Constitutive expression of YFP-ATG8.1 and YFP-ATG8.2 in PCF was
visualised by fluorescent microscopy following RNAi of either
ATG3 or ATG7 for 72 h before
nutrient starvation in PBS for 2.5 h.
(A) Representative images, left hand images FITC filter set,
right hand images DIC FITC merge. Scale bar 5 µm.
(B) The mean number of autophagosomes per cell was
determined by counting >200 cells with data displayed as a mean of three
replicate experiments. Error bars represent standard deviation and
asterisks indicate where data differed significantly from the means of
uninduced controls *p<0.05, ** p<0.01, ***p<0.001.
(C) ATG3 and ATG7 RNAi was
induced for 72 h in cells expressing YFP-ATG8.1 before nutrient
starvation for 3.5 h in PBS (dashed lines) or control treatment in SDM79
media (solid lines). Cells were re-seeded at 0 h in SDM79 + tet (empty
boxes grey line, tet+) and SDM79 -tet (filled boxes black line, tet-)
and growth was determined daily for 96 h. Data represent mean of three
replicate experiments. Error bars show standard deviation and asterisks
indicate where data differed significantly from the means of tet induced
starved and non-starved lines at the 96 h time point ***p<0.001.