Fig. 5

Autophagy induction in OVCAR8-eGFP-LC3B cells is unaffected by Beclin-1 knockdown. OVCAR8-eGFP-LC3 cells were transfected with control siRNA (siNT) or siRNA targeting BECN1, ATG5, ATG7, or ATG5 + 7 and seeded to adherent or non-adherent culture. a Adherent cells were allowed to attach overnight and treated with DMSO or Akti-1/2 (5 μM) the next day. Images were captured 24 h post-treatment using an Olympus IX70 inverted microscope and ImagePro software. A portion of the siBECN1 image was expanded to clearly show an abundance of fluorescent puncta around the nucleus (n). Scale bar: 50 μm. b Transfected cells seeded to non-adherent culture were harvested 24 h following seeding, protein lysates generated, and immunoblot performed for indicated proteins. An anti-GFP antibody was used to detect fused and free eGFP protein. Images and immunoblot are representative of duplicate experiments