Biology subjects

Castillo-Rojas, B.

Publications and source records attributed to Castillo-Rojas, B..

6 recordsLinked to original sources

bioRxiv2026

Genome-wide within-host evolution during prolonged SARS-CoV-2 infection in a person with advanced HIV infection

Within-host evolution during prolonged SARS-CoV-2 infection can be extensive in immunocompromised individuals, but the consequences of mutations that emerge and change in frequency over time remain difficult to predict. We characterized an XBB.1.5.10 infection lasting nearly 90 days in a person with advanced human immunodeficiency virus (HIV) infection. Genome-wide tracking of allele frequencies in samples collected on study days 43, 48, and 70 revealed a Spike N-terminal domain (NTD) deletion spanning residues 138-145 at frequencies of 33%, 80%, and 96%, respectively. A receptor-binding domain (RBD) deletion-insertion spanning residues 373-378 was detected on days 43 and 48 but not on day 70, while nsp12 C464Y was detected at approximately 8% on day 48 and 86% on day 70. We reconstructed representative Spike and non-Spike mutation profiles from the longitudinal samples and tested their effects on cell entry and viral RNA replication. The representative Spike construct for days 43 and 48 increased entry relative to XBB.1.5. Comparison with a matched construct lacking the receptor-binding domain deletion-insertion showed that the deletion-insertion enhanced entry, although it was not detected in the day 70 sample. The day 48 non-Spike profile supported the highest RNA replication among the tested replicons. Together, allele-frequency tracking revealed both transient mutations and mutations detected at progressively higher frequencies across the sampled time points. Functional testing showed that increased entry or RNA replication in vitro did not necessarily predict persistence in vivo.

microbiology↗

bioRxiv2026

Longitudinal linear-epitope profiling of SLE identifies novel antigens associated with disease and disease activity

Systemic lupus erythematosus (SLE) is the prototypical systemic autoimmune disease defined by the production of autoantibodies directed against diverse self-antigens. Antinuclear antibodies (ANA) are central to SLE diagnosis, and anti-double-stranded DNA (anti-dsDNA) antibodies are clinically used to track disease activity. However, these biomarkers provide only partial insight into the full spectrum of autoimmune changes in patients. To expand the known landscape of SLE autoreactivity shared among patients and associated with disease activity, we performed an unbiased characterization of 1,268 SLE plasma samples from 695 SLE patients and 301 healthy controls using phage immunoprecipitation and sequencing (PhIP-seq). This technique preferentially displays linear epitopes, and thus provides a complementary set of autoreactivities to previously discovered autoreactive targets. Despite the marked clinical heterogeneity of SLE, machine learning applied to this data surfaced a shared, reproducible signature that robustly discriminates between SLE and healthy controls in two validation cohorts. Autoreactivities in SLE were significantly enriched for RNA-binding protein targets. Autoreactive proteins containing Basic Acidic Dipeptides (BAD) repeat domains, features associated with biomolecular condensate formation, distinguished patients with high disease activity (by SLEDAI score) from patients with no disease activity. Among all shared autoreactivities, a peptide derived from YTHDF2 emerged as the most prevalent target across SLE cohorts occurring in ~25% of SLE patients and displayed striking sequence similarity to a peptide derived from the Epstein Barr virus (EBV) lytic switch protein BZLF1. Competition assays demonstrated higher-affinity binding to the viral peptide, supporting a model in which antiviral humoral responses to EBV may drive secondary epitope spreading toward molecularly mimicked self-antigens. Remarkably, orthogonal validation using the Luminex bead platform containing a panel of only 10 peptides, including BZLF1, demonstrated strong discrimination between SLE and healthy controls (AUC = 0.95). Together, these findings identify a rich, greatly expanded landscape of autoreactivities in SLE, candidate biomarkers associated with high disease activity, and provide mechanistic insight linking EBV-driven immunity to epitope-specific autoimmunity in lupus.

immunology↗

bioRxiv2026

Humoral Immune Dysregulation Defines High-Risk Cirrhotic Ascites

Fluid accumulates within serosal cavities in diverse disease states, where it creates biologically active and potentially pathologic niches. How these expanded microenvironments reflect and shape disease biology remains incompletely resolved. Here, we investigated human ascites fluid (n = 118), a hallmark of decompensated cirrhosis, to determine whether signatures of microbial exposure and local host response distinguish outcomes. Cirrhotic ascites was characterized by barrier remodeling, depletion of plasma cell-associated programs, and enrichment of oropharyngeal microbes. Surprisingly, detection of oropharyngeal microbes in ascites was not associated with adverse outcomes. Instead, the ascites of cirrhotic patients with poor outcomes was marked by profound plasma cell depletion and significantly reduced dimeric IgA, despite the presence of antigen-experienced B cells. Together, these findings identify local B cell dysregulation as a previously underappreciated component of cirrhosis-associated immune dysfunction. Impaired local humoral immunity, rather than microbial translocation alone, emerges as a feature of high-risk decompensated cirrhosis.

immunology↗

bioRxiv2024

Flavivirus NS1-triggered endothelial dysfunction promotes virus dissemination

The Flaviviridae are a family of viruses that include the important arthropod-borne human pathogens dengue virus (DENV), West Nile virus, Zika virus, Japanese encephalitis virus, and yellow fever virus. Flavivirus nonstructural protein 1 (NS1) is essential for virus replication but is also secreted from virus-infected cells. Extracellular NS1 acts as a virulence factor during flavivirus infection in multiple ways, including triggering endothelial dysfunction and vascular leak via interaction with endothelial cells. While the role of NS1 in inducing vascular leak and exacerbating pathogenesis is well appreciated, if and how NS1-triggered endothelial dysfunction promotes virus infection remains obscure. Flaviviruses have a common need to disseminate from circulation into specific tissues where virus-permissive cells reside. Tissue-specific dissemination is associated with disease manifestations of a given flavivirus, but mechanisms dictating virus dissemination are unclear. Here we show that NS1-mediated endothelial dysfunction promotes virus dissemination in vitro and in vivo. In mouse models of DENV infection, we show that anti-NS1 antibodies decrease virus dissemination, while the addition of exogenous NS1 promotes virus dissemination. Using an in vitro system, we show that NS1 promotes virus dissemination in two distinct ways: (1) promoting crossing of barriers and (2) increasing infectivity of target cells in a tissue- and virus-specific manner. The capacity of NS1 to modulate infectivity correlates with a physical association between virions and NS1, suggesting a potential NS1-virion interaction. Taken together, our study indicates that flavivirus NS1 promotes virus dissemination across endothelial barriers, providing an evolutionary basis for virus-triggered vascular leak. Author SummaryThe Flaviviridae contain numerous medically important human pathogens that cause potentially life-threatening infections. Over half of the worlds population is at risk of flavivirus infection, and this number is expected to increase as climate change expands the habitats of the arthropod vectors that transmit these flaviviruses. There are few effective vaccines and no therapeutics approved for prevention or treatment of flavivirus infection, respectively. Given these challenges, understanding how and why flaviviruses cause pathogenesis is critical for identifying targets for therapeutic intervention. The secreted nonstructural protein 1 (NS1) of flaviviruses is a conserved virulence factor that triggers endothelial dysfunction in a tissue-specific manner. It is unknown if this endothelial dysfunction provides any benefit for virus infection. Here we provide evidence that NS1-triggered endothelial dysfunction facilitates virus crossing of endothelial barriers and augments infection of target cells in vitro and promotes virus dissemination in vivo. This study provides an evolutionary explanation for flaviviruses evolving the capacity to trigger barrier dysfunction and highlights NS1 and the pathways governing endothelial dysfunction, as therapeutic targets to prevent flavivirus dissemination.

microbiology↗

bioRxiv2023

The Inflammasome Pathway is Activated by Dengue Virus Non-structural Protein 1 and is Protective During Dengue Virus Infection

Dengue virus (DENV) is a medically important flavivirus causing an estimated 50-100 million dengue cases annually, some of whom progress to severe disease. DENV non-structural protein 1 (NS1) is secreted from infected cells and has been implicated as a major driver of dengue pathogenesis by inducing endothelial barrier dysfunction. However, less is known about how DENV NS1 interacts with immune cells and what role these interactions play. Here we report that DENV NS1 can trigger activation of inflammasomes, a family of cytosolic innate immune sensors that respond to infectious and noxious stimuli, in mouse and human macrophages. DENV NS1 induces the release of IL-1{beta} in a caspase-1 dependent manner. Additionally, we find that DENV NS1-induced inflammasome activation is independent of the NLRP3, Pyrin, and AIM2 inflammasome pathways, but requires CD14. Intriguingly, DENV NS1-induced inflammasome activation does not induce pyroptosis and rapid cell death; instead, macrophages maintain cellular viability while releasing IL-1{beta}. Lastly, we show that caspase-1/11-deficient, but not NLRP3-deficient, mice are more susceptible to lethal DENV infection. Together, these results indicate that the inflammasome pathway acts as a sensor of DENV NS1 and plays a protective role during infection.

immunology↗

bioRxiv2021

SARS-CoV-2 Spike triggers barrier dysfunction and vascular leak via integrins and TGF-β signaling

Severe COVID-19 is associated with epithelial and endothelial barrier dysfunction within the lung as well as in distal organs. While it is appreciated that an exaggerated inflammatory response is associated with barrier dysfunction, the triggers of this pathology are unclear. Here, we report that cell-intrinsic interactions between the Spike (S) glycoprotein of SARS-CoV-2 and epithelial/endothelial cells are sufficient to trigger barrier dysfunction in vitro and vascular leak in vivo, independently of viral replication and the ACE2 receptor. We identify an S-triggered transcriptional response associated with extracellular matrix reorganization and TGF-{beta} signaling. Using genetic knockouts and specific inhibitors, we demonstrate that glycosaminoglycans, integrins, and the TGF-{beta} signaling axis are required for S-mediated barrier dysfunction. Our findings suggest that S interactions with barrier cells are a contributing factor to COVID-19 disease severity and offer mechanistic insight into SARS-CoV-2 triggered vascular leak, providing a starting point for development of therapies targeting COVID-19 pathogenesis.

microbiology↗