Total RNA was extracted using an RNeasy mini kit (Qiagen, CA). RNA-Seq: The Akt transformed prostate stem cell line was infected with shKlf4 (shKlf4) or non-silencing control (shCon) lentivirus. After 48 hr cells were selected by puromycin for 2 days and passaged for 10 days to allow cells to undergo EMT. The infected shCon and shKlf4 cells were collected for RNA extraction. Another cohort of shKlf4 cells was infected with BFP tagged Doxycycline inducible human KLF4 (shKlf4+KLF4) or BFP tagged Doxycycline inducible empty vector (shKlf4+EV) lentivirus. After 3 days, 100 ng/ml of Dox was added to cultures to induce Klf4 expression while control cultures received no Dox. Two days later, all shKlf4+KLF4 and shKlf4+EV cells with or without Dox treatment were collected for RNA extraction. Total RNA was extracted using an RNeasy mini kit (Qiagen, CA). RNA-seq libraries were prepared with the TruSeq sample preparation kit (Illumina, CA). ChIP-Seq: For ChIP-seq analyses, 1x107 cells of the Akt transformed prostate stem cell line with endogenous Klf4 were fixed in PBS containing 1% of formaldehyde for 8 min at room temperature on a rotating platform. The reaction was quenched with 0.125M glycine for 5 min. After washing twice with cold PBS, cells were collected and the chromatin prepared according to the ChIP-IT High Sensitivity protocol (Active Motif, CA). Cell pellets were re-suspended in 10 ml of chromatin prep buffer supplemented with a proteinase inhibitor cocktail (PIC) and PMSF (100 µM) and nuclei were extracted using a chilled Dounce homogenizer. Nuclei were re-suspended in ChIP buffer supplemented with PIC and PMSF (1 mM) and sonicated using a Diagenode Bioruptor 300 at maximum intensity and pulsed for 30 sec on and 30 sec off for 30 cycles. Prior to ChIP, chromatin size was confirmed by reverse-crosslinking of 2.5% of the input by incubation with 10 µg of RNAse for 30min at 37ºC followed by addition of 20 µg proteinase K and incubation for 30 min at 55ºC and 2 hours at 80ºC, making sure we had achieved an average length between 200-1200. Immunoprecipitation was performed overnight at 4C on 30 μg of sheared chromatin using 4 μg of KLF4 antibody. Chromatin samples were washed 5 times before reversing crosslinks. DNA was then extracted using in a DNA purification column (Active Motif, CA) and eluted in 37 µl of DNA Purification Elution Buffer (Active Motif, CA) for ChIP-seq analysis. Sample quality and DNA concentration was assessed by Qubit (Invitrogen, CA). ChIP-Seq libraries were made using the ThruPLEX DNA-seq kit (Rubicon Genomics, MI) using 200 pg of ChIP DNA or KAPA Hyper Prep kit (Kapa Biosystems, MA) using 1 ng of ChIP DNA. Libraries were size selected prior to PCR amplification using AMPure XP beads (Beckman Coulter, CA). Multiplexed libraries were run on an Illumina HiSeq 2500 instrument, Genome Analyzer using the 50-base pair single read method.