Fig. 3

Identification of gene expression changes upon miR-204 downregulation in the non-tumorigenic HaCaT cell line. HaCaT cells infected with anti-miR-204 lentivirus (shRNA targeting miR-204) or with the corresponding pGreenPuro scrambled hairpin negative control lentiviral supernatant were flow cytometry-sorted and their total RNAs were purified. Differences in gene expression between both conditions were analyzed by the Human Gene ST Arrays platform (Affymetrix). a Ingenuity Pathway Analysis (IPA) identification of the molecular and cellular functions, top canonical pathways and upstream regulators. b In silico identification by IPA of putative direct miR-204 targets differentially regulated in HaCaT cells. c MiR-204 expression relative to RNU48 measured by qRT-PCR in HaCaT cells transfected with miR-204 antagomir or miR-204 mimic along with their appropriate controls. d PTPN11 expression is up-regulated by miR-204 in HaCaT cells, monitored by Western blot. e Relative Renilla luciferase activity derived from the PTPN11 3′ UTR reporter construct monitored after transfection of HaCaT cells with miR-204 antagomir or miR-204 mimic, along with their controls. f Independent validation of miR-204 regulation of some of the genes identified by the expression array, by HaCaT transient transfection of miR-204 antagomir or mimic along with their controls. Mean ± SD of three replicate samples from one representative experiment