scholarly journals Mammalian Abp1, a Signal-Responsive F-Actin–Binding Protein, Links the Actin Cytoskeleton to Endocytosis via the Gtpase Dynamin

2001 ◽  
Vol 153 (2) ◽  
pp. 351-366 ◽  
Author(s):  
Michael M. Kessels ◽  
Åsa E.Y. Engqvist-Goldstein ◽  
David G. Drubin ◽  
Britta Qualmann
Keyword(s):  
Actin Cytoskeleton ◽  
Binding Protein ◽  
Sh3 Domain ◽  
Actin Binding ◽  
Actin Binding Protein ◽  
Domain Interactions ◽  
Endocytic Machinery ◽  
Transferrin Uptake

The actin cytoskeleton has been implicated in endocytosis, yet few molecular links to the endocytic machinery have been established. Here we show that the mammalian F-actin–binding protein Abp1 (SH3P7/HIP-55) can functionally link the actin cytoskeleton to dynamin, a GTPase that functions in endocytosis. Abp1 binds directly to dynamin in vitro through its SH3 domain. Coimmunoprecipitation and colocalization studies demonstrated the in vivo relevance of this interaction. In neurons, mammalian Abp1 and dynamin colocalized at actin-rich sites proximal to the cell body during synaptogenesis. In fibroblasts, mAbp1 appeared at dynamin-rich sites of endocytosis upon growth factor stimulation. To test whether Abp1 functions in endocytosis, we overexpressed several Abp1 constructs in Cos-7 cells and assayed receptor-mediated endocytosis. While overexpression of Abp1's actin-binding modules did not interfere with endocytosis, overexpression of the SH3 domain led to a potent block of transferrin uptake. This implicates the Abp1/dynamin interaction in endocytic function. The endocytosis block was rescued by cooverexpression of dynamin. Since the addition of the actin-binding modules of Abp1 to the SH3 domain construct also fully restored endocytosis, Abp1 may support endocytosis by combining its SH3 domain interactions with cytoskeletal functions in response to signaling cascades converging on this linker protein.

scholarly journals Regulated Interactions between Dynamin and the Actin-Binding Protein Cortactin Modulate Cell Shape

2000 ◽  
Vol 151 (1) ◽  
pp. 187-198 ◽  
Author(s):  
Mark A. McNiven ◽  
Leung Kim ◽  
Eugene W. Krueger ◽  
James D. Orth ◽  
Hong Cao ◽  
...  
Keyword(s):  
Cell Migration ◽  
Actin Cytoskeleton ◽  
Cell Shape ◽  
Binding Protein ◽  
Sh3 Domain ◽  
Actin Binding ◽  
Coat Proteins ◽  
Actin Binding Protein ◽  
Morphological Studies

The dynamin family of large GTPases has been implicated in the formation of nascent vesicles in both the endocytic and secretory pathways. It is believed that dynamin interacts with a variety of cellular proteins to constrict membranes. The actin cytoskeleton has also been implicated in altering membrane shape and form during cell migration, endocytosis, and secretion and has been postulated to work synergistically with dynamin and coat proteins in several of these important processes. We have observed that the cytoplasmic distribution of dynamin changes dramatically in fibroblasts that have been stimulated to undergo migration with a motagen/hormone. In quiescent cells, dynamin 2 (Dyn 2) associates predominantly with clathrin-coated vesicles at the plasma membrane and the Golgi apparatus. Upon treatment with PDGF to induce cell migration, dynamin becomes markedly associated with membrane ruffles and lamellipodia. Biochemical and morphological studies using antibodies and GFP-tagged dynamin demonstrate an interaction with cortactin. Cortactin is an actin-binding protein that contains a well defined SH3 domain. Using a variety of biochemical methods we demonstrate that the cortactin–SH3 domain associates with the proline-rich domain (PRD) of dynamin. Functional studies that express wild-type and mutant forms of dynamin and/or cortactin in living cells support these in vitro observations and demonstrate that an increased expression of cortactin leads to a significant recruitment of endogenous or expressed dynamin into the cell ruffle. Further, expression of a cortactin protein lacking the interactive SH3 domain (CortΔSH3) significantly reduces dynamin localization to the ruffle. Accordingly, transfected cells expressing Dyn 2 lacking the PRD (Dyn 2(aa)ΔPRD) sequester little of this protein to the cortactin-rich ruffle. Interestingly, these mutant cells are viable, but display dramatic alterations in morphology. This change in shape appears to be due, in part, to a striking increase in the number of actin stress fibers. These findings provide the first demonstration that dynamin can interact with the actin cytoskeleton to regulate actin reorganization and subsequently cell shape.

scholarly journals Direct regulation of Arp2/3 complex activity and function by the actin binding protein coronin

2002 ◽  
Vol 159 (6) ◽  
pp. 993-1004 ◽  
Author(s):  
Christine L. Humphries ◽  
Heath I. Balcer ◽  
Jessica L. D'Agostino ◽  
Barbara Winsor ◽  
David G. Drubin ◽  
...  
Keyword(s):  
Binding Protein ◽  
Coiled Coil ◽  
Actin Binding ◽  
Complex Activity ◽  
Actin Binding Protein ◽  
Coiled Coil Domain ◽  
Allele Specific ◽  
And Function

Mechanisms for activating the actin-related protein 2/3 (Arp2/3) complex have been the focus of many recent studies. Here, we identify a novel mode of Arp2/3 complex regulation mediated by the highly conserved actin binding protein coronin. Yeast coronin (Crn1) physically associates with the Arp2/3 complex and inhibits WA- and Abp1-activated actin nucleation in vitro. The inhibition occurs specifically in the absence of preformed actin filaments, suggesting that Crn1 may restrict Arp2/3 complex activity to the sides of filaments. The inhibitory activity of Crn1 resides in its coiled coil domain. Localization of Crn1 to actin patches in vivo and association of Crn1 with the Arp2/3 complex also require its coiled coil domain. Genetic studies provide in vivo evidence for these interactions and activities. Overexpression of CRN1 causes growth arrest and redistribution of Arp2 and Crn1p into aberrant actin loops. These defects are suppressed by deletion of the Crn1 coiled coil domain and by arc35-26, an allele of the p35 subunit of the Arp2/3 complex. Further in vivo evidence that coronin regulates the Arp2/3 complex comes from the observation that crn1 and arp2 mutants display an allele-specific synthetic interaction. This work identifies a new form of regulation of the Arp2/3 complex and an important cellular function for coronin.

scholarly journals nPIST: A Novel Actin Binding Protein of trans-Golgi Network

2018 ◽  
Author(s):  
Swagata Das ◽  
Priyanka Dutta ◽  
Mohit Mazumder ◽  
Soma Seal ◽  
Kheerthana Duraivelan ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Purkinje Cells ◽  
Binding Protein ◽  
Vesicular Trafficking ◽  
Actin Binding ◽  
Perinuclear Region ◽  
Actin Binding Protein ◽  
Golgi Network

Abstractnpist is the neuronal isoform of PIST, a trans-golgi associated protein involved in major modulation of vesicular trafficking. nPIST interacts with glutamate delta2 receptor (GluRδ2) in Purkinje cells. Our study shows nPIST as a novel actin binding protein. Our structure based sequence analysis shows nPIST contains one WH2-like domain. Further our experimental analysis illustrates that fragment of nPIST consisting of WH2-like domain binds to actin. Moreover it was found that nPIST contains several regions involved in interaction with actin. The binding of nPIST to actin through multiple actin binding regions facilitated actin filament stabilization in vitro. In vivo, nPIST localized actin in perinuclear region as a blotch when ectopically expressed.

scholarly journals The Actin Binding Protein Plastin-3 Is Involved in the Pathogenesis of Acute Myeloid Leukemia

Cancers ◽  
2019 ◽  
Vol 11 (11) ◽  
pp. 1663 ◽  
Author(s):  
Arne Velthaus ◽  
Kerstin Cornils ◽  
Jan K. Hennigs ◽  
Saskia Grüb ◽  
Hauke Stamm ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Binding Protein ◽  
Actin Binding ◽  
Bone Marrow Niche ◽  
Actin Binding Protein ◽  
Acute Myeloid

Leukemia-initiating cells reside within the bone marrow in specialized niches where they undergo complex interactions with their surrounding stromal cells. We have identified the actin-binding protein Plastin-3 (PLS3) as potential player within the leukemic bone marrow niche and investigated its functional role in acute myeloid leukemia. High expression of PLS3 was associated with a poor overall and event-free survival for AML patients. These findings were supported by functional in vitro and in vivo experiments. AML cells with a PLS3 knockdown showed significantly reduced colony numbers in vitro while the PLS3 overexpression variants resulted in significantly enhanced colony numbers compared to their respective controls. Furthermore, the survival of NSG mice transplanted with the PLS3 knockdown cells showed a significantly prolonged survival in comparison to mice transplanted with the control AML cells. Further studies should focus on the underlying leukemia-promoting mechanisms and investigate PLS3 as therapeutic target.

scholarly journals Microtubule-associated Protein 2c Reorganizes Both Microtubules and Microfilaments into Distinct Cytological Structures in an Actin-binding Protein-280–deficient Melanoma Cell Line

1997 ◽  
Vol 136 (4) ◽  
pp. 845-857 ◽  
Author(s):  
C. Casey Cunningham ◽  
Nicole Leclerc ◽  
Lisa A. Flanagan ◽  
Mei Lu ◽  
Paul A. Janmey ◽  
...  
Keyword(s):  
Melanoma Cell ◽  
Binding Protein ◽  
Growth Cones ◽  
Actin Binding ◽  
Actin Binding Protein ◽  
Low Concentrations ◽  
Rheologic Property ◽  
Neuronal Growth Cones

The emergence of processes from cells often involves interactions between microtubules and microfilaments. Interactions between these two cytoskeletal systems are particularly apparent in neuronal growth cones. The juvenile isoform of the neuronal microtubule-associated protein 2 (MAP2c) is present in growth cones, where we hypothesize it mediates interactions between microfilaments and microtubules. To approach this problem in vivo, we used the human melanoma cell, M2, which lacks actin-binding protein-280 (ABP-280) and forms membrane blebs, which are not seen in wild-type or ABP-transfected cells. The microinjection of tau or mature MAP2 rescued the blebbing phenotype; MAP2c not only caused cessation of blebbing but also induced the formation of two distinct cellular structures. These were actin-rich lamellae, which often included membrane ruffles, and microtubule-bearing processes. The lamellae collapsed after treatment with cytochalasin D, and the processes retracted after treatment with colchicine. MAP2c was immunocytochemically visualized in zones of the cell that were devoid of tubulin, such as regions within the lamellae and in association with membrane ruffles. In vitro rheometry confirmed that MAP2c is an efficient actin gelation protein capable of organizing actin filaments into an isotropic array at very low concentrations; tau and mature MAP2 do not share this rheologic property. These results suggest that MAP2c engages in functionally specific interactions not only with microtubules but also with microfilaments.

scholarly journals The actin-binding protein Hip1R associates with clathrin during early stages of endocytosis and promotes clathrin assembly in vitro

2001 ◽  
Vol 154 (6) ◽  
pp. 1209-1224 ◽  
Author(s):  
Åsa E.Y. Engqvist-Goldstein ◽  
Robin A. Warren ◽  
Michael M. Kessels ◽  
James H. Keen ◽  
John Heuser ◽  
...  
Keyword(s):  
Binding Protein ◽  
Coiled Coil ◽  
Actin Binding ◽  
Live Cells ◽  
Cell Cortex ◽  
Coated Pits ◽  
Actin Binding Protein ◽  
Real Time Analysis

Huntingtin-interacting protein 1 related (Hip1R) is a novel component of clathrin-coated pits and vesicles and is a mammalian homologue of Sla2p, an actin-binding protein important for both actin organization and endocytosis in yeast. Here, we demonstrate that Hip1R binds via its putative central coiled-coil domain to clathrin, and provide evidence that Hip1R and clathrin are associated in vivo at sites of endocytosis. First, real-time analysis of Hip1R–YFP and DsRed–clathrin light chain (LC) in live cells revealed that these proteins show almost identical temporal and spatial regulation at the cell cortex. Second, at the ultrastructure level, immunogold labeling of ‘unroofed’ cells showed that Hip1R localizes to clathrin-coated pits. Third, overexpression of Hip1R affected the subcellular distribution of clathrin LC. Consistent with a functional role for Hip1R in endocytosis, we also demonstrated that it promotes clathrin cage assembly in vitro. Finally, we showed that Hip1R is a rod-shaped apparent dimer with globular heads at either end, and that it can assemble clathrin-coated vesicles and F-actin into higher order structures. In total, Hip1R's properties suggest an early endocytic function at the interface between clathrin, F-actin, and lipids.

scholarly journals The Actin‐Binding Protein Cofilin and Its Interaction With Cortactin Are Required for Podosome Patterning in Osteoclasts and Bone Resorption In Vivo and In Vitro

2016 ◽  
Vol 31 (9) ◽  
pp. 1701-1712 ◽  
Author(s):  
Detina Zalli ◽  
Lynn Neff ◽  
Kenichi Nagano ◽  
Nah Young Shin ◽  
Walter Witke ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Binding Protein ◽  
Actin Binding ◽  
Actin Binding Protein

Clathrin Hub Expression Dissociates the Actin-Binding Protein Hip1R from Coated Pits and Disrupts Their Alignment with the Actin Cytoskeleton

Traffic ◽  
2001 ◽  
Vol 2 (11) ◽  
pp. 851-858 ◽  
Author(s):  
Elizabeth M. Bennett ◽  
Chih-Ying Chen ◽  
Asa E. Y. Engqvist-Goldstein ◽  
David G. Drubin ◽  
Frances M. Brodsky
Keyword(s):  
Actin Cytoskeleton ◽  
Binding Protein ◽  
Actin Binding ◽  
Coated Pits ◽  
Actin Binding Protein

NUANCE, a giant protein connecting the nucleus and actin cytoskeleton

2002 ◽  
Vol 115 (15) ◽  
pp. 3207-3222 ◽  
Author(s):  
Yen-Yi Zhen ◽  
Thorsten Libotte ◽  
Martina Munck ◽  
Angelika A. Noegel ◽  
Elena Korenbaum
Keyword(s):  
Actin Cytoskeleton ◽  
Transmembrane Domain ◽  
Coiled Coil ◽  
Peripheral Blood Lymphocytes ◽  
Binding Domain ◽  
Actin Binding ◽  
Actin Binding Domain ◽  
Microfilament System

NUANCE (NUcleus and ActiN Connecting Element) was identified as a novel protein with an α-actinin-like actin-binding domain. A human 21.8 kb cDNA of NUANCE spreads over 373 kb on chromosome 14q22.1-q22.3. The cDNA sequence predicts a 796 kDa protein with an N-terminal actin-binding domain, a central coiled-coil rod domain and a predicted C-terminal transmembrane domain. High levels of NUANCE mRNA were detected in the kidney, liver,stomach, placenta, spleen, lymphatic nodes and peripheral blood lymphocytes. At the subcellular level NUANCE is present predominantly at the outer nuclear membrane and in the nucleoplasm. Domain analysis shows that the actin-binding domain binds to Factin in vitro and colocalizes with the actin cytoskeleton in vivo as a GFP-fusion protein. The C-terminal transmembrane domain is responsible for the targeting the nuclear envelope. Thus, NUANCE is the firstα-actinin-related protein that has the potential to link the microfilament system with the nucleus.

scholarly journals In vivo dynamics of the F-actin-binding protein neurabin-II

2000 ◽  
Vol 345 (2) ◽  
pp. 185 ◽  
Author(s):  
David J. STEPHENS ◽  
George BANTING
Keyword(s):  
Binding Protein ◽  
Actin Binding ◽  
Actin Binding Protein
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