Identification of Miranda Protein Domains Regulating Asymmetric Cortical Localization, Cargo Binding, and Cortical Release

• Published in: Molecular and cellular neuroscience 1998-12, Vol.12 (6), p.325-339
• Main Authors: Fuerstenberg, Sal, Peng, Chian-Yu, Alvarez-Ortiz, Pedro, Hor, Tiffany, Doe, Chris Q.
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Binding Sites - physiology; Brain Chemistry; Cell Cycle Proteins - chemistry; Cell Cycle Proteins - genetics; Cell Cycle Proteins - metabolism; Drosophila; Drosophila Proteins; Epithelial Cells - chemistry; Epithelial Cells - cytology; Ganglia, Invertebrate - chemistry; Ganglia, Invertebrate - cytology; Gene Expression - physiology; Insect Proteins - analysis; Insect Proteins - chemistry; Interphase - physiology; Molecular Sequence Data; Mutation; Nerve Tissue Proteins - chemistry; Nerve Tissue Proteins - genetics; Nerve Tissue Proteins - metabolism; Neurons - chemistry; Neurons - cytology; Nuclear Proteins - chemistry; Nuclear Proteins - genetics; Nuclear Proteins - metabolism; Protein Structure, Secondary; Protein Structure, Tertiary; RNA-Binding Proteins - chemistry; RNA-Binding Proteins - genetics; RNA-Binding Proteins - metabolism; Stem Cells - chemistry; Stem Cells - cytology; Transcription Factors - chemistry; Transcription Factors - genetics; Transcription Factors - metabolism
• ISSN: 1044-7431; 1095-9327
• DOI: 10.1006/mcne.1998.0724

An important question in cellular and developmental biology is how a cell divides to produce daughter cells with different fates.Drosophilaneuroblasts are a model system for studying asymmetric cell division: at each division, neuroblasts retain stem cell-like features, whereas their sibling ganglion mother cell (GMC) has a more restricted fate. Establishing neuroblast/GMC differences involves the asymmetric localization of proteins (Inscuteable, Miranda, Prospero, and Staufen) and RNA (prospero). All of these factors are apically localized during interphase, and all except Inscuteable move to the basal cortex at mitosis prior to being partitioned solely into the GMC. In this study, we show that Miranda is colocalized with Staufen and Prospero in neuroblasts, and is required for the asymmetric cortical localization of both proteins. Analysis ofmirandamutants reveals three functional domains within the Miranda protein: (1) an N-terminal domain (1–290 aa) sufficient for association of Miranda with the cell cortex and basal localization in mitotic neuroblasts; (2) a central domain (446–727 aa) necessary for apical localization in interphase neuroblasts as well as for “cargo binding” of Prospero, Staufen, andprosperomRNA; and (3) a C-terminal domain (727–830 aa) necessary for the timely degradation of Miranda and release of its cargo from the cortex of the newborn GMC. In addition, Miranda is asymmetrically localized in epithelial cells that lack Inscuteable and divide symmetrically; thus the mechanism regulating Miranda localization is common to epithelial cells and neuroblasts, and Inscuteable is not an obligate component. Finally, we define a C-terminal domain of Staufen sufficient for Miranda-dependent cortical localization in neuroblasts.