DACH1 protects podocytes from experimental diabetic injury and modulates PTIP-H3K4Me3 activity
Aili Cao, Jianhua Li, Morad Asadi, John M. Basgen, Bingbing Zhu, Zhengzi Yi, Song Jiang, Tomohito Doke, Osama El Shamy, Niralee Patel, Paolo Cravedi, Evren U. Azeloglu, Kirk N. Campbell, Madhav Menon, Steve Coca, Weijia Zhang, Hao Wang, Ke Zen, Zhihong Liu, Barbara Murphy, John C. He, Vivette D. D’Agati, Katalin Susztak, Lewis Kaufman
Aili Cao, Jianhua Li, Morad Asadi, John M. Basgen, Bingbing Zhu, Zhengzi Yi, Song Jiang, Tomohito Doke, Osama El Shamy, Niralee Patel, Paolo Cravedi, Evren U. Azeloglu, Kirk N. Campbell, Madhav Menon, Steve Coca, Weijia Zhang, Hao Wang, Ke Zen, Zhihong Liu, Barbara Murphy, John C. He, Vivette D. D’Agati, Katalin Susztak, Lewis Kaufman
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Research Article Nephrology

DACH1 protects podocytes from experimental diabetic injury and modulates PTIP-H3K4Me3 activity

Abstract

Dachshund homolog 1 (DACH1), a key cell-fate determinant, regulates transcription by DNA sequence–specific binding. We identified diminished Dach1 expression in a large-scale screen for mutations that convert injury-resistant podocytes into injury-susceptible podocytes. In diabetic kidney disease (DKD) patients, podocyte DACH1 expression levels are diminished, a condition that strongly correlates with poor clinical outcomes. Global Dach1 KO mice manifest renal hypoplasia and die perinatally. Podocyte-specific Dach1 KO mice, however, maintain normal glomerular architecture at baseline, but rapidly exhibit podocyte injury after diabetes onset. Furthermore, podocyte-specific augmentation of DACH1 expression in mice protects from DKD. Combined RNA sequencing and in silico promoter analysis reveal conversely overlapping glomerular transcriptomic signatures between podocyte-specific Dach1 and Pax transactivation-domain interacting protein (Ptip) KO mice, with upregulated genes possessing higher-than-expected numbers of promoter Dach1-binding sites. PTIP, an essential component of the activating histone H3 lysine 4 trimethylation (H3K4Me3) complex, interacts with DACH1 and is recruited by DACH1 to its promoter-binding sites. DACH1-PTIP recruitment represses transcription and reduces promoter H3K4Me3 levels. DACH1 knockdown in podocytes combined with hyperglycemia triggers target gene upregulation and increases promoter H3K4Me3. These findings reveal that in DKD, diminished DACH1 expression enhances podocyte injury vulnerability via epigenetic derepression of its target genes.

Authors

Aili Cao, Jianhua Li, Morad Asadi, John M. Basgen, Bingbing Zhu, Zhengzi Yi, Song Jiang, Tomohito Doke, Osama El Shamy, Niralee Patel, Paolo Cravedi, Evren U. Azeloglu, Kirk N. Campbell, Madhav Menon, Steve Coca, Weijia Zhang, Hao Wang, Ke Zen, Zhihong Liu, Barbara Murphy, John C. He, Vivette D. D’Agati, Katalin Susztak, Lewis Kaufman

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Figure 8

DACH1 knockdown in podocytes combined with hyperglycemia triggers target gene derepression and increases promoter H3K4Me3 levels.

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DACH1 knockdown in podocytes combined with hyperglycemia triggers target...
(A) Western blot comparing levels of DACH1 protein between stably transduced human podocyte cell lines that each express a distinct DACH1-targeted shRNA or scrambled shRNA control plasmid. (B) qPCR for 4 DACH1-target genes (NELL2, NTRK3, PAMR1, and MYCL1) in DACH1 knockdown shRNA no. 1 and control transduced podocytes with and without hyperglycemia. Under basal conditions, DACH1 knockdown (KD) podocytes showed modest transcriptional derepression of each of the 4 genes compared with control shRNA podocytes (top panel). *P < 0.003; **P < 0.0004; ***P < 0.0004. In control podocytes cultured in high glucose, none of the 4 genes demonstrated increased mRNA expression levels compared with identical cells grown in mannitol (middle panel). †P < 0.005; ††P < 0.0007. In DACH1 KD podocytes grown in high glucose, however, transcriptional derepression was augmented dramatically (lower panel). #P < 0.015; ##P < 0.003; ###P < 0004; ####P < 0.0001. (C) ChIP-qPCR was performed using an H3K4Me3-specific antibody and primers specific for the promoter of NELL2 comparing chromatin extracted from DACH1 KD and control podocytes with and without high glucose. The combination of DACH1 KD with hyperglycemia caused an increase in levels of promoter bound H3K4Me3. Primer pair 1: *P < 0.002; **P < 0.001; ***P < 0.001. Primer pair 2: †P < 0.015; ††P < 0.004; †††P < 0.001. (D) ChIP qPCR as in C, but with primers specific for the NTRK3 promoter. Primer pair 1: *P < 0.015; **P < 0.003; ***P < 0.002. Primer pair 2: †P < 0.012; ††P < 0.004; †††P < 0.004. (B) Two-tailed Student’s t test. (C and D) One-way ANOVA and Tukey’s post hoc test.