Conditions have been devised that permit study of the entry of Bordetella pertussis extracellular adenylate cyclase into a variety of host cells in culture. It is important to guard against errors deriving from the extracellular production of cAMP formed from secreted ATP. Extracellularly generated cAMP appears not to enter the cells although transport in the opposite direction can occur in some cell lines. Purified enzyme does not enter cells but crude preparations show rapid, dose-dependent huge accumulations of intracellular cAMP not originating from the outside but ascribable to cyclase that has crossed the plasma membrane. This entry appears not to occur by classical endocytosis as judged by inhibitor studies with chloroquin, dansyl cadaverine, monensin ammonium chloride or methyl amine chloride. These agents are effective protectors against other toxins such as diptheria toxin. Adenylate cyclase may require the participation of microtubules (rather than microfilaments) and appears to be quite different from the entry of other bacterial toxins. The data suggest the existence of a helper factor that is currently being identified.
