Search bioRxiv⌕ Search

bioRxiv · 10.1101/2023.03.27.534273

APOL1 kidney risk variants in glomerular diseases modeled in transgenic mice

Abstract

APOL1 high-risk variants partially explain the high kidney disease prevalence among African ancestry individuals. Many mechanisms have been reported in cell culture models, but few have been demonstrated in mouse models. Here we characterize two models: (1) HIV- associated nephropathy (HIVAN) Tg26 mice crossed with bacterial artificial chromosome (BAC)/APOL1 transgenic mice and (2) interferon-{psi} administered to BAC/APOL1 mice. Both models showed exacerbated glomerular disease in APOL1-G1 compared to APOL1-G0 mice. HIVAN model glomerular bulk RNA-seq identified synergistic podocyte-damaging pathways activated by the APOL1-G1 allele and by HIV transgenes. Single-nuclear RNA-seq revealed podocyte-specific patterns of differentially-expressed genes as a function of APOL1 alleles. Eukaryotic Initiation factor-2 pathway was the most activated pathway in the interferon-{psi} model and the most deactivated pathway in the HIVAN model. HIVAN mouse model podocyte single-nuclear RNA-seq data showed similarity to human focal segmental glomerulosclerosis (FSGS) glomerular bulk RNA-seq data. Furthermore, single-nuclear RNA-seq data from interferon-{psi} mouse model podocytes (in vivo) showed similarity to human FSGS single-cell RNA- seq data from urine podocytes (ex vivo) and from human podocyte cell lines (in vitro) using bulk RNA-seq. These data highlight differences in the transcriptional effects of the APOL1-G1 risk variant in a model specific manner. Shared differentially expressed genes in podocytes in both mouse models suggest possible novel glomerular damage markers in APOL1 variant-induced diseases. Transcription factor Zbtb16 was downregulated in podocytes and endothelial cells in both models, possibly contributing to glucocorticoid-resistance. In summary, these findings in two mouse models suggest both shared and distinct therapeutic opportunities for APOL1 glomerulopathies. Significance statementCoding variants in APOL1, encoding apolipoprotein L1, contribute to kidney disease in individuals with African ancestry. The mechanisms for glomerular injury remain incompletely understood. We studied two transgenic mouse models, HIV-associated nephropathy and interferon-{psi} administration. Using glomerular and single-nuclear RNA sequencing, we identified genes differentially expressed among mice with kidney risk alleles (G1) and the common variant (G0). Both models exhibited up-regulation of genes that indicated podocyte damage with risk alleles compared to the common variant. One gene down-regulated in both models was Zbtb16, encoding a transcription factor, that may contribute to glucocorticoid-resistance. Overall, the findings suggest both shared and distinct alterations in the two disease models.

Source connections

Explore related subjects

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Yoshida, T., Latt, K. Z., Santo, B. A., Shrivastav, S., Zhao, Y., Fenaroli, P., Chung, J.-Y., Hewitt, S. M., Tutino, V. M., Sarder, P., Rosenberg, A. Z., Winkler, C. A., Kopp, J. B.. 2023-03-27. APOL1 kidney risk variants in glomerular diseases modeled in transgenic mice. https://doi.org/10.1101/2023.03.27.534273

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

NAE1-Dependent Protein Neddylation Preserves Endothelial Identity and Vascular Integrity

Background: Endothelial dysfunction is a central driver of cardiovascular and inflammatory diseases, yet the post-translational mechanisms that preserve endothelial homeostasis remain incompletely understood. Protein neddylation, the covalent conjugation of a ubiquitin-like modifier, regulates diverse cellular processes, yet its physiological role in the vascular endothelium remains unknown. This study investigated whether protein neddylation is required to preserve endothelial identity and vascular homeostasis. Methods: We generated tamoxifen-inducible endothelial-specific Nae1 knockout mice to inhibit neddylation and combined bulk RNA sequencing, single-cell and single-nucleus transcriptomics, quantitative proteomics, biochemical analyses, and gain- and loss-of-function approaches to define the role of endothelial neddylation in vascular homeostasis and inflammatory injury. Results: Endothelial-specific Nae1 deletion caused rapid mortality associated with vascular leakage, platelet accumulation, inflammation, and multi-organ injury. Multi-omics analyses demonstrated profound loss of endothelial identity, characterized by suppression of core endothelial programs and activation of inflammatory, procoagulant, and pyroptotic pathways. Single-cell analyses revealed progressive endothelial dysfunction culminating in depletion of the endothelial population and remodeling of the vascular niche. Mechanistically, endothelial neddylation deficiency activated gasdermin D (GSDMD)- and gasdermin E (GSDME)-dependent pyroptosis, whereas dual inhibition of GSDMD and GSDME markedly attenuated inflammatory transcriptomic remodeling, vascular injury, hepatocyte death, immune cell infiltration, and platelet accumulation. Translational analyses demonstrated reduced endothelial neddylation in experimental endotoxemia and decreased expression of neddylation pathway components in human atherosclerosis and COVID-19 datasets. Conversely, restoration of endothelial neddylation partially reversed inflammatory endothelial transcriptomic reprogramming in vivo. Conclusions: NAE1-dependent protein neddylation is an essential regulator of endothelial identity and vascular integrity. Loss of endothelial neddylation promotes gasdermin-dependent pyroptosis and thrombo-inflammatory vascular injury, whereas restoration of the neddylation pathway mitigates inflammatory endothelial dysfunction. These findings identify endothelial neddylation as a fundamental mechanism maintaining vascular homeostasis and a potential therapeutic target for cardiovascular and inflammatory diseases.

pathology↗

EZH2 inhibition stimulates viral mimicry in resting splenic B cells

In mammalian cells expression of repetitive genomic sequences is repressed by heterochromatin, underscoring the potential threat of repeat expression to cellular homeostasis. However, the specific consequences of ectopic repeat expression remains unclear. Here we demonstrate that EZH2 inhibitors stimulate repeat misexpression and cell death in resting splenic B cells. We show that B cells are uniquely sensitive to these agents because of high levels of H3K27me3 at repeats and correspondingly low DNA methylation. We generated a pattern recognition receptor loss-of-function mouse model called RIC with mutations in Rigi, Ifih1 (MDA5), and Cgas to specifically block the consequences of repeat misexpression. In both WT and RIC mutant B cells, EZH2 inhibition caused focused loss of H3K27me3 at repetitive elements and upregulated their expression. However, expression of inflammatory chemokines and cell death were interrupted by the RIC mutations. Furthermore, the chemokine expression patterns induced by EZH2 inhibitors resemble the B cell response to Epstein-Barr virus infection. This study demonstrates a viral mimicry effect induced by pharmacological activation of repeat expression that induces inflammation and B cell death.

pathology↗

NON-TOXIC ACID-FREE GLYOXAL FIXATIVE FOR VETERINARY HISTOPATHOLOGY, IMMUNOHISTOCHEMISTRY AND MOLECULAR ANALYSIS

Formaldehyde fixation is worldwide the most used system for histopathological examination. However, its toxicity is well known, and preservation of proteins and nucleic acids is not optimal. Alternative fixatives warranting similar morphological quality of tissues and costs, but lacking toxicity and allowing better preservation of proteins and nucleic acids would therefore increase both safety of operators and quality of molecular analysis in pathology. This multi-institutional study aimed to compare the morphological, histochemical, immunohistochemical (IHC), and molecular analyses outcomes of a newly patented, non-toxic, acid-free Glyoxal (GAF) fixative with neutral buffered formaldehyde (NBF). Tissues from a total of 73 subjects were analyzed, including 13 necropsies. Gross features were preserved after GAF fixation, with no tissue hardening or discoloration. Cellular ultrastructure was also better preserved with GAF and histology and histochemistry on GAF-fixed samples showed good results when compared to NBF-fixed samples, with the exception of loss of tinctorial affinity of erythrocytes and mast cell granules. IHC analyses also showed comparable results with only slight and rare protocol adjustment. DNA and RNA yields were higher from GAF-fixed samples (P<0.05) and the tested genes (p53 and COX1) were better amplified. RNA scope showed positive results for c-KIT expression in GAF-fixed mast cell tumors. Based on these data, the non-toxic GAF fixative allows good macroscopical, histological and immunohistochemical analyses of tissue samples, including on-field application, and better molecular analyses when compared to NBF. This represents a promising possibility for teaching, diagnostic, and research in veterinary pathology.

pathology↗