Multiscale genetic architecture of donor-recipient differences reveals intronic LIMS1 mismatches associated with kidney transplant survival
Zeguo Sun, … , Peter S. Heeger, Madhav C. Menon
Zeguo Sun, … , Peter S. Heeger, Madhav C. Menon
Published September 7, 2023
Citation Information: J Clin Invest. 2023;133(21):e170420. https://doi.org/10.1172/JCI170420.
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Multiscale genetic architecture of donor-recipient differences reveals intronic LIMS1 mismatches associated with kidney transplant survival

Abstract

Donor-recipient (D-R) mismatches outside of human leukocyte antigens (HLAs) contribute to kidney allograft loss, but the mechanisms remain unclear, specifically for intronic mismatches. We quantified non-HLA mismatches at variant-, gene-, and genome-wide scales from single nucleotide polymorphism (SNP) data of D-Rs from 2 well-phenotyped transplant cohorts: Genomics of Chronic Allograft Rejection (GoCAR; n = 385) and Clinical Trials in Organ Transplantation-01/17 (CTOT-01/17; n = 146). Unbiased gene-level screening in GoCAR uncovered the LIMS1 locus as the top-ranked gene where D-R mismatches associated with death-censored graft loss (DCGL). A previously unreported, intronic, LIMS1 haplotype of 30 SNPs independently associated with DCGL in both cohorts. Haplotype mismatches showed a dosage effect, and minor-allele introduction to major-allele-carrying recipients showed greater hazard of DCGL. The LIMS1 haplotype and the previously reported LIMS1 SNP rs893403 are expression quantitative trait loci (eQTL) in immune cells for GCC2 (not LIMS1), which encodes a protein involved in mannose-6-phosphase receptor (M6PR) recycling. Peripheral blood and T cell transcriptome analyses associated the GCC2 gene and LIMS1 SNPs with the TGF-β1/SMAD pathway, suggesting a regulatory effect. In vitro GCC2 modulation impacted M6PR-dependent regulation of active TGF-β1 and downstream signaling in T cells. Together, our data link LIMS1 locus D-R mismatches to DCGL via GCC2 eQTLs that modulate TGF-β1–dependent effects on T cells.

Authors

Zeguo Sun, Zhongyang Zhang, Khadija Banu, Ian W. Gibson, Robert B. Colvin, Zhengzi Yi, Weijia Zhang, Bony De Kumar, Anand Reghuvaran, John Pell, Thomas D. Manes, Arjang Djamali, Lorenzo Gallon, Philip J. O’Connell, John Cijiang He, Jordan S. Pober, Peter S. Heeger, Madhav C. Menon

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Figure 8

GCC2 modulates generation of active TGF-β1 and downstream signaling in lymphocytes and epithelial cell lines.

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GCC2 modulates generation of active TGF-β1 and downstream signaling in ...
To investigate the cellular role of GCC2, we overexpressed either a GFP-tagged GCC2 (GCC2-OE) or a GFP-control expression plasmid in HEK-293T cells (A–C) and Jurkat T cells (D–F), followed by extraction of protein lysates, subcellular fractionation, and immunoblotting. Representative immunoblots of cellular fractions probed for GCC2, CI-M6PR, calreticulin, and GAPDH are displayed for HEK-293T (A) and Jurkat T cells (D). Dot plots show corresponding densitometric quantifications of CI-M6PR in the MFs (normalized to calreticulin) from these respective cell lines (B and E). Dot plots show corresponding ratios of active (LAP cleaved) to total TGF-β1 levels (both in pg/mL normalized to control in each paired set and analyzed by paired, 2-tailed t test) in GCC2-OE and controls in HEK-293T (C) and Jurkat T cells (F) supernatants assayed by ELISA after 24 hours serum starvation (n ≥ 4 sets). (G) Dot plots show qPCR results for IKZF2, FOXP3, and IFNG mRNA normalized to GAPDH in GCC2-OE and control Jurkat T cells (n = 3 sets). (H) Schema showing role of LIMS1 locus mismatch (A allele at rs893403) in activation of SMAD signaling pathway in response to increased GCC2 levels. GCC2 overexpression leads to increased levels of membrane-bound CI-M6PR levels via Golgi-to-endosome trafficking causing higher levels of active TGF-β1 and ultimately SMAD pathway activation (figure created using BioRender). *P < 0.05, **P < 0.01 by 2-tailed, unpaired t test. MF and CF, membrane and cytoplasmic fraction of lysate; TE, total extract; CI-M6PR, cation-independent mannose-6-phosphate receptor; IMCD, rat inner medullary collecting duct cells; TGF-β, TGF-β1.