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Significance
Two common variants in the APOL1 gene explain most of the high rate of kidney disease in people of recent African ancestry. However, not all APOL1 high-risk individuals develop kidney disease. Here we identified the UBD locus as a genetic modifier of APOL1 kidney disease using admixture mapping. Focal segmental glomerulosclerosis patie...
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Context 1
... also compared the genome-wide ancestries between different groups. We replicated our findings by genotyping an additional 62 FSGS samples ( Fig. S1 and Table S1). ...
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... each individual, after local ancestry inference, we de- termined genome-wide ancestry and local ancestry at loci throughout the genome. The APOL1 high-risk genotype indi- viduals studied included individuals with FSGS, AA individuals with ESRD, AA individuals from the Jackson Heart Study with or without CKD, and high-risk individuals imputed from two collec- tions of samples available through the dbGAP database without kidney phenotype information but genotyped using the same plat- form as we used for the FSGS discovery and ESRD groups (phs000507 and phs000560) (Table S1). ...
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... this study, we demonstrate that individuals with FSGS and a high-risk APOL1 genotype are enriched for recent African ancestry at the UBD locus. The moderate number of APOL1 high-risk Tet: EV G0 G1 G2 EV G0 G1 G2 - - - - + + + + 25KD-- 22KD-- 46KD-- 32KD-- B Fig. 3. Induction of APOL1 G1 and G2 leads to up-regulation of UBD transcripts but reduced UBD protein levels. (A) Quantitative PCR of UBD and GAPDH transcripts after APOL1 induction in T-REx-293 cells with tet (50 ng/mL; eight replicates). ...
Citations
... Since the original publication in 2010, attempts have been made by different investigators to identify genetic modifiers of APOL1-associated kidney diseases. Nevertheless, most of these associations have not been validated or replicated [9][10][11][12]. On the other hand, viral infections, exposure to alpha interferon, intra renal high expression of APOL1 protein from high risk genotype (pre-clinical models) have all been demonstrated and robustly replicated as environmental disease modifiers [13][14][15][16][17][18]. ...
... A commonly used cell model to examine the cytotoxic mechanisms of APOL1 variants are HEK-293 cells. In some cases, APOL1-G1 and G2 expression elicited greater cytotoxicity than G0 [8][9][10][11][12] , but not in others 13,14 . ...
... It also remains to be demonstrated whether cation conductance actually correlates with cytotoxicity for all the haplotypes in the relevant cell type, podocytes. Another issue is that cytotoxicity is clearly dependent on APOL1 expression levels 10,13,21 and the majority of reports (including Lannon et al.) 8,[10][11][12][13]15,17 show total APOL1 expression, rather than APOL1 levels at the plasma membrane, where cation channel activity occurs 9,14 . This is potentially important because APOL1 variants may fold differently compared to G0 22,23 and might thus be differentially retained in the endoplasmic reticulum versus being transported to the cell surface 11 . ...
Homozygous Apolipoprotein L1 (APOL1) variants G1 and G2 cause APOL1-mediated kidney disease, purportedly acting as surface cation channels in podocytes. APOL1-G0 exhibits various single nucleotide polymorphisms, most commonly haplotype E150K, M228I and R255K (“KIK”; the Reference Sequence is “EMR”), whereas variants G1 and G2 are mostly found in a single “African” haplotype background (“EIK”). Several labs reported cytotoxicity with risk variants G1 and G2 in KIK or EIK background haplotypes, but used HEK-293 cells and did not verify equal surface expression. To see if haplotype matters in a more relevant cell type, we induced APOL1-G0, G1 and G2 EIK, KIK and EMR at comparable surface levels in immortalized podocytes. G1 and G2 risk variants (but not G0) caused dose-dependent podocyte death within 48h only in their native African EIK haplotype and correlated with K⁺ conductance (thallium FLIPR). We ruled out differences in localization and trafficking, except for possibly greater surface clustering of cytotoxic haplotypes. APOL1 surface expression was required, since Brefeldin A rescued cytotoxicity; and cytoplasmic isoforms vB3 and vC were not cytotoxic. Thus, APOL1-EIK risk variants kill podocytes in a dose and haplotype-dependent manner (as in HEK-293 cells), whereas unlike in HEK-293 cells the KIK risk variants did not.
... The UBD gene, located in the 6q21.3 region of the chromosome, is unique in encoding a ubiquitin-like protein that directly targets substrates for proteasomal degradation (26). UBD dysregulation has been reported in various cancers and can promote tumor progression through multiple pathways (27). ...
Increasing evidence suggests that pseudogenes play crucial roles in various cancers, yet their functions and regulatory mechanisms in glioma pathogenesis remain enigmatic. In the present study, a novel pseudogene was identified, UBDP1, which is significantly upregulated in glioblastoma and positively correlated with the expression of its parent gene, UBD. Additionally, high levels of these paired genes are linked with a poor prognosis for patients. In the present study, clinical samples were collected followed by various analyses including microarray for long non-coding RNAs, reverse transcription-quantitative PCR, fluorescence in situ hybridiza-tion and western blotting. Cell lines were authenticated and cultured then subjected to various assays for proliferation, migration, and invasion to investigate the molecular mechanisms. Bioinformatic tools identified miRNA targets, and luciferase reporter assays validated these interactions. A tumor xenograft model in mice was used for in vivo studies. In vitro and in vivo studies have demonstrated that UBDP1, localized in the cytoplasm, functions as a tumor-promoting factor influencing cell proliferation, migration, invasion and tumor growth. Mechanistic investigations have indicated that UBDP1 exerts its oncogenic effects by decoying miR-6072 from UBD mRNA, thus forming a competitive endogenous RNA network, which results in the enhanced oncogenic activity of UBD. The present findings offered new insights into the role of pseudogenes in glioma progression, suggesting that targeting the UBDP1/miR-6072/UBD network may serve as a potential therapeutic strategy for glioma patients.
... variants for APOL1-mediated kidney disease and, particularly, FSGS, remains elusive. The few reported in the literature still require validation 12,13 . ...
African Americans have a significantly higher risk of developing chronic kidney disease, especially focal segmental glomerulosclerosis -, than European Americans. Two coding variants (G1 and G2) in the APOL1 gene play a major role in this disparity. While 13% of African Americans carry the high-risk recessive genotypes, only a fraction of these individuals develops FSGS or kidney failure, indicating the involvement of additional disease modifiers. Here, we show that the presence of the APOL1 p.N264K missense variant, when co-inherited with the G2 APOL1 risk allele, substantially reduces the penetrance of the G1G2 and G2G2 high-risk genotypes by rendering these genotypes low-risk. These results align with prior functional evidence showing that the p.N264K variant reduces the toxicity of the APOL1 high-risk alleles. These findings have important implications for our understanding of the mechanisms of APOL1-associated nephropathy, as well as for the clinical management of individuals with high-risk genotypes that include the G2 allele.
... ; https://doi.org/10.1101/2023.08.02.23293554 doi: medRxiv preprint genetic variants for APOL1-mediated kidney disease and, particularly, FSGS, remains elusive. The few reported in the literature still require validation 12,13 . ...
Black Americans have a significantly higher risk of developing chronic kidney disease (CKD), especially focal segmental glomerulosclerosis (FSGS), than European Americans. Two coding variants (G1 and G2) in the APOL1 gene play a major role in this disparity. While 13% of Black Americans carry the high-risk recessive genotypes, only a fraction of these individuals develops FSGS or kidney failure, indicating the involvement of additional disease modifiers.
Here, we show that the presence of the APOL1 p.N264K missense variant, when co-inherited with the G2 APOL1 risk allele, substantially reduces the penetrance of the G1G2 and G2G2 high-risk genotypes by rendering these genotypes low-risk. These results align with prior functional evidence showing that the p.N264K variant reduces the toxicity of the APOL1 high-risk alleles. These findings have important implications for our understanding of the mechanisms of APOL1-associated nephropathy, as well as for the clinical management of individuals with high-risk genotypes that include the G2 allele.
... 28 Genetic modifiers such as SMOC2, DEF1B, UBD, NUDT7, and GSTB1 have also been identified. [29][30][31][32][33] In addition, environmental factors such as air pollution have been implicated in the development of AMKD in patients carrying APOL1 risk variants, probably due to cellular stress mechanisms. [34][35][36][37] In sum, AMKD has diverse clinical manifestations and it is likely that complex epistatic and environmental interactions result in differential pathological cellular programming. ...
... In addition to those mentioned previously, other mechanisms implicated in AMKD that have been suggested include activation of protein kinase R, 69 which appears during viral infections and inhibits protein synthesis, increased autophagic cell death (mediated by the BCL2homology 3 domain within the pore-forming domain of APOL1), 70 or the alteration of the ubiquitin-proteasome system, which reduces ubiquitin levels prolonging the intracellular retention of proteins (including APOL1 itself). 31 In addition, APOL1 G1 and G2 variants have been shown to have high affinity for suPAR activated avb3 integrin on podocytes in the progression of chronic kidney disease. 71 These and the aforementioned mechanisms are summarized in Figure 1. ...
Apolipoprotein L1 (APOL1) high-risk variants confer an increased risk for the development and progression of kidney disease among individuals of recent African ancestry. Over the past several years, significant progress has been made in understanding the pathogenesis of APOL1-mediated kidney diseases (AMKD), including genetic regulation, environmental interactions, immunomodulatory, proinflammatory and apoptotic signaling processes, as well as the complex role of APOL1 as an ion channel. Collectively, these findings have paved the way for novel therapeutic strategies to mitigate APOL1-mediated kidney injury. Precision medicine approaches are being developed to identify subgroups of AMKD patients who may benefit from these targeted interventions, fueling hope for improved clinical outcomes. This review summarizes key mechanistic insights in the pathogenesis of AMKD, emergent therapies, and discusses future challenges.
... 11,12 Additionally, the effect of APOL1 risk genotype is highly variable between specific kidney diseases and self-declared Black ancestry is associated with more rapid progression independent of APOL1 genotype, suggesting the presence of modifiers. 14,16,17 One known modifier is elevations in interferon levels, which induce APOL1 expression in podocytes, potentially explaining the HIV and SARS-CoV-2 associated collapsing glomerulopathy seen in individuals with high-risk APOL1 genotypes. 5,10,[13][14][15] Recent studies have implicated the NLRP3 inflammasome and pyroptosis in interferon induced APOL1 cytotoxicity, suggesting these pathways as potential modifiers for APOL1-associated phenotypes. ...
... 14,16,17 One known modifier is elevations in interferon levels, which induce APOL1 expression in podocytes, potentially explaining the HIV and SARS-CoV-2 associated collapsing glomerulopathy seen in individuals with high-risk APOL1 genotypes. 5,10,[13][14][15] Recent studies have implicated the NLRP3 inflammasome and pyroptosis in interferon induced APOL1 cytotoxicity, suggesting these pathways as potential modifiers for APOL1-associated phenotypes. 16,17 Several studies have attempted to identify genetic modifiers of the effect of APOL1 risk genotype on kidney disease but no strong signals have been identified and replicated. ...
... The gene-based collapsing analyses did not identify significant modifiers of the effect of APOL1 genotype, including the previously suggested modifiers AHDC1 and UBD. 14,23 The suggestive signals in NLRP1, and the inflammasome gene-set are noteworthy because this pathway has been implicated in APOL1 mediated cytotoxicity. 61 The cytotoxicity of APOL1 risk variants may be due to increased expression following viral infections, inflammation and interferon signaling. ...
Background:
APOL1 genotype has significant effects on kidney disease development and progression that vary among specific causes of kidney disease, suggesting the presence of effect modifiers.
Methods:
We assessed the risk of kidney failure and eGFR decline rate in patients with chronic kidney disease (CKD) carrying high-risk (N=239) and genetically matched low-risk (N=1187) APOL1 genotypes. Exome sequencing revealed monogenic kidney diseases. Exome-wide association studies and gene-based and gene-set based collapsing analyses evaluated genetic modifiers of the effect of APOL1 genotype on CKD.
Results:
Compared to genetic ancestry-matched CKD cases with low-risk APOL1 genotypes, those with high-risk APOL1 genotypes had a higher risk of kidney failure (HR= 1.58), a higher decline in eGFR (6.55 vs 3.63 mL/min/1.73m2/year), and were younger at time of kidney failure (45.1 vs53.6 years), with the G1/G1 genotype demonstrating the highest risk. The rate for monogenic kidney disorders was lower among CKD patients with high-risk APOL1 genotypes (2.5%) compared to those with low-risk genotypes (6.7%). Gene-set analysis identified an enrichment of rare missense variants in the inflammasome pathway in individuals with high-risk APOL1 genotypes and CKD (OR = 1.90).
Conclusions:
In this genetically matched cohort, high-risk APOL1 genotypes were associated with an increased risk of kidney failure and eGFR decline rate, with a graded risk between specific high-risk genotypes and a lower rate of monogenic kidney disease. Rare missense variants in the inflammasome pathway may act as genetic modifiers of APOL1 effect on kidney disease.
... UBD genotypes associated with lower UBD expression may correlate with risk of APOL1-nephropathy. [104,105] GSTM1 Individuals APOL1-nephropathy and null GSTM1 alleles have increased rate kidney disease progression, [106] Pharmacologic Therapeutic IFN FSGS and collapsing nephropathy have been seen almost exclusively in people with high-risk APOL1 genotypes [61,62] Infectious HIV HIVAN is extremely rare in people without high-risk APOL1 genotypes [58,107] COVID-19 COVAN has been reported almost exclusively in people with high-risk APOL1 genotypes [63] Inflammatory Lupus Greater risk of ESKD and collapsing lupus nephritis [108,109] ...
Background: Genetic variants in APOL1 are a major contributor to the increased risk of kidney disease in people of recent African ancestry.
Summary: Two alleles in the APOL1 gene, referred to as G1 and G2, confer increased risk of kidney disease under a recessive model of risk inheritance. Disease risk is inherited as a recessive trait: People with genotypes G1/G1, G2/G2, and G1/G2 (i.e. a risk allele from each parent) have increased risk for what we refer to here as APOL1-associated kidney disease. In the United States, about 13% of the self-identified African-American population has a high-risk genotype. As we discuss below, APOL1 is an unusual disease gene. Most studies to date have suggested that the G1 and G2 variants have toxic, gain-of-function effects on the encoded protein.
Key Message: In this article, we review key concepts critical to understanding APOL1-associated kidney disease, emphasizing ways in which it is highly atypical for a human disease-causing gene.
... The mechanisms by which the APOL1 variants modulate the risk of kidney diseases is complex and still obscure. However, some of the proposed mechanisms largely supported by animal and in vitro studies include podocyte injury by the increased APOL1 expression through interferon, APOL1 inflammatory-mediated apoptosis leading to proteinuria and glomerular scarring, inhibition of protein synthesis by the APOL1 G1 and G2 variants through activation of protein kinase R, APOL1-triggered mitochondrial dysfunction, and dysregulation of ubiquitin D (UBD)-a ubiquitin-like modifier protein [29][30][31]. An additional mechanistic may be the triggering of cell lysis and intracellular loss of potassium by G1 and G2 APOL1 risk alleles [32]. ...
Background:
The risk of various types of kidney disease is significantly increased in the presence of APOL1 high-risk genotype (carriage of two risk alleles), particularly HIV-associated nephropathy (HIVAN). However, there are discrepancies in the existing evidence about the level of association between APOL1 high-risk genotype and the risk of kidney diseases in people living with HIV (PLWHIV).
Methods:
This systematic review and meta-analysis was conducted to assess the relationship between the APOL1 genotypes and kidney disease in the HIV population. An a priori protocol registered on PROSPERO (ID: CRD42021253877), was followed by a systematic search of five electronic databases. Database-specific search terms were used to identify observational studies that evaluated the outcomes chosen in the review, based on a set of prespecified eligibility criteria. Using a random effect model, the odds ratios (ORs) and their corresponding 95% confidence intervals (CIs) were pooled for the meta-analysis.
Results:
After screening 4418 citations, 14 articles comprising 11,069 participants were included in this review. The risk of chronic kidney disease (CKD) in the HIV positive population was significantly increased in the presence of two APOL1 risk alleles (OR 4.65 [95% CI 3.51-6.15]). Also, a significant association was observed between the carriage of two risk APOL1 variants and proteinuria (OR 2.58 [95% CI 2.05-3.25]), HIVAN (OR 16.67 [95% CI 10.22-27.19]), and progression to end-stage kidney disease (ESKD) hazard ratio: 1.79 (95% CI 1.20-2.66).
Conclusion:
This review highlights a strong association between the presence of two risk APOL1 variants and an increased risk of kidney disease in PLWHIV, and provides a more precise estimate of the effect size, with smaller 95% CIs for CKD, HIVAN, and progression to ESKD.
... APOL1 (OMIM gene locus #603743) variants G0 (African Common, or AfCom), G1 (rs73885319, rs60910145) and G2 cDNAs (rs71785313) were as previously described [10,26,33,42,57]. (See Shah et al. [42] for complete APOL1 cDNA sequences). ...
... Attached cells at 50-80% confluency were transfected with 0.5 µg plasmid cDNA and 1 µl each of Lipofectamine P3000 and Lipofectamine reagent (Ther-moFisher) per well in OptiMEM (Life Technologies) as per manufacturers' instructions. Stable, tetracyclineinducible, transgenic HEK T-Rex-293 cell lines with human APOL1 G0, G1, or G2 integrated at a defined locus [10,26,33,42,57] were grown in Dulbecco's modified Eagle's medium (DMEM, Corning) supplemented with 10% tetracycline system-approved fetal bovine serum (Atlanta Biologicals), 0.2 mg/ml zeocin, 2 ug/ml blasticidin and 1% antibiotic-antimycotic at 37 °C and 5% CO 2 . Cells were induced to overexpress APOL1 by 6-24 h exposure to 50 ng/ml doxycycline, as indicated. ...
Two heterozygous missense variants (G1 and G2) of Apolipoprotein L1 (APOL1) found in individuals of recent African ancestry can attenuate the severity of infection by some forms of Trypanosoma brucei. However, these two variants within a broader African haplotype also increase the risk of kidney disease in Americans of African descent. Although overexpression of either variant G1 or G2 causes multiple pathogenic changes in cultured cells and transgenic mouse models, the mechanism(s) promoting kidney disease remain unclear. Human serum APOL1 kills trypanosomes through its cation channel activity, and cation channel activity of recombinant APOL1 has been reconstituted in lipid bilayers and proteoliposomes. Although APOL1 overexpression increases whole cell cation currents in HEK-293 cells, the ion channel activity of APOL1 has not been assessed in glomerular podocytes, the major site of APOL1-associated kidney diseases. We characterize APOL1-associated whole cell and on-cell cation currents in HEK-293 T-Rex cells and demonstrate partial inhibition of currents by anti-APOL antibodies. We detect in primary human podocytes a similar cation current inducible by interferon-γ (IFNγ) and sensitive to inhibition by anti-APOL antibody as well as by a fragment of T. brucei Serum Resistance-Associated protein (SRA). CRISPR knockout of APOL1 in human primary podocytes abrogates the IFNγ-induced, antibody-sensitive current. Our novel characterization in HEK-293 cells of heterologous APOL1-associated cation conductance inhibited by anti-APOL antibody and our documentation in primary human glomerular podocytes of endogenous IFNγ-stimulated, APOL1-mediated, SRA and anti-APOL-sensitive ion channel activity together support APOL1-mediated channel activity as a therapeutic target for treatment of APOL1-associated kidney diseases.
