Biology subjects

Lima, T.

Publications and source records attributed to Lima, T..

5 recordsLinked to original sources

bioRxiv2024

Serum proteomics reveals high-affinity and convergent antibodies by tracking SARS-CoV2 hybrid immunity to emerging variants of concern

The rapid spread of SARS-CoV2 and continuing impact on human health has prompted the need for effective and rapid development of monoclonal antibody therapeutics. In this study, we interrogate polyclonal antibodies in serum and B cells from whole blood of three donors with SARS-CoV2 immunity to find high-affinity anti-SARS-CoV2 antibodies to escape variants. Serum IgG antibodies were selected by affinity to the receptor-binding domain (RBD) and non-RBD sites on the spike protein of Omicron subvariant B.1.1.529 from each donor. Antibodies were analyzed by bottom-up mass spectrometry, and matched to single- and bulk-cell sequenced repertoires for each donor. Antibodies observed in serum were recombinantly expressed, and characterized to assess domain binding, cross-reactivity between different variants, and capacity to inhibit RBD binding to host protein. Donors infected with early Omicron subvariants had serum antibodies with subnanomolar affinity to RBD that show binding activity to a newer Omicron subvariant BQ.1.1. The donors also showed a convergent immune response. Serum antibodies and other single- and bulk-cell sequences were similar to publicly reported anti-SARS-CoV-2 antibodies, and characterized serum antibodies had the same variant-binding and neutralization profiles as their reported public sequence. The serum antibodies analyzed were a subset of anti-SARS-CoV2 antibodies in the B cell repertoire, which demonstrates significant dynamics between the B cells and circulating antibodies in peripheral blood.

immunology↗

bioRxiv2023

Inhibition of CERS1 in skeletal muscle exacerbates age-related muscle dysfunction

Age-related muscle wasting and dysfunction render the elderly population vulnerable and incapacitated, while underlying mechanisms are poorly understood. Here, we implicate the CERS1 enzyme of the de novo sphingolipid synthesis pathway in the pathogenesis of age-related skeletal muscle impairment. In humans, CERS1 abundance declines with aging in skeletal muscle cells and, correlates with biological pathways involved in muscle function and myogenesis. Furthermore, CERS1 is upregulated during myogenic differentiation. Pharmacological or genetic inhibition of CERS1 in aged mice blunts myogenesis and deteriorates aged skeletal muscle mass and function, which is associated with the occurrence of morphological features typical of inflammation and fibrosis. Ablation of the CERS1 orthologue lagr-1 in C. elegans similarly exacerbates the age-associated decline in muscle function and integrity. We discover genetic variants reducing CERS1 expression in human skeletal muscle and Mendelian randomization analysis in the UK biobank cohort shows that these variants reduce muscle grip strength and overall health. In summary, our findings link age-related impairments in muscle function to a reduction in CERS1, thereby underlining the importance of the sphingolipid biosynthesis pathway in age-related muscle homeostasis.

physiology↗

bioRxiv2022

A lysosomal surveillance response (LySR) that reduces proteotoxicity and extends healthspan

Lysosomes are cytoplasmic organelles central for the degradation of macromolecules to maintain cellular homeostasis and health. Here, we discovered an adaptive lysosomal transcriptional response that we termed the Lysosomal Surveillance Response (LySR). Typified by the induction of a large group of transcripts involved in lysosomal function and proteolysis, the LySR can be triggered by silencing of specific vacuolar H+-ATPase subunits in Caenorhabditis elegans. Notably, LySR activation enhances the clearance of protein aggregates in worm models of Alzheimers and Huntingtons disease and amyotrophic lateral sclerosis, thereby boosting fitness and extending lifespan. The GATA transcription factor, ELT-2, regulates the LySR program as well as its associated beneficial effects. In mammalian cells, overexpression of GATA4/GATA6, the mammalian orthologs of ELT-2, is sufficient to induce the expression of multiple lysosome-specific proteases and alleviate proteotoxicity. Activating the LySR pathway may therefore represent an attractive mechanism to reduce proteotoxicity and, as such, potentially extend healthspan. HighlightsO_LIRNAi of specific v-ATPase subunits extends C. elegans lifespan and activates LySR C_LIO_LIGATA transcription factor ELT-2 regulates LySR and LySR-associated lifespan extension C_LIO_LILySR activation reduces protein aggregates and extends worm healthspan C_LIO_LIOverexpression of GATA4/GATA6 alleviates amyloid-{beta} proteotoxicity in mammalian cells C_LI

cell biology↗

bioRxiv2022

Neutrophil IL-1β secretion induced by ExoS expressing Pseudomonas aeruginosa is dependent on NLRP3 and Gasdermin D

Macrophages infected with Gram-negative bacteria expressing Type III secretion system (T3SS) activate the NLRC4 inflammasome, resulting in Gasdermin D (GSDMD)-mediated IL-1{beta} secretion and pyroptosis. Here we examined inflammasome signaling in neutrophils infected with Pseudomonas aeruginosa strain PAO1 that expresses the T3SS effectors ExoS and ExoT. IL-1{beta} secretion by neutrophils required the T3SS needle and translocon proteins and GSDMD. In macrophages, PAO1 and mutants lacking ExoS and ExoT ({Delta}exoST) stimulated NLRC4 for IL-1{beta} secretion. While IL-1{beta} release from{Delta} exoST infected neutrophils was also NLRC4-dependent, this was redirected to NLRP3-dependence by PAO1 infection via the ADP ribosyl transferase activity of ExoS. Genetic and pharmacologic approaches revealed that NLRP3, but not NLRC4, was essential for bacterial killing and limiting disease severity in a murine model of P. aeruginosa corneal infection. This reveals a novel role for ExoS ADPRT in regulating inflammasome subtype usage by neutrophils versus macrophages and an unexpected role for NLRP3 in P. aeruginosa keratitis.

microbiology↗

bioRxiv2019

Serum proteomics expands on identifying more high-affinity antibodies in immunized rabbits than deep B-cell repertoire sequencing alone

AO_SCPLOWBSTRACTC_SCPLOWRabbits are a model for immunology studies, and monoclonal antibodies developed from rabbits have been sought after to empower immunoassays in a variety of applications. High-throughput characterization of circulating serum antibodies in response to specific antigens is highly impactful for both humoral immunology studies and antibody development. A combination of high throughput sequencing of antibody transcripts from B cells and proteomic analysis of serum antibodies, an approach referred to as immunoproteogenomics, is applied to profile the immune response of rabbits to {beta}-galactosidase (Beta-gal) in both recombinant antigen and peptide antigen immunization formats. The use of intact protein antigen resulted in observing 56.3% more heavy chains CDR3s in serum than immunization with peptide antigens. Additionally, sampling peripheral blood mononuclear cells (PBMCs) for B-cell repertoire sequencing at different time points throughout the immunization was found to capture 47.8%-72.8% of total proteomically observed heavy chain CDR3s, and would serve well in replacing sequencing the B cell rich, but more difficult to access spleen or bone marrow compartments. Despite B-cell repertoire sequencing to depths of 2M to 10M reads, we found proteomic evidence supporting at least 10% of serum antibodies are still missed. Further improvements to proteomic analysis techniques would enable more precise characterization of antibodies circulating in serum and determine antibody protein sequences missed by repertoire sequencing.

immunology↗