Biology subjects

Kline, I.

Publications and source records attributed to Kline, I..

4 recordsLinked to original sources

bioRxiv2025

Human Cytomegalovirus Infection Reduces an Endogenous Antiviral Fatty Acid by Promoting Host Metabolism

Some viruses, including human cytomegalovirus (HCMV), induce the synthesis of fatty acids and lipids to ensure that the lipid environment of infected cells supports virus replication. HCMV infection broadly reprograms metabolism to ensure central carbon metabolism provides the metabolites required for anabolic synthesis of nucleotides, proteins, and lipids while also meeting the energy demands placed on the infected cells. While HCMV infection increases the levels of most very long chain fatty acids (VLCFA), we found that the levels of erucic acid (EA), a C22:1 monounsaturated VLCFA, are reduced. Treating infected cells with EA disrupted a late step in virus replication, resulting in the release of virions with reduced infectivity. Moreover, we used lipidomics to determine that EA-treated cells had elevated levels of lipids containing a combination of a C22:1 tail and a VLC polyunsaturated fatty acid tail (VLC-PUFA). We demonstrate that fatty acid elongase 5 (ELOVL5) mediated production of VLC-PUFAs is stimulated by HCMV infection. ELOVL5 aided the increase in lipids with C22:1 plus VLC-PUFA tails following EA treatment and reduced the overall level of C22:1 in HCMV-infected cells. Moreover, we found that ELOVL5 mollified EA inhibition of HCMV replication, suggesting ELOVL5 plays a critical role in reducing the level of an endogenous FA with antiviral properties. Our study provides insight into how infection may increase the synthesis of an antiviral metabolite or FA and how the virus may evade their antiviral effect by promoting their metabolism.

microbiology↗

bioRxiv2025

Human cytomegalovirus promotes de novo PC synthesis during early virus replication

Human cytomegalovirus (HCMV) infection reprograms metabolism, including lipid synthesis. While several metabolite-related pathways have been demonstrated to have altered activity in infected cells, the alteration of lipid-related pathways by HCMV has not been examined beyond fatty acid synthesis and elongation. In this study, we addressed this lack of understanding by focusing on phosphatidylcholine (PC), a class of lipids we previously showed is increased by HCMV infection in human foreskin fibroblasts. Here, we found that HCMV infection increases the abundance of PCs in several different fibroblasts and, similarly, in endothelial and epithelial cells. Additionally, HCMV elevates PC levels regardless of the level of confluency, type of growth medium, and presence of serum. Next, we investigated if HCMV alters the activity in the three PC synthesis pathways. We demonstrate that HCMV infection promotes the activity in the de novo PC synthesis pathway using a 13C-choline isotopic tracer and liquid chromatography high resolution tandem mass spectrometry (LC-MS/MS). Infection did not alter the activity in the other two pathways. Moreover, we examined the kinetics of PC remodeling by HCMV and found that the de novo synthesis pathway is promoted and the PC lipidome shifts 24 hours post infection. That led us to examining if the early stages of replication are sufficient to alter PC levels. After inhibiting late virus replication, we found that HCMV alters the PC lipidome independent of late gene expression. Overall, this work suggests that an immediate-early or early viral protein promotes the reprogramming of host lipid metabolism to ensure the synthesis of a lipidome necessary to support HCMV replication. IMPORTANCEHuman cytomegalovirus (HCMV) is a common herpesvirus that establishes a lifelong and persistent infection in its human host (1). HCMV infection in most people does not cause overt disease (1). However, in immunocompromised individuals, severe CMV-associated disease can lead to permanent disabilities and even death (1, 2). Additionally, congenital CMV is the leading infectious cause of birth defects (3, 4). Viruses have evolved to hijack host metabolic pathways to facilitate their replication cycle. In this study, we determine that HCMV promotes the activity in the de novo pathway of phosphatidylcholine (PC) synthesis. We demonstrate that the activity in the other PC synthesis pathways, the PEMT and Lands cycle, is unaltered by HCMV infection. Moreover, we found that HCMV infection alters metabolic activity to increase the PC lipidome before 48 hpi. Additionally, we demonstrate that changes in PC lipids during virus replication is independent of late gene expression. Together, our findings demonstrate that infection promotes the de novo PC pathway to increase PC lipids during the early stages of virus replication.

microbiology↗

bioRxiv2021

The p97-UBXD8 complex modulates ER-Mitochondria contact sites by modulating membrane lipid saturation and composition

The intimate association between the endoplasmic reticulum (ER) and mitochondrial membranes at ER-mitochondria contact sites serves as a platform for several critical cellular processes, in particular lipid synthesis. Enzymes involved in lipid biosynthesis are enriched at contacts and membrane lipid composition at contacts is distinct relative to surrounding membranes. How contacts are remodeled and the subsequent biological consequences of altered contacts such as perturbed lipid metabolism remains poorly understood. Here we show that the p97 AAA-ATPase and its ER-tethered ubiquitin-X domain adaptor 8 (UBXD8) regulate the prevalence of ER-mitochondria contacts. The p97-UBXD8 complex localizes to contacts and loss of this complex increases contacts in a manner that is dependent on p97 catalytic activity. Quantitative proteomics of purified contacts demonstrates alterations in proteins regulating lipid metabolism upon loss of UBXD8. Furthermore, lipidomics studies indicate significant changes in distinct lipid species in UBXD8 knockout cells. We show that loss of p97-UBXD8 results in perturbed contacts due to an increase in membrane lipid saturation via SREBP1 and the lipid desaturase SCD1. Aberrant contacts in p97-UBXD8 loss of function cells can be rescued by supplementation with unsaturated fatty acids or overexpression of SCD1. Perturbation of contacts and inherent lipid synthesis is emerging as a hallmark to a variety of human disorders such as neurodegeneration. Notably, we find that the SREBP1-SCD1 pathway is negatively impacted in the brains of mice with p97 mutations that cause neurodegeneration. Our results suggest that contacts are exquisitely sensitive to alterations to membrane lipid composition and saturation in a manner that is dependent on p97-UBXD8.

cell biology↗

bioRxiv2021

Human Cytomegalovirus uses a Host Stress Response to Balance the Elongation of Saturated/Monounsaturated and Polyunsaturated Very Long Chain Fatty Acids

Stress and virus infection are known to regulate lipid metabolism in cells. Human cytomegalovirus infection induces fatty acid (FA) elongation and increases the cellular abundance of lipids with very long-chain FA tails (VLCFAs). While reprogramming of metabolism can be stress-related, the role of stress in HCMV reprogramming of lipid metabolism is poorly understood. In this study, we engineered cells to knockout PKR-like ER kinase (PERK) in the ER stress pathway and measured lipid changes using lipidomics to determine if PERK is needed for lipid changes associated with HCMV infection. We found that in HCMV-infected cells, PERK promotes the increase in the levels of phospholipids with saturated FA (SFA) and monounsaturated FA (MUFA) VLCFAs tails. Consistent with the SFA/MUFA lipidome changes, PERK enhances the protein levels of FA elongase 7 (ELOVL7), which elongates SFA and MUFA VLCFAs. Additionally, we found that increases in the elongation of polyunsaturated fatty acids (PUFAs) associated with HCMV infection was independent of PERK and that lipids with PUFA tails accumulated in HCMV-infected PERK knockout cells. Consistent with the PUFA lipidome changes, the protein levels of ELOVL5, which elongates PUFAs, are increased by HCMV infection through a PERK-independent mechanism. These observations show that PERK differentially regulates ELOVL7 and ELOVL5, creating a balance between the synthesis of lipids with SFA/MUFA tails and PUFA tails. Additionally, we found that PERK was necessary for virus replication and the infectivity of released viral progeny. Overall, our findings indicate that PERK--and more broadly, ER stress--may be necessary for membrane biogenesis needed to generate infectious HCMV virions. IMPORTANCEHCMV is a common herpesvirus that establishes lifelong persistent infections. While infection is asymptomatic in most people, HCMV causes life-threatening illnesses in immunocompromised people, including transplant recipients and cancer patients. Additionally, HCMV infection is a leading cause of congenital disabilities. HCMV replication relies on lipid synthesis. Here, we demonstrated that the ER stress mediator, PERK, controls fatty acid (FA) elongation and cellular abundance of several types of lipids following HCMV infection. Specifically, we found that PERK promotes FA elongase 7 synthesis of lipids with saturated/monounsaturated very long-chain FA tails which are important for building the viral membrane of infectious HCMV virions. Overall, our study shows that PERK is an essential host factor that supports HCMV replication and promotes lipidome changes caused by HCMV infection.

microbiology↗