Programmed Synthesis of Tubulin for the Flagella that Develop During Cell Differentiation in Naegleria gruberi

Amebae of Naegleria gruberi differentiate into flagellates when transferred from growth medium to nonnutrient buffer. Experiments were performed to determine whether the tubulin that forms the flagellar microtubules pre-exists in amebae or is synthesized during differentiation. Amebae prelabeled uniformly with [(35)S]methionine were allowed to differentiate in the presence and in the absence of exogenous unlabeled methionine. In the presence of unlabeled methionine the flagellar tubulin contained only 30% as much [(35)S]methionine as in its absence. Thus at least 70% of the tubulin was synthesized de novo. Isotope dilution and pulse experiments showed that flagellar tubulin synthesis began one-third of the way through differentiation, before any morphological change had occurred. Flagellar tubulin antigen, as measured using a specific antiserum, also began to increase one-third of the way through differentiation and increased 35- to 55-fold during the course of differentiation. These experiments demonstrate that most if not all of the flagellar tubulin is synthesized de novo during differentiation.