Biology subjects

Mears, J.

Publications and source records attributed to Mears, J..

6 recordsLinked to original sources

bioRxiv2026

Genetic demultiplexing and transcript start site identification from nanopore sequencing of 10x Genomics multiome libraries

Short-read Illumina sequencing of 10x Genomics single-nucleus multiome libraries captures only the 3 end of RNA transcripts, losing transcription start site (TSS) information. Here we demonstrate nanopore sequencing of 10x multiome libraries, which enables the profiling of full length transcripts. We show concordance with common short-read sequencing based workflows including successful genetic demultiplexing of nanopore data despite its higher error rate. We compare TSS identified using nanopore sequencing of multiome cDNA to those identified using a short-read 5 assay, and provide an optimized approach for the preprocessing of nanopore reads prior to TSS identification. We find that nanopore sequencing of multiome cDNA captures a median of 63% of the TSS detected by the 5 assay.

bioinformatics↗

bioRxiv2025

Deep profiling of lupus nephritis kidneys reveals dynamic changes in myeloid cells associated with disease progression

ObjectivesLupus nephritis (LN) is a common, potentially fatal manifestation of systemic lupus erythematosus. We aim to gain new insights into the immune responses underlying LN and their relation to the histologic heterogeneity observed in this disease, focusing on myeloid cells. MethodsSingle-cell RNA-sequencing (scRNA-seq) was used to profile dissociated kidney samples from 156 LN patients and 30 healthy individuals. Spatial transcriptomics (ST), utilizing a gene panel designed to capture all myeloid subsets identified in the scRNA-seq data, was applied to kidney samples acquired from 6 LN patients and 2 healthy controls. ResultsWe generated a full catalog of the myeloid subsets found in LN kidneys. Our analyses indicated that an increase in irreversible tissue damage, as measured by the NIH chronicity index (CI), is associated with a gradual switch of the local immune response from one dominated by monocytes and macrophages to one featuring expanded CD4+ T, GZMK+CD8+ T, B and dendritic cells, with a parallel decrease in the interferon response. In proliferative/mixed LN only, the degree of active inflammation correlates with expansion of disease-specific macrophage (DMac) subsets, which later contract as the CI increases. Trajectory analysis of the scRNA-seq data suggested that DMacs arise from both infiltrating monocytes and tissue-resident macrophages; this was supported by the ST data, as well as cell cultures. DMacs are indicated to interact with parietal epithelial cells, promoting the development of glomerulosclerosis. ConclusionsWe suggest a detailed picture of the changes in the kidney immune mechanisms in LN as this disease progresses.

immunology↗

bioRxiv2025

The Legionella effector RidL promotes mitochondrial fragmentation through phosphorylation activation of the large GTPase Drp1

Intracellular pathogens such as Legionella pneumophila secrete effector proteins that manipulate host cell processes to promote bacterial survival. One such effector, RidL, is known to inhibit retrograde trafficking by interacting with the retromer complex via its N-terminal domain. Here, we identify a second function of RidL mediated by its C-terminal domain, which directly binds to the mitochondrial fission GTPase Drp1 and related large GTPases. In vitro, RidL reduces Drp1 GTPase activity and disrupts its oligomerization. During infection, RidL localizes to mitochondria, enhances the accumulation of Drp1 and the outer membrane protein Tom20, and impairs mitochondrial dynamics and function. Moreover, in L. pneumophila-infected cells, RidL promotes phosphorylation of Drp1 at Ser616, leading to Drp1 activation and mitochondrial fragmentation. These findings establish RidL as a bifunctional effector that targets both the retromer complex and Drp1 through distinct domains. By interfering with host mitochondrial dynamics, RidL enables L. pneumophila to remodel host organelles and optimize conditions for intracellular replication.

microbiology↗

bioRxiv2025

A population-scale atlas of blood and tissue in lupus nephritis

One Sentence SummaryA single-cell atlas of paired blood and tissue samples from Lupus Nephritis patients and healthy controls identified stromal and immune populations within renal tissue, including the scar-associated macrophage populations, which correlate with and may drive renal disease activity. Lupus nephritis (LN), a severe manifestation of Systemic Lupus Erythematosus (SLE), is a heterogeneous disease driven by diverse immune and tissue cell types. We obtained 538K single-cell and 140K single-nuclear profiles from kidney biopsies of 155 LN patients and 30 pre-implantation transplant biopsy controls, along with 325K single-cell blood profiles overlapping many of these patients. We identified key tissue cell types and cell states, and immune cell states; we were able to determine cell states that were tissue specific, and those that were present in the blood. We observed that LN pathological features are significantly associated with cell states using differential gene expression and Covarying Neighborhood Analysis (CNA). These analyses revealed broad changes in cell states associated with irreversible chronic tissue damage. After controlling for the effects of ongoing tissue damage, we observed that expansion of key glomerular and Scar Associated Macrophages (SAMs) populations tracked with increasing inflammatory disease activity. SAMs appear to drive LN fibrosis and, in active disease, infiltrate the glomeruli more than other myeloid cells. These observations strongly support that therapeutic targeting of myeloid populations may offer an as-of-yet unproven strategy to prevent renal inflammation and ongoing kidney damage in LN.

molecular biology↗

bioRxiv2022

Cellular deconstruction of inflamed synovium defines diverse inflammatory phenotypes in rheumatoid arthritis

Rheumatoid arthritis (RA) is a prototypical autoimmune disease that causes destructive tissue inflammation in joints and elsewhere. Clinical challenges in RA include the empirical selection of drugs to treat patients, inadequate responders with incomplete disease remission, and lack of a cure. We profiled the full spectrum of cells in inflamed synovium from patients with RA with the goal of deconstructing the cell states and pathways characterizing pathogenic heterogeneity in RA. Our multicenter consortium effort used multi-modal CITE-seq, RNA-seq, and histology of synovial tissue from 79 donors to build a >314,000 single-cell RA synovial cell atlas with 77 cell states from T, B/plasma, natural killer, myeloid, stromal, and endothelial cells. We stratified tissue samples into six distinct cell type abundance phenotypes (CTAPs) individually enriched for specific cell states. These CTAPs demonstrate the striking diversity of RA synovial inflammation, ranging from marked enrichment of T and B cells (CTAP-TB) to a congregation of specific myeloid, fibroblast, and endothelial cells largely lacking lymphocytes (CTAP-EFM). Disease-relevant cytokines, histology, and serology metrics are associated with certain CTAPs. This comprehensive RA synovial atlas and molecular, tissue-based CTAP stratification reveal new insights into RA pathology and heterogeneity, which could lead to novel targeted-treatment approaches in RA.

genomics↗

bioRxiv2022

DRP1-mediated mitochondrial fission is essential to maintain cristae morphology and bioenergetics

Mitochondria and peroxisomes are both dynamic signaling organelles that constantly undergo fission. While mitochondrial fission is known to coordinate cellular metabolism, proliferation, and apoptosis, the physiological relevance of peroxisome dynamics and the implications for cell fate are not fully understood. DRP1 (dynamin-related protein 1) is an essential GTPase that executes both mitochondrial and peroxisomal fission. Patients with de novo heterozygous missense mutations in the gene that encodes DRP1, DNM1L (Dynamin 1 Like), present with encephalopathy due to defective mitochondrial and peroxisomal fission (EMPF1). EMPF1 is a devastating neurodevelopmental disease with no effective treatment. To interrogate the mechanisms by which DRP1 mutations cause cellular dysfunction, we used human-derived fibroblasts from patients with mutations in DRP1 who present with EMPF1. As expected, patient cells display elongated mitochondrial morphology and lack of fission. Patient cells display a lower coupling efficiency of the electron transport chain, increased proton leak, and upregulation of glycolysis. In addition to these metabolic abnormalities, mitochondrial hyperfusion results in aberrant cristae structure and hyperpolarized mitochondrial membrane potential, both of which are tightly linked to the changes in metabolism. Peroxisome structure is also severely elongated in patient cells and results in a potential functional compensation of fatty acid oxidation. Understanding the mechanism by which DRP1 mutations cause these metabolic changes will give insight into the role of mitochondrial dynamics in cristae maintenance and the metabolic capacity of the cell, as well as the disease mechanism underlying EMPF1.

cell biology↗