Biology subjects

Zschocke, J.

Publications and source records attributed to Zschocke, J..

6 recordsLinked to original sources

bioRxiv2025

Biochemical signatures from dried blood spots in untargeted metabolics are influenced by matrix effects

Dried blood spots (DBS) represent a convenient clinical sample material, offering low infection risk, easy transport, and long-term metabolite stability. However, applying samples such as whole blood, serum or plasma onto filter paper introduces an additional matrix, potentially affecting metabolite extraction. Here, we compare metabolite recovery from liquid samples and their filter paper analogue using both targeted (acylcarnitines, amino acids) and untargeted metabolomics. Significant matrix effects were observed for some compounds, especially for dicarboxylic acylcarnitines (C3DC-C6DC) and specific amino acids (cystine, cystathionine). We did not identify specific metabolite characteristics that may predicted altered recovery. In a cohort of 229 authentic DBS samples -- including patients diagnosed with inherited metabolic disorders, obesity or under a ketogenic diet -- untargeted profiling combined with random-forest machine learning led to an effective stratification. Notably, C4DC, despite strong matrix effects, was ranked in the top ten variables of this random-forest model. With adequate validation, DBS can be safely used for diagnostic purposes despite possible matrix effects, but care must be taken in the comparison of values obtained when different sample materials are used.

biochemistry↗

bioRxiv2024

TXNIP mediates LAT1/SLC7A5 endocytosis to reduce amino acid uptake in cells entering quiescence

Entry and exit from cellular quiescence require dynamic adjustments in nutrient acquisition, yet the mechanisms by which quiescent cells downregulate amino acid (AA) transport remain poorly understood. Here, we demonstrate that cells entering quiescence select plasma membrane-resident AA transporters for endocytosis and lysosomal degradation, to match AA uptake with reduced translation. We identify the -arrestin TXNIP as a key regulator of AA uptake during quiescence, since it mediates the endocytosis of the SLC7A5-SLC3A2 (LAT1-4F2hc) transporter complex in response to reduced AKT signaling. Mechanistically, TXNIP interacts with HECT-type ubiquitin ligases to facilitate transporter ubiquitination. Loss of TXNIP disrupts this regulation, resulting in dysregulated AA uptake, sustained mTORC1 signaling, and accelerated quiescence exit. A novel TXNIP loss-of-function mutation in a patient with severe metabolic disease further supports its role in nutrient homeostasis and human health. These findings highlight TXNIPs role in controlling SLC7A5-SLC3A2 mediated AA acquisition with implications for quiescence biology and disease.

cell biology↗

bioRxiv2024

Mitochondrial cardiolipin metabolism controlled by tafazzin enables ferroptosis

Mitochondria are important producers of reactive oxygen species, which are involved in triggering ferroptosis, a lipid peroxidation driven form of cell death. Paradoxically, in the rare inherited metabolic disease Barth Syndrome, we discovered a protection from erastin-induced ferroptosis, despite intrinsically elevated mitochondrial ROS levels. The affected transacylase tafazzin, which is mutated in Barth Syndrome, is pivotal for remodeling of the dimeric phospholipid cardiolipin. They unique to mitochondria and essential for shaping their membrane functionalities. We investigated which downstream effects of the pathogenic membrane alterations are responsible for the protective effect against ferroptosis. We found that while iron metabolism, the unsaturation of membrane lipids, and the metabolic activity of the cells were modifying factors, they were not causal. However, we observed that cardiolipin abnormalities are not limited to impair only inner, but also outer mitochondrial membrane protein complexes. Specifically, they impact abundance and oligomerization of voltage-dependent anion channels (VDAC) in response to oxidative stress. We found that tafazzin deficiency via alteration of cardiolipins affects VDAC functionality, thereby modulating small molecule transport and signaling between mitochondria and the remaining cell. This is in line with a reduction of mitochondria-associated membranes (MAM) sites that are formed through VDACs and trapping ROS in mitochondria where they are unable to contribute to ferroptosis. These findings demonstrate that the mitochondrial membrane architecture impacting on subcellular small molecule distribution crucially impact on the manifestation of cell fate decisions, including ferroptosis.

molecular biology↗

bioRxiv2020

Unequivocal mapping of molecular ether lipid species by LC-MS/MS in plasmalogen-deficient mice

Deficient ether lipid biosynthesis in rhizomelic chondrodysplasia punctata and other disorders is associated with a wide range of severe symptoms including small stature with proximal shortening of the limbs, contractures, facial dysmorphism, congenital cataracts, ichthyosis, spasticity, microcephaly, and mental disability. Mouse models are available but show less severe symptoms. In both humans and mice it has remained elusive which of the symptoms can be attributed to lack of plasmanyl or plasmenyl ether lipids. The latter compounds, better known as plasmalogens, harbor a vinyl ether double bond conferring special chemical and physical properties. Discrimination between plasmanyl and plasmenyl ether lipids is a major analytical challenge, especially in complex lipid extracts with many isobaric species. Consequently, these lipids are often neglected also in recent lipidomic studies. Here we present a comprehensive LC-MS/MS based approach that allows unequivocal distinction of these two lipid subclasses based on their chromatographic properties. The method was validated using a novel plasmalogen-deficient mouse model which lacks plasmanylethanolamine desaturase and therefore cannot form plasmenyl ether lipids. We demonstrate that plasmanylethanolamine desaturase deficiency causes an accumulation of plasmanyl species, a little studied but biologically important substance class.

biochemistry↗

bioRxiv2020

Fatty Acyl Availability Modulates Cardiolipin Composition and Alters Mitochondrial Function in HeLa Cells

The molecular assembly of cells depends not only on their balance between anabolism and catabolism, but to a large degree also on the building blocks available in the environment. For cultivated mammalian cells, this is largely determined by the composition of the growth medium used. Here we study the impact of medium lipids on mitochondrial membrane architecture and function by combining LC-MS/MS lipidomics and functional tests with lipid supplementation experiments in an otherwise serum- and lipid-free cell culture model. We demonstrate that the composition of mitochondrial cardiolipins (CL) strongly depends on the lipid environment in cultured cells and prefers the incorporation of essential linoleic acid over other fatty acids. Simultaneously, the mitochondrial respiratory complex I activity was altered, whereas the matrix-localized enzyme citrate synthase was unaffected. This suggests a link between membrane composition and respiratory capacity. In summary, we find a strong dependency of central mitochondrial features on the type of lipids contained in the growth medium. Thus, this underlines the importance of considering these factors when using and establishing cell culture models in biomedical research. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=76 SRC="FIGDIR/small/937433v2_ufig1.gif" ALT="Figure 1"> View larger version (17K): org.highwire.dtl.DTLVardef@1960166org.highwire.dtl.DTLVardef@135d267org.highwire.dtl.DTLVardef@88868forg.highwire.dtl.DTLVardef@13a3c83_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2017

A homozygous variant in mitochondrial RNase P subunit PRORP is associated with Perrault syndrome characterized by hearing loss and primary ovarian insufficiency

Perrault syndrome is a rare autosomal recessive condition characterised by sensorineural hearing loss in both sexes and primary ovarian insufficiency in 46 XX, females. It is genetically heterogeneous with biallelic variants in six genes identified to date (HSD17B4, HARS2, LARS2, CLPP, C10orf2 and ERAL1). Most genes possessing variants associated with Perrault syndrome are involved in mitochondrial translation. We describe a consanguineous family with three affected individuals homozygous for a novel missense variant c.1454C>T; p.(Ala485Val) in KIAA0391, encoding proteinaceous RNase P (PRORP), the metallonuclease subunit of the mitochondrial RNase P complex, responsible for the 5-end processing of mitochondrial precursor tRNAs. In RNase P activity assays, RNase P complexes containing the PRORP disease variant produced ~35-45% less 5-processed tRNA than wild type PRORP. Consistently, the accumulation of unprocessed polycistronic mitochondrial transcripts was observed in patient dermal fibroblasts, leading to an observable loss of steady-state levels of mitochondrial oxidative phosphorylation components. Expression of wild type KIAA0391 in patient fibroblasts rescued tRNA processing. Immunohistochemistry analyses of the auditory sensory epithelium from postnatal and adult mouse inner ear showed a high level of PRORP in the efferent synapses and nerve fibres of hair cells, indicating a possible mechanism for the sensorineural hearing loss observed in affected individuals. We have identified a variant in an additional gene associated with Perrault syndrome. With the identification of this disease-causing variant in KIAA0391, reduced function of each of the three subunits of mitochondrial RNase P have now been associated with distinct clinical presentations.\n\nAuthor SummaryPerrault syndrome is a rare genetic condition which results in hearing loss in both sexes and ovarian dysfunction in females. Perrault syndrome may also cause neurological symptoms in some patients. Here, we present the features and genetic basis of the condition in three sisters affected by Perrault syndrome. The sisters did not have pathogenic variants in any of the genes previously associated with Perrault syndrome. We identified a change in the gene KIAA0391, encoding PRORP, a subunit of the mitochondrial RNase P complex. Mitochondrial RNase P is a key enzyme in RNA processing in mitochondria. Impaired RNA processing reduces protein production in mitochondria, which we observed in patient cells along with high levels of unprocessed RNA. When we expressed wild type PRORP in patient cells, the RNA processing improved. We also investigated PRORP localisation in the mouse inner ear and found high levels in the synapses and nerve fibers that transmit sound. It may be that disruption of RNA processing in the mitochondria of these cells causes hearing loss in this family.

genetics↗