Supplementary Information from Methylation of HSP70 Orchestrates Its Binding to and Stabilization of BCL2 Mrna and Renders Pancreatic Cancer Cells Resistant to Therapeutics

Supplementary Figures and Tables and Methods 8 Supplemental Figures Figure S1. PRMT1 is overexpressed in multiple gastrointestinal tumors and is associated with patient overall survival. Figure S2. Genetic alterations and inflammatory factors contribute to increased PRMT1 expression during pancreatic cancer (PDA) progression. Figure S3. Altered expression and activity of PRMT1 impacts pancreatic cancer (PDA) cell migration, invasion, and growth. Figure S4. PRMT1 and HSP70 proteins exhibit subcellular co-localization and binding motifs of PRMT1-HSP70 interaction are evident. Figure S5. Arginine methylation sites of HSP70 are identified. Figure S6. Arginine methylation of HSP70 protects cells against various stresses. Figure S7. Arginine methylation of HSP70 promotes its mRNA binding and stabilization activity. Figure S8. Arginine methylation of heat shock proteins mediates therapeutic resistance in pancreatic cancer. 3 Supplemental Tables Table S1. Clinicopathologic parameters and PRMT1 expression in the pancreatic cancer patients Table S2. HA-PRMT1 and HA-ctrl binding proteins identified in mass spectrometry Table S3. Primers and peptides used in the study. Detailed Supplemental Methods for Cell Lines and Cell Culture; Plasmids and Cloning Procedures; Antibodies, siRNAs, Primers and Compounds; shRNA construct and transfection; Western blot Analysis; Alcian Blue Staining; Cell Scratch-Wound Healing Assay; Cell Invasion Assay; In Vitro Cytotoxicity and Cell Viability Assay; Immunoprecipitation Assay; Protein Purification from Bacterial or Mammalian Cells; Immunofluorescence Assay; In Vitro Methylation Assay; Protein Sequence Alignment; Mass Spectrometry Analysis; Generation of Gene Knockout Cell Lines; Annexin V-Propidium Iodide Assay; Quantitative Real-time PCR Assay; Ribonucleoprotein Immunoprecipitation; mRNA Decay Assay; Protein-Nucleic Acid Binding Assay; and Luciferase Assay.