pgen.1009094.g008.tif (739.71 kB)

In vitro functional validation of chromatin remodelers in immortalized keratinocytes.

figure

posted on 2021-08-16, 17:39 authored by Aziz Aiderus, Justin Y. Newberg, Liliana Guzman-Rojas, Ana M. Contreras-Sandoval, Amanda L. Meshey, Devin J. Jones, Felipe Amaya-Manzanares, Roberto Rangel, Jerrold M. Ward, Song-Choon Lee, Kenneth Hon-Kim Ban, Keith Rogers, Susan M. Rogers, Luxmanan Selvanesan, Leslie A. McNoe, Neal G. Copeland, Nancy A. Jenkins, Kenneth Y. Tsai, Michael A. Black, Karen M. Mann, Michael B. Mann

Assessment of keratinocyte transformation in response to stable shRNA-mediated knockdown of CREBBP, KMT2C and NCOA2 in HaCaT cells (A) confirmed by Taq-Man showed cellular proliferation changes (B) with CREBBP knockdown relative to HaCaT cells stably transduced with a non-targeting shRNA (shNTC). Statistical significance was tested by one-way ANOVA (P = 0.0024) followed by Sidak’s multiple comparisons adjustment (shNTC vs. shCREBBP, P = 0.0022; shNTC vs. shKMT2C, P = 0.19; shNTC vs. shNCOA2, P = 0.99; n = 6 per group). Colony formation assays in soft agar (C) showed significant differences in the number and size of colonies from HaCaT cells with target gene knockdown compared to the non-targeting control for (D) CREBBP knockdown vs. shNTC, P = 0.018; (E) KMT2C knockdown vs. shNTC, P = 0.039; (F) NCOA2 knockdown vs. shNTC, P = 0.31. Colonies were stained with crystal violet solution and quantitated; statistical significance was tested using an unpaired t-test; n = 3 per group.