Biology subjects

Gullberg, R. C.

Publications and source records attributed to Gullberg, R. C..

5 recordsLinked to original sources

bioRxiv2026

Distinct virus-derived circular RNA molecule influences host response during SARS-CoV-2 infection

Virus-derived circular RNA molecules (VcircRNAs) are expressed by many RNA viruses during infection. Putative functions include modulating viral replication and interacting with the host immune response. Some function as non-coding RNA fragments that regulate gene expression through binding to complementary RNA sequences, whereas others contain internal ribosomal entry site (IRES) sequences or non-canonical modifications that allow them to be translated. Here, we confirm the expression of a distinct SARS-CoV-2 VcircRNA molecule, circ7b8N, that has not been previously identified. We found that circ7b8N is expressed and detectable in cell culture infections and in acute infections across SARS-CoV-2 variants and shows promise for detection in post-acute clinical samples. Conservation of circ7b8N junctions is limited to the nearest phylogenetic relatives within the betacoronavirus genus but are present in other human and bat-infecting coronaviruses. Host cell gene expression is modulated by the treatment with circ7b8N agnostic of viral infection. The discovery and subsequent confirmation of circ7b8N expressed by SARS-CoV-2 provides a new biomarker for infection, and its conservation across variants suggests functional importance. Author SummaryCircular RNAs are a well-documented class of molecules expressed by mammalian cells. However, circular RNA molecules expressed by RNA viruses remain largely uncharacterized regarding their generation, specific functions, and roles in host-pathogen interactions. Our computational predictions discovered thousands of distinct circular RNA molecules expressed by SARS-CoV-2. Among these, we confirmed the presence of circ7b8N, the most abundant SARS-CoV-2-derived circular RNA identified in our sequencing data. We found that circ7b8N localizes outside the nucleus and is detectable in clinical samples collected both during and after acute SARS-CoV-2 infection. Although overexpression of circ7b8N was not found to alter viral titers, it modulated the expression of host genes related to immune response activation and membrane remodeling. This suggests that circ7b8N may simultaneously provide pro- and anti-viral functions independent of influencing viral replication. Phylogenetic analyses of coronaviruses suggest that the expression of circ7b8N is a relatively recent evolutionary event, and it is conserved across SARS-CoV-2 variants from the first five years of the pandemic. The abundant presence of circ7b8N across variants in both sequencing data and clinical samples implies it plays a multifaceted role in SARS-CoV-2 pathogenesis.

microbiology↗

bioRxiv2026

A pooled CRISPR screen reveals genes critical for erythroblast enucleation

Terminal erythroid differentiation involves dramatic cellular remodeling that culminates in the expulsion of the nucleus, a process known as enucleation. While enucleation is conserved across mammals and is crucial for the generation of fully functional erythrocytes, the mechanisms governing this process have remained largely unknown, in part because the absence of genetic material in mature, enucleated red blood cells hinders genetic experimentation. Here, we performed a pooled, forward-genetic CRISPR-Cas9 screen in enucleated red blood cells derived from primary human hematopoietic stem cells to identify genes required for enucleation. We found that Chloride Intracellular Channel 3 (CLIC3) and Vesicle-associated membrane protein 8 (VAMP8) are both necessary for terminal erythroid differentiation, yet likely act through different mechanisms. Knockdown of CLIC3 led to a delay in erythroblast differentiation, culminating in impaired enucleation. We found that the knockdown cells had increased p53 and p21 and exhibited cell cycle alterations, suggesting CLIC3 plays a crucial role in coordinating cell cycle progression during erythropoiesis. In comparison, VAMP8-depleted cells initially appear to undergo accelerated differentiation but then display a specific defect in enucleation. Transcriptional analysis of the VAMP8-knockdown cells suggested dysregulation of pathways for vesicle trafficking and actin binding, and imaging of late-stage erythroblasts revealed impaired nuclear polarization and disorganized actin. This work provides a new approach for functional genomics in enucleated cells and reveals novel factors important for terminal erythroid differentiation and enucleation. Key pointsO_LIA CROPseq-based CRISPR-Cas9 screen enables functional genomics in enucleated primary human red blood cells. C_LIO_LIChloride Intracellular Channel 3 (CLIC3) and Vesicle Associated Membrane Protein 8 (VAMP8) were identified as critical for terminal erythroid differentiation and enucleation, likely acting through two distinct mechanisms. C_LI

cell biology↗

bioRxiv2024

Perturbed pediatric serum metabolome in mild and severe dengue disease

Dengue viruses (DENVs) are the most prevalent arboviruses affecting humans. Four billion people are at risk of infection and this burden is rapidly increasing due to geographic expansion of the mosquito vector. Infection with any of the four serotypes of DENV can result in a self-limiting disease but debilitating febrile illness (DF), and some infections progress to severe disease with manifestations such as hemorrhage and shock. DENV infection drives the metabolic state of host cells for viral benefit and induces a host-immune response that has metabolic implications that link to disease. In this study, a dynamic metabolic response to DENV infection and disease was measured in 535 pediatric patient sera using liquid chromatography-mass spectrometry. The metabolome was interrogated to discover biochemical pathways and identify key metabolites perturbed in severe dengue disease. A biomarker panel of thirty-two perturbed metabolites was utilized to classify DF, and severe dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) with high sensitivity and specificity equating to a balanced accuracy of 96.9%. Some metabolites that were structurally confirmed here belong to important biochemical pathways of omega-3 and omega-6 fatty acids, sphingolipids, purines, and tryptophan metabolism. A previously reported trend between serotonin and platelets in DHF patients has been expanded upon here to reveal a major depletion of serotonin, but not platelets, in DSS patients. This study differentiated and classified DF and DHF/DSS using a serum metabolic biomarker panel based on perturbed biochemical pathways that have potential implications for severe dengue disease. One sentence summary: Metabolic biomarkers distinguish dengue hemorrhagic fever and dengue shock syndrome from dengue fever and lend insight to severe disease pathology

systems biology↗

bioRxiv2022

Shifting a Cellular Metabolic Landscape Identifies a Refractory Environment for Flavivirus Replication

Host-targeted therapeutics to control viral infection are gaining prominence given the vulnerability of viral replication at select host-interaction points and the limited possibility of developing drug resistant mutants. Nevertheless, the chemical and biological impact of many host-targeted therapeutics on both the cell and virus has not been elucidated and remains a key complication. Previously, it has been demonstrated that inhibition of fatty acid metabolism has significant antiviral potential. Here, we use a multidisciplinary approach to demonstrate how inhibition of fatty acid biosynthesis creates a metabolically refractory environment that drives viral dependence on alternate metabolic pathways for survival. By profiling the global metabolic landscape following inhibition of fatty acid biosynthesis, we identified additional biochemical pathways that, when inhibited in combination with fatty acid biosynthesis, displayed increased antiviral potential. Our studies also demonstrated that there was a direct link between changes in cellular chemical composition and the ultrastructural membrane architecture induced by viral gene products. Utilizing inhibitors to change these metabolic environments significantly impacted early viral replication and disrupted the membrane architecture critical for the viral life cycle. Here, we have defined at a molecular level how shifting metabolic landscapes can be exploited to identify combinations of therapeutics that have a greater antiviral effect. Author SummaryDengue viruses are transmitted by Aedes aegypti mosquitoes which are prevalent in the tropical and subtropical regions of the world. These viruses cause over 350 million infections annually. There are no antivirals to combat infection and the only vaccine available is suboptimal. Since these viruses are obligate pathogens, they hijack lipid metabolic pathways in host cells to drive new lipid synthesis critically required for their replication. Mechanisms of how lipid synthesis impacts viral replication is unknown. These viruses also rearrange cellular membranes to form platforms for assembly of viral replication complexes. Here, for the first time, we show that virus-hijacking of de novo fatty acid biosynthesis pathways is required for the formation of membranous replication platforms and if inhibited disrupted synthesis of replicative form viral RNA. Importantly, these inhibitors drastically rearranged the metabolic landscape of the cell resulting in an activation of compensatory nucleotide synthesis pathways that allowed the virus to survive at a low level through the inhibition. However, if both pathways were inhibited in combination, infectious virus release was reduced to below detection limits. The study demonstrates how understanding the metabolic landscape altered by specific inhibitors can lead to the discovery of compensatory metabolic pathways and targets that in combination can enhance intervention efficacy.

microbiology↗

bioRxiv2021

Acyl-CoA Thioesterases; a rheostat that controls activated fatty acids modulates dengue virus serotype 2 replication

During infection with dengue viruses (DENVs), the lipid landscape within host cells is significantly altered to assemble membrane platforms that support viral replication and particle assembly. Fatty acyl-CoAs are key intermediates in the biosynthesis of complex lipids that form these membranes. They also function as key signaling lipids in the cell. Here, we carried out loss of function studies on acyl-CoA thioesterases (ACOTs), a family of enzymes that hydrolyze fatty acyl-CoAs to free fatty acids and coenzyme A, to understand their influence on the lifecycle of DENVs. Loss of function of the type I ACOTs 1 (cytoplasmic) and 2 (mitochondrial) together significantly increased DENV serotype 2 (DENV2) viral replication and infectious particle release. However, isolated knockdown of mitochondrial ACOT2 significantly decreased DENV2 protein translation, genome replication, and infectious virus release. Furthermore, loss of ACOT7 function, a mitochondrial type II ACOT, similarly suppressed DENV2. As ACOT1 and ACOT2 are splice variants, these data suggest that location (cytosol and mitochondria, respectively) rather than function of these proteins may account for the differences in DENV2 infection phenotype. Additionally, loss of mitochondrial ACOT2 and ACOT7 expression also altered the expression of several ACOTs located in multiple organelle compartments within the cell highlighting a complex relationship between ACOTs in the DENV2 virus lifecycle.

microbiology↗