Skip to main content

Advertisement

Fig. 4 | BMC Cancer

Fig. 4

From: A germline mutation in Rab43 gene identified from a cancer family predisposes to a hereditary liver-colon cancer syndrome

Fig. 4

Loss of Rab43 retards cell growth and proliferation through impairment of Akt activation (a) Generation of Rab43 knockout HepG2 cell line by Crispr-Cas9 gene editing, a Rab43 knockout cell line was established and verified by Sanger DNA sequencing with three different reading frame shift mutations as shown in (a). The red arrow indicates the location of a deoxycytidine triphosphate insertion. b Western blot confirmed Rab43 protein is completely lost in the Crispr-Rab43 HepG2 cells. c-d Rab43 knockout HepG2 cells display an enlarged and flattened phenotype with slow growth, some cells exhibit multiple nuclei as indicated by a black arrow. The slow growth of Rab43 knockout cells could be reversed by over-expressing a Crispr-resistant Rab43 gene. e-g Activation of Akt and Akt signaling is impaired in Rab43 knockout HepG2 cells. Basal level of Akt activation was barely detectable in Crispr-Rab43 cells without insulin stimulation by Western blotting in (e). The indicated cells were serum-starved overnight and re-stimulated with 100 nM insulin for 30 min. The expression levels of Akt-S473, Akt and phos-p70S6K were detected by Western blot in (f), quantified by densitometry scanning, normalized to GAPDH signal and plotted in (g). Quantification data were based on three independent experiments. The activation differences of Akt and phos-p70S6K between V2 control and Crispr-Rab43 cells post insulin stimulation were statistically significant (*p < 0.05 by Student’s t test)

Back to article page