Neuronal cell fate decisions are directed in Drosophila by NUMB, a signaling adapter protein with two protein-protein interaction domains: a phosphotyrosinebinding domain and a proline-rich region (PRR) that functions as an SH3-binding domain. Here we show that there are at least four human NUMB isoforms and that these serve two distinct developmental functions in the neuronal lineage: differentiation (but not proliferation) is promoted by human NUMB protein isoforms with a type I (short) PRR. In contrast, proliferation (but not differentiation) is directed by isoforms that have a type II (long) PRR. The two types of PRR may promote distinct intracellular signaling pathways downstream of the NOTCH receptor during mammalian neurogenesis.During Drosophila peripheral nervous system development, a sensory organ precursor cell divides twice to produce the four cells that form a functional sensory organ (neuron, sheath, hair, and socket). NUMB functions in this lineage to direct specific binary cell fate choices: the IIb vs. IIa fate at the first division and at the second division, neuron vs. sheath as well as hair vs. socket (1). Absence of NUMB results in production of two IIa cells that then divide to give four sockets while ectopic expression of NUMB generates two IIb cells and, subsequently, four neurons and no hairs (2). The fate choices directed by Drosophila NUMB occur in a stereotypical lineage within which NUMB functions solely to control binary differentiation decisions and not the proliferation dynamics of the cells. NUMB has been hypothesized to function in directing cell fate choice by directly interacting with NOTCH and inhibiting NOTCH function (3).The recent cloning of a mammalian NUMB homologue suggested an evolutionarily conserved function for mammalian NUMB (4, 5). We showed that ectopic expression of this mammalian NUMB protein (mNUMB) promotes neuronal commitment in both Drosophila and cultured mammalian cells (4). Recently, it has been observed that a human NUMB homologue (hNUMB) is transported to the nucleus and associates with the modulator of mitogenesis, MDM2 (6, 7). Our previous studies showed no mitogenic component to murine NUMB function (4). Here we report the existence of four hNUMB isoforms. Based on the structure of the proline-rich region (PRR), the NUMB isoforms regulate either cell fate or cellular proliferation, but not both, during mammalian neurogenesis.

We compared overexpression of the magnetotactic bacterial gene MagA with the modified mammalian ferritin genes HF + LF, in which both heavy and light subunits lack iron response elements. Whereas both expression systems have been proposed for use in non-invasive, magnetic resonance (MR) reporter gene expression, limited information is available regarding their relative potential for providing gene-based contrast. Measurements of MR relaxation rates in these expression systems are important for optimizing cell detection and specificity, for developing quantification methods, and for refinement of gene-based iron contrast using magnetosome associated genes. We measured the total transverse relaxation rate (R2*), its irreversible and reversible components (R2 and R2′, respectively) and the longitudinal relaxation rate (R1) in MDA-MB-435 tumor cells. Clonal lines overexpressing MagA and HF + LF were cultured in the presence and absence of iron supplementation, and mounted in a spherical phantom for relaxation mapping at 3 Tesla. In addition to MR measures, cellular changes in iron and zinc were evaluated by inductively coupled plasma mass spectrometry, in ATP by luciferase bioluminescence and in transferrin receptor by Western blot. Only transverse relaxation rates were significantly higher in iron-supplemented, MagA- and HF + LF-expressing cells compared to non-supplemented cells and the parental control. R2* provided the greatest absolute difference and R2′ showed the greatest relative difference, consistent with the notion that R2′ may be a more specific indicator of iron-based contrast than R2, as observed in brain tissue. Iron supplementation of MagA- and HF + LF-expressing cells increased the iron/zinc ratio approximately 20-fold, while transferrin receptor expression decreased approximately 10-fold. Level of ATP was similar across all cell types and culture conditions. These results highlight the potential of magnetotactic bacterial gene expression for improving MR contrast.