Biology subjects

Hintzen, J. C. J.

Publications and source records attributed to Hintzen, J. C. J..

3 recordsLinked to original sources

bioRxiv2026

Mitochondrial copper stabilizes lipoylated TCA cycle proteins to sustain metabolism and proliferation

Copper (Cu) is an essential cofactor for mitochondrial cytochrome c oxidase, yet whether it directly regulates mitochondrial metabolism beyond respiration remains unclear. Here we show that mitochondrial Cu, delivered by SLC25A3, is required to maintain the stability of lipoylated TCA cycle proteins. Loss of Slc25a3 or pharmacological Cu depletion selectively destabilized the lipoylated E2 subunits of mitochondrial dehydrogenases and the lipoylation enzymes LIPT1 and LIPT2, an effect not reproduced by acute electron transport chain inhibition. Mechanistically, we find that Cu directly engages the reduced lipoyl moiety using chemical probes and synthetic peptide approaches. Cu depletion impaired PDH and OGDH activity, rewired TCA cycle metabolism, and imposed a dependence on pyruvate carboxylase for anaplerosis. This metabolic defect depleted aspartate, suppressed mTORC1 signaling, and limited proliferation. Conversely, selective delivery of Cu to the mitochondria restored lipoylation, TCA cycle function, and cell growth. Together, these findings identify mitochondrial Cu as a structural regulator of the lipoylation machinery and reveal a direct link between Cu homeostasis and central carbon metabolism.

cell biology↗

bioRxiv2025

Extended nuclear glycosylation is a common post-translational modification

In eukaryotes, glycans modify proteins in the secretory pathway and the extracellular space. Aside from nucleocytoplasmic O-GlcNAc, glycosylation is not considered a relevant post-translational modification in other cellular compartments. Here, we challenge this long-standing paradigm by showing that extended O-glycans are commonly found on intranuclear proteins. Through comprehensive genetic and biochemical analyses, we conclusively demonstrate that these O-glycans stem from the secretory pathway, yet are found on nuclear proteins across mammalian cell lines and primary cells. Using knock-out cell lines, we show mechanistically that nuclear glycans are shuttled to the nucleus via active vesicular transport. We identify several of these intranuclear glycoproteins as RNA-binding proteins, including KHSRP/FUBP2, RBM12, and RPP30. Lastly, we show that site-specific glycosylation of RPP30 is crucial for effective tRNA processing. Overall, our findings suggest a much broader role for glycosylation in regulating cellular functions and open up investigation into the role of glycans in more biological processes.

molecular biology↗

bioRxiv2025

Targeted Sortase A Inhibition by Novel Peptidomimetic Antivirulents against Staphylococcal Infections

Antibiotic resistance is a critical public health issue, causing resistant bacterial strains to be increasingly difficult to control. Antivirulence therapies, which target bacterial virulence factors rather than kill bacteria, present a promising approach. Sortase enzymes, particularly SrtA, are crucial for Gram-positive bacterial virulence by anchoring surface proteins essential for bacterial adhesion and biofilm formation to the bacterial outer cell wall. This study evaluates the selectivity of the peptidomimetic inhibitor BzLPRDSar towards various Gram-positive bacteria. The BzLPRDSar significantly inhibited biofilm formation in multidrug-resistant S. aureus and S. epidermidis. Conversely, it showed variable and generally lower selectivity to Gram-positive species such as E. faecalis, B. cereus and S. agalactiae. The selectivity towards Staphylococcus species is attributed to conserved structural elements in the SrtA enzyme, particularly the {beta}7/{beta}8 loop region with a key tryptophan, likely facilitating strong binding interactions with the inhibitor. ImportanceThis study addresses the pressing issue of antibiotic resistance by exploring antivirulence therapy as an innovative alternative to conventional antibiotics, focusing on inhibiting bacterial virulence rather than bacterial growth. By evaluating the selectivity of the peptidomimetic inhibitor BzLPRDSar against various Gram-positive bacteria, the study highlights its potent selectivity in inhibiting biofilm formation in multidrug-resistant S. aureus and S. epidermidis. The findings underscore the potential of targeting conserved structural elements in bacterial Sortase enzymes, particularly in Staphylococcal species, to develop more selective and effective antivirulence therapies.

microbiology↗