Investigating the Role of Vasodilator Stimulated Phosphoprotein in Protein Scavenging of Pancreatic Ductal Adenocarcinoma

Abstract

Pancreatic ductal adenocarcinoma (PDAC) is a lethal malignant disease with extremely poor prognosis and limited therapeutic options. Recent studies have shown that constitutive KRAS signaling which drives most of PDAC induces a process called protein scavenging. During this process, cancer cells take up extracellular proteins such as serum albumin by macropinocytosis, and subsequent degradation in lysosome yields monomeric amino acids. Although protein scavenging has emerged as an important mechanism to secure a carbon source in nutrient-poor tumor microenvironment, detailed understanding of mechanism and genes involved is lacking. A genome wide CRISPR- Cas9 screen on a murine derived KRPC cell line identified VASP as one of the essential genes for protein scavenging, and genetic knockout of VASP protein using CRISPR decreased rate of protein scavenging measured by a quantitative flux analysis. To further confirm the role of VASP in protein scavenging, we generated rescue cell lines by expressing wild type(WT) VASP as well as phosphorylation-defective Y39F VASP in VASP KO cell lines. We show that VASP re-expression rescues growth of KRPCA cells by increasing protein scavenging selectively in leucine deplete medium supplemented with bovine serum albumin in which cells depend largely on scavenging for proliferation. In addition, while Y39F VASP also rescued growth, the effect size was lower than that of WT VASP, indicating importance of phosphorylation at tyrosine 39 for proper function of VASP. Findings in this study are robust follow-up of the CRISPR screen, and they demonstrate the importance of VASP in regulating protein scavenging in PDAC.