FLUORESCENT RECOVERY AFTER PHOTOBLEACHING (FRAP) OF A FLUORESCENT TRANSFERRIN INTERNALIZED IN THE LATE TRANSFERRIN ENDOCYTIC COMPARTMENT OFLIVING A431 CELLS - THEORY

In previous works, other authors characterized a compartment (LCT) of A431 carcinoma cells in which markers of transferrin endocytose had ac cumulated during a long chase period. This compartment, was essentiall y formed by large stationary vacuoles. A few small vesicles budded fro m these vacuoles, rapidly saltated along microtubules and eventually f used with other vacuoles, causing an intracellular transport of the ma rker bound to the limiting membrane (M. De Brabander, R. Nuygens, H. G eerts, C.R. Hopkins, Cell. Mot. Cytoskel. 9 (1988) 30). In the present paper, we derived the fluorescence recovery after photobleaching (FRA P) of a fluorescent marker of LCT. We assumed that the rate of the int racellular transport of the marker was controlled by the fission-fusio n process between vesicles and vacuoles. We showed that the concentrat ion of a bleached fluorescent marker was a decreasing exponential func tion of the time elapsed from the beginning of the recovery phase. The rate constant of this exponential was equal to the product of the ves icle surface by the number of vesicles which fused with a unit of vacu ole surface during one second. If a fraction of the marker spontaneous ly reactivated itself with a much higher rate constant of reaction tha n the rate constant of the transport process, the fractional FRAP of t he marker was the sum of the fractional FRAP of both processes occurri ng separately. In a companion paper (F. Azizi, P. Wahl, Biochim. Bioph ys. Acta 1327 (1997) 75-88), our FRAP experiments will be described an d analysed with the mathematical expressions derived in the present pa per. (C) 1997 Elsevier Science B.V.