A HETEROTRIMERIC G(I3)-PROTEIN CONTROLS AUTOPHAGIC SEQUESTRATION IN THE HUMAN COLON-CANCER CELL-LINE HT-29

Human colon cancer HT-29 cells exhibit a differentiation dependent autophagic lysosomal pathway that is responsible for the degradation of a pool of newly synthesized N-linked glycoproteins in undifferentiated cells. In the present study, we have investigated the molecular control of this degradative pathway in undifferentiated HT-29 cells. For this purpose, we have modulated the function and expression of the heterotrimeric G-proteins (G(8) and G(i)) in these cells. After pertussis toxin treatment which ADP-ribosylates heterotrimeric G(i)-proteins, we observed an inhibition of autophagic sequestration and the complete restoration of the passage of N-linked glycoproteins through the Golgi complex. In contrast, autophagic sequestration was not reduced by cholera toxin, which acts on heterotrimeric G(s)-proteins. Further insights on the nature of the pertussis toxin-sensitive alpha subunit controlling autophagic sequestration were obtained by cDNA transfections of alpha(i) subunits. Overexpression of the alpha(i3), subunit increased autophagic sequestration and degradation in undifferentiated cells, whereas overexpression of the alpha(i2) subunit, the only other pertussis toxin-sensitive alpha subunit expressed in HT-29 cells, did not alter the rate of autophagy.