Onocgenic cell marking and single-cell transcriptomics reveal cell type and time-resolved actions of von Hippel Lindau inactivation in the kidney

Defining the first events in oncogenesis and the cellular responses they entrain, even in advance of morphological abnormality, is a fundamental challenge in understanding cancer initiation. As a paradigm to address this we studied the tumor suppressor von Hippel Lindau (VHL), whose loss ultimately drives clear cell renal cancer. Vhl inactivation was directly coupled to activation of a tdTomato reporter within a single allele, allowing accurate visualization of affected cells in their native context and retrieval from the kidney for single-cell RNA sequencing. This strategy uncovered cell-type specific responses to Vhl inactivation, defined a novel proximal tubular class with oncogenic potential, and revealed longer term adaptive changes in the renal epithelium and the interstitium. Oncogenic cell marking also revealed markedly heterogeneous cellular effects including time-limited proliferation and elimination of specific cell types. Striking heterogeneity in these responses suggests that very early events define the tissue specificity of VHL tumor suppressor action. Overall design: Vhl(wt/flox) (Control) and Vhl(KO/flox) (KO) mice carrying the renal tubular epithelium specific Pax8-CreERT2 were dosed with 5x 2 mg tamoxifen and harvested either at an early (1-3 weeks) or late (4-12 months) timepoints after dosing. Cre-mediated recombination marked Vhl recombination with the expression of tdTomato. tdTomato-positve and tdTomato-negative cells were retrieved by FACS and subjected to scRNA-seq using the 10X Chromium NextGEM platform.