Research has previously shown that, without methionine supplements, neural tube proteins of rat embryos cultured on bovine sera were hypomethylated and neural tubes failed to close. In the present study, to identify the proteins that became methylated during neurulation, rat embryos were first cultured on methionine-deficient bovine serum for 40 hr, then incubated with puromycin for 1 hr, and, finally, incubated with [methyl-14 C]methionine and puromycin for 5 hr. On the basis of molecular weights, isoelectric points, and Western immunoblots, the methyl-14 C-labeled proteins were identified as actin, ␣␤-tubulin, and neurofilament L. Indirect immunof luorescence studies indicated that without the addition of methionine to the culture, localization of actin and ␣␤-tubulin in the basal cytoplasm did not occur and these neuroepithelial cells lost their columnar morphology.