A Western Blot B Colony Scattering Assay VEC ARTN siCONT siARTN VEC

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A Western Blot B Colony Scattering Assay VEC ARTN siCONT siARTN VEC ARTN siCONT siARTN BT549 ARTN β-ACTIN 28 kDa 42 kDa BT549 MDA-MB-231 ARTN 28 kDa β-ACTIN 42 kDa MDA-MB-231 C Filamentous Actin siCONT siARTN BT549 BT549 MDA-MB-231 VEC siCONT ARTN siARTN VEC ARTN D 3D Matrigel morphology MDA-MB-231 BT549 VEC ARTN siCONT siARTN additional file 2: ARTEMIN (ARTN) modulates the morphology of estrogen receptor (ER) negative mammary carcinoma (ER- MC) cells. (A) Western blot analyses for ARTN in BT549 and MDA-MB-231 cells with forced expression or depletion of ARTN. β-ACTIN was used as loading control for cell lysates. The sizes of detected protein bands in kiloDalton (kDa) are shown on the right.(B) BT549-ARTN and MDA-MB-231-ARTN cells with their respective vector (VEC) cells and BT549-siARTN and MDA-MB-231-siARTN with their respective control (siCONT) cells were seeded at very low cell density (1000 cells plated in 10cm petri dish). Colonies formed by each cell type, were categorised and counted as per the extent of colony scattering(100X magnification). Arrows indicates loose (VEC/siCONT cells), scattered (ARTN cells) and compact (siARTN cells) colonies in BT549 and MDA-MB-231 cells respectively. (C) F-actin was visualised in BT549 and MDA-MB-231 cells with forced expression or depletion of ARTN (400X magnification), Bar, 20µm. (D) Representative photomicrographic pictures of 3D matrigel colonies formed by BT549 and MDA-MB-231 cells with forced expression or depletion of ARTN. Bar, 125μm.