ASH2L Controls Ureteric Bud Morphogenesis through the Regulation of RET/GFRA1 Signaling Activity in a Mouse Model

Abstract

Significance statement: Causes of congenital anomalies of the kidney and urinary tract (CAKUT) remain unclear. The authors investigated whether and how inactivation of Ash2l -which encodes a subunit of the COMPASS methyltransferase responsible for genome-wide histone H3 lysine K4 (H3K4) methylation-might contribute to CAKUT. In a mouse model, inactivation of Ash2l in the ureteric bud (UB) lineage led to CAKUT-like phenotypes. Removal of ASH2L led to deficient H3K4 trimethylation, which slowed cell proliferation at the UB tip, delaying budding and impairing branching morphogenesis. The absence of ASH2L also downregulated the expression of Ret , Gfra1 , and Wnt11 genes involved in RET/GFRA1 signaling. These findings identify ASH2L-mediated H3K4 methylation as an upstream epigenetic regulator of signaling crucial for UB morphogenesis and indicate that deficiency or dysregulation of these processes may lead to CAKUT.

Background: Ureteric bud (UB) induction and branching morphogenesis are fundamental to the establishment of the renal architecture and are key determinants of nephron number. Defective UB morphogenesis could give rise to a spectrum of malformations associated with congenital anomalies of the kidney and urinary tract (CAKUT). Signaling involving glial cell line-derived neurotrophic factor and its receptor rearranged during transfection (RET) and coreceptor GFRA1 seems to be particularly important in UB development. Recent epigenome profiling studies have uncovered dynamic changes of histone H3 lysine K4 (H3K4) methylation during metanephros development, and dysregulated H3K4 methylation has been associated with a syndromic human CAKUT.

Methods: To investigate whether and how inactivation of Ash2l , which encodes a subunit of the COMPASS methyltransferase responsible for genome-wide H3K4 methylation, might contribute to CAKUT, we inactivated Ash2l specifically from the UB lineage in C57BL/6 mice and examined the effects on genome-wide H3K4 methylation and metanephros development. Genes and epigenome changes potentially involved in these effects were screened using RNA-seq combined with Cleavage Under Targets and Tagmentation sequencing.

Results: UB-specific inactivation of Ash2l caused CAKUT-like phenotypes mainly involving renal dysplasia at birth, which were associated with deficient H3K4 trimethylation. Ash2l inactivation slowed proliferation of cells at the UB tip, delaying budding and impairing UB branching morphogenesis. These effects were associated with downregulation of Ret , Gfra1 , and Wnt11 , which participate in RET/GFRA1 signaling.

Conclusions: These experiments identify ASH2L-dependent H3K4 methylation in the UB lineage as an upstream epigenetic regulator of RET/GFRA1 signaling in UB morphogenesis, which, if deficient, may lead to CAKUT.