Abstract. Basic substances and acidic ionophores that increase the lysosomal pH in cultured macrophages (Ohkuma, S., and B. Poole, 1978, Proc. NatL Acad. Sci. USA., 75:3327-3331; Poole, B., and S. Ohkuma, 1981, d. Cell Biol., 90:665-669) inhibited the digestion of heat-denatured acetylated bovine serum albumin (BSA) taken up by the cells. For several substances, the shift in pH sufficed to explain the inhibition of proteolysis. Additional effects, presumably on enzyme activities, have to be postulated for tributylamine, amantadine, and chloroquine. Sodium fluoride (10 mM) had no significant effect on the breakdown of BSA by macrophages.The breakdown of endogenous macrophage proteins, whether short lived or long lived, was inhibited ~40% by 10 mM NaF and 30%, or sometimes less in the case of long-lived proteins, by 100 #M chloroquine. When the cells were supplied with BSA, a mixture of cell proteins, or even inert endocytosible materims, the breakdown of endogenous long-lived proteins and the inhibitory effect of chloroquine on this process were selectively reduced. Inhibition of endocytosis by cytochalasins B or D did not affect the chloroquine-sensitive breakdown of endogenous proteins, indicating that the proteins degraded by this process were truly endogenous and not taken in from the outside by cellular cannibalism. On the other hand, when macrophage proteins were supplied extracellulady, their breakdown occurred at the same rate for short-lived and long-lived proteins, and it was strongly inhibited by chloroquine and not by NaF.It is concluded from these results that the breakdown of endogenous proteins, both short-lived and long-lived, probably takes place partly (~30%) in lysosomes and partly through one or more nonlysosomal mechanism(s) unaffected by chloroquine and presumably susceptible to inhibition by fluoride. A difference must exist between short-lived and long-lived proteins in the manner in which they reach lysosomes or are handled by these organelles; this difference would account for the selective effect of the supply of endocytosible materials on the lysosomal processing of longlived proteins.
