Volume 8 issue 6 June 2007
ENLARGEOSOME TRAFFIC: EXOCYTOSIS TRIGGERED BY VARIOUS SIGNALS IS FOLLOWED BY ENDOCYTOSIS, MEMBRANE SHEDDING OR BOTH.
E Cocucci, G Racchetti, P Podini and J Meldolesi

SI 1. [Ca²⁺]_i responses induced in PC12-27 cells by ionomycin (2 mM) and ATP (500 µM). PMA (30 nM) failed to induce any [Ca²⁺]_i change.
Data are expressed in terms of fura-2 340/380 ratio.

Figure 1 (jpg)

SI 2. Effects of TX-100 and trypsin (Tr) on the recovery of the enlargeosome marker, d/A, (top row) and annexin2 (bottom row) in enriched shedding bodies.
Parallel aliquots of shedding bodies, resuspended from filtered pellets, were incubated after no treatment (NT) and after treatment with TX-100 (0.5%), trypsin (Tr, 10 mg/ml, 10 min, 22°) or both, and analyzed without previous re-centrifugation. Notice that TX-100 has no effect on the recovery of both d/A and annexin2, whereas trypsin affects d/A, demonstrating its external orientation in the body membrane. Annexin2, on the other hand, is affected by trypsin only after TX-100 permeabilization of the membrane, as expected for a protein segregated within the body.

Figure 2 (jpg)