptor-target genes results in demethylation of trimethyl H3K9 and in stimulation of androgen receptor-dependent transcription. Conversely, knockdown of JMJD2C inhibits androgen-induced removal of trimethyl H3K9, transcriptional activation and tumour cell proliferation. Importantly, JMJD2C colocalizes with androgen receptor and LSD1 in normal prostate and in prostate carcinomas. JMJD2C and LSD1 interact and both demethylases cooperatively stimulate androgen receptor-dependent gene transcription. In addition, androgen receptor, JMJD2C and LSD1 assemble on chromatin to remove methyl groups from mono, di and trimethylated H3K9. Thus, our data suggest that specific gene regulation requires the assembly and coordinate action of demethylases with distinct substrate specificities.
Gene regulation in eukaryotes requires the coordinate interaction of chromatin-modulating proteins with specific transcription factors such as the androgen receptor. Gene activation and repression is ...specifically regulated by histone methylation status at distinct lysine residues. Here we show that lysine-specific demethylase 1 (LSD1; also known as BHC110) co-localizes with the androgen receptor in normal human prostate and prostate tumour. LSD1 interacts with androgen receptor in vitro and in vivo, and stimulates androgen-receptor-dependent transcription. Conversely, knockdown of LSD1 protein levels abrogates androgen-induced transcriptional activation and cell proliferation. Chromatin immunoprecipitation analyses demonstrate that androgen receptor and LSD1 form chromatin-associated complexes in a ligand-dependent manner. LSD1 relieves repressive histone marks by demethylation of histone H3 at lysine 9 (H3-K9), thereby leading to de-repression of androgen receptor target genes. Furthermore, we identify pargyline as an inhibitor of LSD1. Pargyline blocks demethylation of H3-K9 by LSD1 and consequently androgen-receptor-dependent transcription. Thus, modulation of LSD1 activity offers a new strategy to regulate androgen receptor functions. Here, we link demethylation of a repressive histone mark with androgen-receptor-dependent gene activation, thus providing a mechanism by which demethylases control specific gene expression.
The mitochondrial outer membrane contains translocase complexes for the import of precursor proteins. The translocase of the outer membrane complex functions as a general preprotein entry gate, ...whereas the sorting and assembly machinery complex mediates membrane insertion of β-barrel proteins of the outer membrane. Several α-helical outer membrane proteins are known to carry multiple transmembrane segments; however, only limited information is available on the biogenesis of these proteins. We report that mitochondria lacking the mitochondrial import protein 1 (Mim1) are impaired in the biogenesis of multispanning outer membrane proteins, whereas overexpression of Mim1 stimulates their import. The Mim1 complex cooperates with the receptor Tom70 in binding of precursor proteins and promotes their insertion and assembly into the outer membrane. We conclude that the Mim1 complex plays a central role in the import of α-helical outer membrane proteins with multiple transmembrane segments.
Kinetics of RNA-LNP delivery and protein expression Müller, Judith A.; Schäffler, Nathalie; Kellerer, Thomas ...
European journal of pharmaceutics and biopharmaceutics,
April 2024, 2024-Apr, 2024-04-00, 20240401, Letnik:
197
Journal Article
Recenzirano
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Lipid nanoparticles (LNPs) employing ionizable lipids are the most advanced technology for delivery of RNA, most notably mRNA, to cells. LNPs represent well-defined core–shell ...particles with efficient nucleic acid encapsulation, low immunogenicity and enhanced efficacy. While much is known about the structure and activity of LNPs, less attention is given to the timing of LNP uptake, cytosolic transfer and protein expression. However, LNP kinetics is a key factor determining delivery efficiency. Hence quantitative insight into the multi-cascaded pathway of LNPs is of interest to elucidate the mechanism of delivery. Here, we review experiments as well as theoretical modeling of the timing of LNP uptake, mRNA-release and protein expression. We describe LNP delivery as a sequence of stochastic transfer processes and review a mathematical model of subsequent protein translation from mRNA. We compile probabilities and numbers obtained from time resolved microscopy. Specifically, live-cell imaging on single cell arrays (LISCA) allows for high-throughput acquisition of thousands of individual GFP reporter expression time courses. The traces yield the distribution of mRNA life-times, expression rates and expression onset. Correlation analysis reveals an inverse dependence of gene expression efficiency and transfection onset-times. Finally, we discuss why timing of mRNA release is critical in the context of codelivery of multiple nucleic acid species as in the case of mRNA co-expression or CRISPR/Cas gene editing.
The intermembrane space of mitochondria contains the specific mitochondrial intermembrane space assembly (MIA) machinery that operates in the biogenesis pathway of precursor proteins destined to this ...compartment. The Mia40 component of the MIA pathway functions as a receptor and binds incoming precursors, forming an essential early intermediate in the biogenesis of intermembrane space proteins. The elements that are crucial for the association of the intermembrane space precursors with Mia40 have not been determined. In this study, we found that a region within the Tim9 and Tim10 precursors, consisting of only nine amino acid residues, functions as a signal for the engagement of substrate proteins with the Mia40 receptor. Furthermore, the signal contains sufficient information to facilitate the transfer of proteins across the outer membrane to the intermembrane space. Thus, here we have identified the mitochondrial intermembrane space sorting signal required for delivery of proteins to the mitochondrial intermembrane space.
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