A Novel Hook-Related Protein Family and the Characterization of Hook-Related Protein 1

Institute for Molecular Bioscience, University of Queensland, Brisbane, QLD 4072, Australia.
Traffic (Impact Factor: 4.35). 07/2005; 6(6):442-58. DOI: 10.1111/j.1600-0854.2005.00289.x
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The spatial organization of organelles within a cell is dependent on microtubules. Recently, members of the Hook family of proteins have been proposed to function in linking organelles to microtubules. We report the identification of a completely novel protein family, the Hook-related protein (HkRP) family, from which the Hook proteins have diverged. Bioinformatic analysis of the HkRP family revealed several conserved domains, including a unique C-terminal HkRP domain. The central region of each protein is comprised of an extensive coiled-coil domain, and the N-terminus contains a putative microtubule-binding domain. This domain has been shown to bind microtubules in the Hook protein and show that the HkRP1 protein is microtubule-associated. While endogenous HkRP1 has no distinct organelle association, expression of the C-terminal membrane-binding domain suggests a function of the HkRP1 in early endosome. Ultrastructural studies reveal that expression of the C-terminal HkRP1 domain causes an accumulation of internal membranes with an electron-dense coat. Co-localization studies show a concomitant redistribution of the early endosome marker sorting-nexin 1 but not the early endosome antigen-1 (EEA1). The steady-state distribution of the epidermal growth factor receptor is also specifically disrupted by expression of the C-terminal domain. We propose that HkRP1 is involved in the process of tubulation of sorting nexin-1 positive membranes from early endosome subdomains.

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    • "Since in both mecA 101 and mecA 20 mutations were introduced in a region that is common to all splice variants, both mecA alleles are expected to affect all Girdin isoforms. Both human and Drosophila Girdin have been reported to have N termini that are similar to the microtubule-binding domain of the Hook protein, an extensive central coiled-coil domain that is conserved in family-HkRP and more diverse C termini that specify the binding partners, such as actin (Simpson et al. 2005; Enomoto et al. 2006; Puseenam et al. 2009). "
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    ABSTRACT: The dendrite of the sensory neuron is surrounded by support cells and is composed of two specialized compartments: the inner segment and the sensory cilium. How the sensory dendrite is formed and maintained is not well understood. Hook-related proteins (HkRP) like Girdin, DAPLE, and Gipie are actin-binding proteins, implicated in actin organization and in cell motility. Here, we show that the Drosophila melanogaster single member of the Hook-related protein family, Girdin, is essential for sensory dendrite formation and function. Mutations in girdin were identified during a screen for fly mutants with no mechanosensory function. Physiological, morphological, and ultra-structural studies of girdin mutant flies indicate that the mechanosensory neurons innervating external sensory organs (bristles) initially form a ciliated dendrite that degenerates shortly after, followed by the clustering of their cell bodies. Importantly, we observed that Girdin is expressed transiently during dendrite morphogenesis in three previously unidentified actin-based structures surrounding the inner segment tip and the sensory cilium. These actin structures are largely missing in girdin. Defects in cilia are observed in other sensory organs such as those mediating olfaction and taste, suggesting that Girdin has a general role in forming sensory dendrites in Drosophila. These suggest that Girdin functions temporarily within the sensory organ and that this function is essential for the formation of the sensory dendrites via actin structures. Copyright © 2015, The Genetics Society of America.
    Genetics 06/2015; DOI:10.1534/genetics.115.178954 · 5.96 Impact Factor
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    • "Previous study indicated girdin as the component of dynamin complex (Simpson et al, 2005). To confirm this finding, the endogenous interaction between girdin and dynamin was investigated by co-immunoprecipitation (Co-IP). "
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    ABSTRACT: In clathrin-mediated endocytosis (CME), specificity and selectivity for cargoes are thought to be tightly regulated by cargo-specific adaptors for distinct cellular functions. Here, we show that the actin-binding protein girdin is a regulator of cargo-selective CME. Girdin interacts with dynamin 2, a GTPase that excises endocytic vesicles from the plasma membrane, and functions as its GTPase-activating protein. Interestingly, girdin depletion leads to the defect in clathrin-coated pit formation in the center of cells. Also, we find that girdin differentially interacts with some cargoes, which competitively prevents girdin from interacting with dynamin 2 and confers the cargo selectivity for CME. Therefore, girdin regulates transferrin and E-cadherin endocytosis in the center of cells and their subsequent polarized intracellular localization, but has no effect on integrin and epidermal growth factor receptor endocytosis that occurs at the cell periphery. Our results reveal that girdin regulates selective CME via a mechanism involving dynamin 2, but not by operating as a cargo-specific adaptor.
    The EMBO Journal 07/2014; 33(18). DOI:10.15252/embj.201488289 · 10.43 Impact Factor
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    • "Recently, Girdin, also named Akt phosphorylation enhancer (APE) or Gα-interacting vesicle-associated protein (GIV), was identified as an Akt or Gα binding protein in yeast two hybrid screens [6] [7] [8] or by computer homology search for Hook related proteins (HkRP1) [9]. So far, the main function ascribed to Girdin is to enhance and prolong the Akt kinase activity although the mechanism is still unclear. "
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    ABSTRACT: Akt kinases are important mediators of the insulin signal, and some Akt substrates are directly involved in glucose homeostasis. Recently, Girdin has been described as an Akt substrate that is expressed ubiquitously in mammals. Cells overexpressing Girdin show an enhanced Akt activity. However, not much is known about Girdin's role in insulin signaling. We therefore analyzed the role of Girdin in primary human myotubes and found a correlation between Girdin expression and insulin sensitivity of the muscle biopsy donors, as measured by a hyperinsulinemic-euglycemic clamp. To understand this finding on a cellular level, we then investigated the function of Girdin in C2C12 mouse myoblasts. Girdin knock-down reduced Akt and insulin receptor substrate-1 phosphorylation. In contrast, stable overexpression of Girdin in C2C12 cells strikingly increased insulin sensitivity through a massive upregulation of the insulin receptor and enhanced tyrosine phosphorylation of insulin receptor substrate-1. Furthermore, Akt and c-Abl kinases were constitutively activated. To investigate medium-term insulin responses we measured glucose incorporation into glycogen. The Girdin overexpressing cells showed a high basal glycogen synthesis that peaked already at 1 nM insulin. Taken together, we characterized Girdin as a new and major regulator of the insulin signal in myoblasts and skeletal muscle.
    Biochimica et Biophysica Acta 07/2013; 1833(12). DOI:10.1016/j.bbamcr.2013.07.012 · 4.66 Impact Factor
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