Biology subjects

Norden, P. R.

Publications and source records attributed to Norden, P. R..

5 recordsLinked to original sources

bioRxiv2026

Functional partitioning of lipoic acid decouples cellular abundance from mitochondrial utilization

-Lipoic acid (LA) is widely included in "mitochondrial cocktails" recommended to patients with primary mitochondrial disorders, yet its mechanism of action remains unclear. Here, we define the intracellular availability and functional utilization of LA in mammalian cells. We show that LA exists in two functionally distinct cellular pools: a low-abundance free pool and a protein-bound pool generated through mitochondrial fatty acid synthesis (mtFAS). Disruption of the mtFAS pathway abolishes protein lipoylation and impairs oxidative phosphorylation without altering free LA levels. Conversely, supplementation with exogenous LA markedly increases free intracellular LA without restoring protein lipoylation, mitochondrial respiration, or cell proliferation. Instead, the cellular effects of LA supplementation resemble those of the antioxidant N-acetylcysteine. These findings clarify the mechanism of action of a widely used mitochondrial supplement and identify a fundamental disconnect between cellular LA abundance and mitochondrial utilization, challenging the rationale for using LA supplementation to restore mitochondrial function.

biochemistry↗

bioRxiv2026

Cells Engage Endogenous Malonate Synthesis to Drive Mitochondrial Metabolism

Malonate is often described as an endogenous inhibitor of complex II of the electron transport chain. However, the cellular source of malonate is unclear, and current knowledge concerning its metabolism is limited to the action of a single enzyme, Acyl-CoA Synthetase Family Member 3 (ACSF3), which converts malonate to malonyl-CoA in the mitochondrial matrix. One potential route of malonate metabolism downstream of ACSF3 is its consumption by the mitochondrial fatty acid synthesis (mtFAS) pathway. However, studies examining the link between ACSF3 and mtFAS have yielded conflicting results. We developed a novel mass spectrometry approach to perform stable isotope tracing into products of mtFAS, and found that while malonate is in fact a carbon source for mtFAS, ACSF3 is not required for malonate incorporation into mtFAS products. Using this method to trace other nutrients into mtFAS, we also found evidence of acetyl-CoA carboxylase 1 (ACC1)-dependent malonate synthesis from glucose. We further show that ACC1 is required for optimal mtFAS activity, with downstream effects on oxidative phosphorylation. Together these findings establish the malonate as a regulated endogenous intermediate that supports mtFAS activity and mitochondrial oxidative function.

biochemistry↗

bioRxiv2024

Mitochondrial Phosphopantetheinylation is Required for Oxidative Function

4-phosphopantetheinyl (4PP) groups are essential co-factors added to target proteins by phosphopantetheinyl transferase (PPTase) enzymes. Although mitochondrial 4PP-modified proteins have been described for decades, a mitochondrially-localized PPTase has never been found in mammals. We discovered that the cytoplasmic PPTase aminoadipate semialdehyde dehydrogenase phosphopantetheinyl transferase (AASDHPPT) is required for mitochondrial respiration and oxidative metabolism. Loss of AASDHPPT results in failed 4PP modification of the mitochondrial acyl carrier protein and blunted activity of the mitochondrial fatty acid synthesis (mtFAS) pathway. We found that in addition to its cytoplasmic localization, AASDHPPT localizes to the mitochondrial matrix via an N-terminal mitochondrial targeting sequence contained within the first 20 amino acids of the protein. Our data show that this novel mitochondrial localization of AASDHPPT is required to support mtFAS activity and oxidative function. We further identify five variants of uncertain significance in AASDHPPT that are likely pathogenic in humans due to loss of mtFAS activity.

molecular biology↗

bioRxiv2022

Endothelial FOXC1 and FOXC2 promote intestinal regeneration after ischemia-reperfusion injury

Intestinal ischemia induces mucosal damage while simultaneously activating intestinal stem cells (ISCs), which subsequently regenerate the damaged intestinal epithelium. However, whether paracrine factors secreted from vascular endothelial cells (ECs) - blood and lymphatic ECs (BECs and LECs, respectively) - regulate ISC-mediated regeneration have yet to be elucidated. Here, we identify FOXC1 and FOXC2 as essential regulators of paracrine signaling in regeneration of the small intestine after ischemia-reperfusion (I/R) injury. EC- and LEC-specific deletions of Foxc1, Foxc2, or both in mice augment I/R-induced intestinal damage by causing defects in vascular regrowth, expression of the chemokine CXCL12 and the Wnt activator R- spondin 3 in BECs and LECs, respectively, and activation of Wnt signaling in ISCs. Treatment with CXCL12 and R-spondin 3 rescues the I/R-induced intestinal damage in EC- and LEC-Foxc mutant mice, respectively. This study provides evidence that FOXC1 and FOXC2 are required for intestinal regeneration by stimulating paracrine CXCL12 and Wnt signaling.

molecular biology↗

bioRxiv2022

Differential roles of neural crest- and endothelial-derived FOXC2 in trabecular meshwork and Schlemms Canal in glaucomatous pathology

Impaired development and maintenance of the Schlemms Canal (SC) is associated with perturbed aqueous humor outflow regulation and glaucoma progression. Key molecular mechanisms, such as ANGPT/TIE2, PROX1, and VEGF-C/VEGFR-3 regulate SC development and maintenance, but mechanisms of paracrine signaling from neighboring tissues, including the trabecular meshwork (TM) are poorly understood. Here, we show Foxc2 is critical within the neural crest (NC)-derived TM and SC endothelium for development of the aqueous humor outflow pathway. In mice, NC- specific deletion of Foxc2 results in abnormal anterior eye segment development, including impaired SC morphogenesis and functional maintenance, loss of SC identity, and impaired maintenance of intraocular pressure (IOP). Visible light optical coherence tomography angiography analysis also demonstrated functional impairment of the SC in response to changes in IOP in NC-Foxc2-/- mice, suggesting increased TM stiffness. Utilization of single-cell RNA-sequencing (scRNA-seq) analysis then identified that this phenotype is predominately characterized by transcriptional changes associated with extracellular matrix organization and stiffness in TM-associated cell clusters, including increased matrix metalloproteinase (MMP) expression, which can generate soluble TIE2 that acts as an ANGPT trap. As FOXC2 is also critically involved in development of the lymphatic vasculature in other tissues, we also show that endothelial-specific deletion of Foxc2 resulted in impaired SC morphogenesis due to loss of TIE2 expression, which was rescued by deletion of the TIE2 phosphatase VE-PTP. Thus, NC-Foxc2 is critical for development of the TM, and both NC- and endothelial-Foxc2 are key for maintenance of SC identity and its morphogenesis.

developmental biology↗