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 aut ophagic 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 contro l of this degradative pathway in undifferentiated HT-29 cells. For thi s purpose, we have modulated the function and expression of the hetero trimeric G-proteins (G(8) and G(i)) in these cells. After pertussis to xin treatment which ADP-ribosylates heterotrimeric G(i)-proteins, we o bserved an inhibition of autophagic sequestration and the complete res toration of the passage of N-linked glycoproteins through the Golgi co mplex. In contrast, autophagic sequestration was not reduced by choler a toxin, which acts on heterotrimeric G(s)-proteins. Further insights on the nature of the pertussis toxin-sensitive alpha subunit controlli ng autophagic sequestration were obtained by cDNA transfections of alp ha(i) subunits. Overexpression of the alpha(i3), subunit increased aut ophagic sequestration and degradation in undifferentiated cells, where as overexpression of the alpha(i2) subunit, the only other pertussis t oxin-sensitive alpha subunit expressed in HT-29 cells, did not alter t he rate of autophagy.