Biology subjects

Susztak, K.

Publications and source records attributed to Susztak, K..

4 recordsLinked to original sources

bioRxiv2018

DNMT1 in Six2 progenitor cells is essential for transposable element silencing and kidney development

Cytosine methylation (5mC) plays a key role in maintaining progenitor cell self-renewal and differentiation. Here, we analyzed the role of 5mC in kidney development by genome-wide methylation, expression profiling, and by systematic genetic targeting of DNA methyltransferases (Dnmt) and Ten-eleven translocation methylcytosine hydroxylases (Tet).\n\nIn mice, nephrons differentiate from Six2+ progenitor cells, therefore we created animals with genetic deletion of Dnmt1, 3a, 3b, Tet1, and Tet2 in the Six2+ population (Six2Cre/Dnmt1flox/flox, Six2Cre/Dnmt3aflox/flox, Six2Cre/Dnmt3bflox/flox, Six2Cre/Tet2flox/flox and Tet1-/-). Animals with conditional deletion of Dnmt3a, 3b, Tet1 and Tet2 showed no significant structural or functional renal abnormalities. On the other hand, Six2Cre/Dnmt1flox/flox mice died within 24hrs of birth. Dnmt1 knock-out animals had small kidneys and significantly reduced nephron number. Genome-wide methylation analysis indicated marked loss of methylation mostly on transposable elements. RNA sequencing detected endogenous retroviral (ERV) gene transcripts and early embryonic genes. Increase in levels of interferon (and RIG-I signaling) and apoptosis (Trp53) in response to ERV activity likely contributed to the phenotype development. Once epithelial differentiation was established, loss of Dnmt1, 3a, 3b, Tet1 or Tet2 in glomerular epithelial cells did not lead to functional or structural differences at baseline or following toxic glomerular injury.\n\nGenome-wide cytosine methylation and gene expression profiling showed that Dnmt1-mediated DNA methylation is essential for kidney development by preventing regression of progenitor cells into a primitive undifferentiated state and demethylation of transposable elements.\n\nSignificanceCytosine methylation of regulatory regions (promoters and enhancers) has been proposed to play a key role in establishing gene expression and thereby cellular phenotype. DNMT1 is the key enzyme responsible for maintaining methylation patterns during DNA replication. While the role of Dnmt1 has been described in multiple organs, here we identified a novel, critically important mechanism how Dnmt1 controls tissue progenitors. The greatest methylation difference in Dnmt1 knock-out mice was observed on transposable elements (TE), which resulted in increase of endogenous retroviruses and cell death. We believe that release of TE was a critically overlooked component of phenotype development in previous studies that our comprehensive genome wide methylation analysis allowed us to identify.\n\nCompeting interestsThe Susztak lab receives research support from Biogen, Boehringer Ingelheim, Celgene, GSK, Merck, Regeneron and ONO Pharma for work not related to this manuscript.

developmental biology↗

bioRxiv2018

Bulk Tissue Cell Type Deconvolution with Multi-Subject Single-Cell Expression Reference

We present MuSiC, a method that utilizes cell-type specific gene expression from single-cell RNA sequencing (RNA-seq) data to characterize cell type compositions from bulk RNA-seq data in complex tissues. When applied to pancreatic islet and whole kidney expression data in human, mouse, and rats, MuSiC outperformed existing methods, especially for tissues with closely related cell types. MuSiC enables characterization of cellular heterogeneity of complex tissues for identification of disease mechanisms.

bioinformatics↗

bioRxiv2018

Jagged1/Notch2 Controls Kidney Fibrosis via Tfam-mediated Metabolic Reprogramming

While Notch signaling has been proposed to play a key role in fibrosis, the direct molecular pathways targeted by Notch signaling and the precise ligand and receptor pair that are responsible for kidney disease remain poorly defined.\n\nIn this study, we found that JAG1 and NOTCH2 showed the strongest correlation with the degree of interstitial fibrosis in a genome wide expression analysis of a large cohort of human kidney samples. RNA sequencing analysis of kidneys of mice with folic acid nephropathy, unilateral ureteral obstruction, or APOL1-associated kidney disease indicated that Jag1 and Notch2 levels were higher in all analyzed kidney fibrosis models. Mice with tubule-specific deletion of Jag1 or Notch2 (Kspcre/Jag1flox/flox, and Kspcre/Notch2flox/flox) had no kidney-specific alterations at baseline, but showed protection from folic acid induced kidney fibrosis. Tubule-specific genetic deletion of Notch1 and global knock-out of Notch3 had no effect on fibrosis. In vitro chromatin immunoprecipitation experiments and genome-wide expression studies identified the mitochondrial transcription factor A (Tfam) as a direct Notch target. Re-expression of Tfam in tubule cells prevented Notch-induced metabolic and profibrotic reprogramming. Kidney tubule specific deletion of Tfam resulted in perinatal lethality.\n\nIn summary, Jag1/Notch2 plays a key role in kidney fibrosis development by regulating Tfam expression and metabolic reprogramming.

pathology↗

bioRxiv2017

Comprehensive single cell RNAseq analysis of the kidney reveals novel cell types and unexpected cell plasticity

A key limitations to understand kidney function and disease development has been that specific cell types responsible for specific homeostatic kidney function or disease phenotypes have not been defined at the molecular level.\n\nTo fill this gap, we characterized 57,979 cells from healthy mouse kidneys using unbiased single-cell RNA sequencing. We show that genetic mutations that present with similar phenotypes mostly affect genes that are expressed in a single unique differentiated cell type. On the other hand, we found unexpected cell plasticity of epithelial cells in the final segment of the kidney (collecting duct) that is responsible for final composition of the urine. Using computational cell trajectory analysis and in vivo linage tracing, we found that, intercalated cells (that secrete protons) and principal cells (that maintain salt, water and potassium balance) undergo a Notch mediated interconversion via a newly identified transitional cell type. In disease states this transition is shifted towards the principal cell fate. Loss of intercalated cells likely contributes to metabolic acidosis observed in kidney disease.\n\nIn summary, single cell analysis advanced a mechanistic description of kidney diseases by identifying a defective homeostatic cell lineage.\n\nOne Sentence SummaryA comprehensive single cell atlas of the kidney reveals a transitional cell type and cell plasticity determined by Notch signaling which is defective in chronic kidney disease.

cell biology↗