Biology subjects

Kolaczkowski, O. M.

Publications and source records attributed to Kolaczkowski, O. M..

3 recordsLinked to original sources

bioRxiv2026

HSD17B11 maintains FSP1 localization on lipid droplets to support ferroptosis defense

Ferroptosis is a regulated form of cell death driven by iron-dependent lipid peroxidation, and lipid droplets (LDs) are increasingly recognized as important regulators of this process. Consistent with this role, the anti-ferroptotic factor ferroptosis suppressor protein 1 (FSP1) is known localizing on LDs through N-myristoylation-dependent membrane targeting, where it protects LD lipids from peroxidation. Here, we identify the LD protein HSD17B11 as an additional factor required for maintaining both FSP1 localization on LDs and cellular FSP1 abundance. Silver staining followed by mass spectrometry analysis of purified LD proteins identified reduced LD-associated FSP1 in HSD17B11-deficient cells, which was further validated by immunoblotting and imaging analyses. Mechanistically, HSD17B11 physically interacted with FSP1 and was required to preserve FSP1 association with LDs. Mutational analyses further demonstrated that both FSP1 N-myristoylation and an intact HSD17B11 interaction interface are necessary for LD targeting. Correspondingly, HSD17B11 deficiency reduced LD-associated and total cellular FSP1 levels and increased cellular sensitivity to lipid oxidative stress. Together, our findings identify HSD17B11 as a previously unrecognized regulator of LD-associated FSP1 and reveal an additional mechanism controlling compartmentalized ferroptosis defense.

cell biology↗

bioRxiv2024

Manipulation of Host Cholesterol by SARS-CoV-2

Cholesterol homeostasis relies on lysosomes, which release free cholesterol from degraded lipids. We show that SARS-CoV-2 blocks lysosomal cholesterol export through its protein ORF3a. ORF3a binds the HOPS subunit VPS39, and disrupting this interaction restores cholesterol trafficking. Two mechanisms underlie this defect. First, ORF3a-VPS39 interaction traps the sorting receptor CI-MPR and the retrieval complex retromer in endosomes/lysosomes, impairing trafficking of the cholesterol transporter NPC2. Retromer deletion reproduced these defects. Second, ORF3a reduces bis(monoacylglycerol)phosphates (BMPs), lysosomal lipids required for cholesterol export. Lipidomics and proteomics revealed altered metabolism of BMP precursors, mitochondrial phosphatidylglycerols (PGs), and reduced mitochondrial proteins at lysosomes. ORF3a-VPS39 interaction decreased lysosome-mitochondrion membrane contact sites (MCS), excluding autophagy or mitochondrion-derived vesicles as routes for PG transfer. VPS39 deletion decreased the MCS and BMPs. These findings identify VPS39 as a regulator of NPC2 trafficking and BMP biosynthesis and reveal that ORF3a disrupts both pathways to block cholesterol egress.

cell biology↗

bioRxiv2024

Synergistic Role of Amino Acids in Enhancing mTOR Activation Through Lysosome Positioning

Lysosome positioning, or lysosome cellular distribution, is critical for lysosomal functions in response to both extracellular and intracellular cues. Amino acids, as essential nutrients, have been shown to promote lysosome movement toward the cell periphery. Peripheral lysosomes are involved in processes such as lysosomal exocytosis, cell migration, and metabolic signaling--functions that are particularly important for cancer cell motility and growth. However, the specific types of amino acids that regulate lysosome positioning, their underlying mechanisms, and their connection to amino acid-regulated metabolic signaling remain poorly understood. In this study, we developed a high-content imaging system for unbiased, quantitative analysis of lysosome positioning. We examined the 15 amino acids present in cell culture media and found that 10 promoted lysosome redistribution toward the cell periphery to varying extents, with aromatic amino acids showing the strongest effect. This redistribution was mediated by promoting outward transport through SLC38A9-BORC-kinesin 1/3 axis and simultaneously reducing inward transport via inhibiting the recruitment of Rab7 and JIP4 onto lysosomes. When examining the effects of amino acids on mTOR activation--a central regulator of cell metabolism--we found that the amino acids most strongly promoting lysosome dispersal, such as phenylalanine, did not activate mTOR on their own. However, combining phenylalanine with arginine, which activates mTOR without affecting lysosome positioning, synergistically enhanced mTOR activity. This synergy was lost when lysosomes failed to localize to the cell periphery, as observed in kinesin 1/3 knockout (KO) cells. Furthermore, breast cancer cells exhibited heightened sensitivity to phenylalanine-induced lysosome dispersal compared to noncancerous breast cells. Inhibition of LAT1, the amino acid transporter responsible for phenylalanine uptake, reduced peripheral lysosomes and impaired cancer cell migration and proliferation, highlighting the importance of lysosome positioning in these coordinated cellular activities. In summary, amino acid-regulated lysosome positioning and mTOR signaling depend on distinct sets of amino acids. Combining lysosome-dispersing amino acids with mTOR-activating amino acids synergistically enhances mTOR activation, which may be particularly relevant in cancer cells.

cell biology↗