During asymmetric cell division, protein determinants are segregated into one of the two daughter cells. The Numb protein acts as a segregating determinant during both mouse and Drosophila development. In flies, Numb localizes asymmetrically and is required for cell-fate specification in the central and peripheral nervous systems, as well as during muscle and heart development. Whether its asymmetric segregation is important to the performance of these functions is not firmly established. Here, we demonstrate that Numb acts both in a localization-dependent and in a localization-independent manner. We have generated numb mutants that affect only the asymmetric localization of the protein during mitosis. We demonstrate that asymmetric segregation of Numb into one of the two daughter cells is absolutely essential for cell-fate specification in the Drosophila peripheral nervous system. Numb localization is also essential in MP2 neuroblasts in the central nervous system and during muscle development. Surprisingly, in dividing ganglion mother cells or during heart development, Numb function is independent of its ability to segregate asymmetrically in mitosis. Our results suggest that two classes of asymmetric cell division exist, each with different requirements for asymmetric inheritance of cell-fate determinants.