Fig. 2

Lentiviral mediated HEYL overexpression in patient-derived colorectal cancer cells. a Morphology of patient-derived colorectal cancer spheroid cultures (M1 and NM1). Phase contrast, scale bar: 200 μm. b Schematic overview of relevant functional sequences of the lentiviral HEYL overexpression and control vectors. CMV, human cytomegaly virus promotor; IRES, internal ribosome entry site; eGFP, enhanced green fluorescent protein; WPRE, woodchuck hepatitis posttranscriptional regulatory element. c The lentivirally encoded gene product GFP is expressed in patient-derived CRC spheroid cultures. Phase contrast and GFP fluorescence, scale bar: 200 μm. d NM1 and M1 cells lack endogenous HEYL mRNA expression. HEYL mRNA is detectable in patient-derived CRC spheroid cultures after transduction with the lentiviral HEYL overexpression vector, but not with the control vector. Relative expression levels were quantified via qRT-PCR as fold-expression calibrated to HEK-293 T control with GAPDH as internal control. Depicted are mean and standard deviation (n = 3). e HEYL protein is weakly expressed in patient-derived CRC spheroid cultures with a 1.7-fold and a 2.1-fold higher optical density for the M1 and NM1 HEYL-transduced cells compared to control, respectively. Western Blot with α-Tubulin used as internal control; HEYL band at ca. 35 kDa (upper band); HEYL-overexpressing adherent HEK-293 T were used as a positive (pos.) control