Copine A is expressed in prestalk cells and regulates slug phototaxis and thermotaxis in developing Dictyostelium

Flegel, Kerry; Pineda, Jaimie M; Smith, Tasha S.; Laszczyk, Ann M.; Price, J. M.; Karasiewicz, Kristen M; Damer, Cynthia Kay

doi:10.1111/j.1440-169x.2011.01300.x

Cited by 10 publications

(13 citation statements)

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“…Previously, we focused our studies on one of the copine proteins in Dictyostelium , CpnA [ 2 , 12 – 14 ]. In the present study, we have expanded our investigations to include four additional copines.…”

Section: Discussionmentioning

confidence: 99%

“…Vegetative cells lacking the cpnA gene display defects in cytokinesis and contractile vacuole function [ 12 ]. Developing cpnA − cells are delayed in aggregation, form larger than normal slugs, exhibit defects in slug thermotaxis and phototaxis, and fail to culminate into normal fruiting bodies [ 12 – 14 ]. GFP-tagged Copine A (GFP-CpnA) was shown to bind to membranes in a calcium-dependent manner and transiently translocate from the cytoplasm to the plasma membrane and intracellular vesicles in starved cells.…”

Section: Introductionmentioning

confidence: 99%

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Cyclic AMP signaling in Dictyostelium promotes the translocation of the copine family of calcium-binding proteins to the plasma membrane

et al. 2018

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BackgroundCopines are calcium-dependent phospholipid-binding proteins found in many eukaryotic organisms and are thought to be involved in signaling pathways that regulate a wide variety of cellular processes. Copines are characterized by having two C2 domains at the N-terminus accompanied by an A domain at the C-terminus. Six copine genes have been identified in the Dictyostelium genome, cpnA – cpnF.ResultsIndependent cell lines expressing CpnA, CpnB, CpnC, CpnE, or CpnF tagged with green fluorescent protein (GFP) were created as tools to study copine protein membrane-binding and localization. In general, the GFP-tagged copine proteins appeared to localize to the cytoplasm in live cells. GFP-tagged CpnB, CpnC, and CpnF were also found in the nucleus. When cells were fixed or when live cells were treated with calcium ionophore, the GFP-tagged copine proteins were found associated with the plasma membrane and vesicular organelles. When starved Dictyostelium cells were stimulated with cAMP, which causes a transitory increase in calcium concentration, all of the copines translocated to the plasma membrane, but with varying magnitudes and on and off times, suggesting each of the copines has distinct calcium-sensitivities and/or membrane-binding properties. In vitro membrane binding assays showed that all of the GFP-tagged copines pelleted with cellular membranes in the presence of calcium; yet, each copine displayed distinct calcium-independent membrane-binding in the absence of calcium. A lipid overlay assay with purified GFP-tagged copine proteins was used to screen for specific phospholipid-binding targets. Similar to other proteins that contain C2 domains, GFP-tagged copines bound to a variety of acidic phospholipids. CpnA, CpnB, and CpnE bound strongly to PS, PI(4)P, and PI(4,5)P2, while CpnC and CpnF bound strongly to PI(4)P.ConclusionsOur studies show that the Dictyostelium copines are soluble cytoplasmic and nuclear proteins that have the ability to bind intracellular membranes. Moreover, copines display different membrane-binding properties suggesting they play distinct roles in the cell. The transient translocation of copines to the plasma membrane in response to cAMP suggests copines may play a specific role in chemotaxis signaling.Electronic supplementary materialThe online version of this article (10.1186/s12860-018-0160-5) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Cyclic AMP signaling in Dictyostelium promotes the translocation of the copine family of calcium-binding proteins to the plasma membrane

et al. 2018

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“…We are studying the function of copines using the model organism Dictyostelium discoideum . We have identified six copine genes in Dictyostelium discoideum and have focused our studies on one of the copine proteins, CpnA [10,11,12,13]. Cells lacking the cpnA gene ( cpnA − ) were previously shown to have defects in cytokinesis, contractile vacuole function, and development [11].…”

Section: Introductionmentioning

confidence: 99%

“…When cpnA − cells were starved, they were delayed in aggregation to form the mound and then arrested at the slug stage [11]. The slugs formed by cpnA − cells were bigger than normal slugs, and they were not able to carry out normal phototaxis and thermotaxis [13].…”

Section: Introductionmentioning

confidence: 99%

Copine A Interacts with Actin Filaments and Plays a Role in Chemotaxis and Adhesion

Buccilli

Ilacqua

Han

et al. 2019

Cells

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Copines make up a family of calcium-dependent, phospholipid-binding proteins found in numerous eukaryotic organisms. Copine proteins consist of two C2 domains at the N-terminus followed by an A domain similar to the von Willebrand A domain found in integrins. We are studying copine protein function in the model organism, Dictyostelium discoideum, which has six copine genes, cpnA-cpnF. Previous research showed that cells lacking the cpnA gene exhibited a cytokinesis defect, a contractile vacuole defect, and developmental defects. To provide insight into the role of CpnA in these cellular processes, we used column chromatography and immunoprecipitation to isolate proteins that bind to CpnA. These proteins were identified by mass spectrometry. One of the proteins identified was actin. Purified CpnA was shown to bind to actin filaments in a calcium-dependent manner in vitro. cpnA− cells exhibited defects in three actin-based processes: chemotaxis, cell polarity, and adhesion. These results suggest that CpnA plays a role in chemotaxis and adhesion and may do so by interacting with actin filaments.

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“…We have focused our studies on one of the copines, CpnA. Our studies on cpnA knockout (KO) mutants ( cpnA − ) in Dictyostelium suggest that CpnA is involved in many cellular functions including cytokinesis, adhesion, and chemotaxis, and developmental functions including aggregation, slug phototaxis and thermotaxis, culmination, and stalk cell formation [12–15]. CpnA binds to membranes in a calcium‐dependent manner and specifically binds to acidic phospholipids with strongest binding to phosphatidylserine and phosphatidylinositol phosphate [11,16].…”

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Copine A regulates the size and exocytosis of contractile vacuoles and postlysosomes in Dictyostelium

2020

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Copines are a family of cytosolic proteins that associate with membranes in a calcium‐dependent manner and are found in many eukaryotic organisms. Dictyostelium discoideum has six copine genes (cpnA‐cpnF), and cells lacking cpnA(cpnA−) have defects in cytokinesis, chemotaxis, adhesion, and development. CpnA has also been shown to associate with the plasma membrane, contractile vacuoles (CV), and organelles of the endolysosomal pathway. Here, we use cpnA− cells to investigate the role of CpnA in CV function and endocytosis. When placed in water, cpnA− cells made abnormally large CVs that took longer to expel. Visualization of CVs with the marker protein GFP‐dajumin indicated that cpnA− cells had fewer CVs that sometimes refilled before complete emptying. In endocytosis assays, cpnA− cells took up small fluorescent beads by macropinocytosis at rates similar to parental cells. However, cpnA− cells reached a plateau sooner than parental cells and had less fluorescence at later time points. p80 antibody labeling of postlysosomes (PL) indicated that there were fewer and smaller PLs in cpnA− cells. In dextran pulse‐chase experiments, the number of PLs peaked earlier in cpnA− cells, and the PLs did not become as large and disappeared sooner as compared to parental cells. PLs in cpnA− cells were also shown to have more actin coats, suggesting CpnA may play a role in actin filament disassembly on PL membranes. Overall, these results indicate that CpnA is involved in the regulation of CV size and expulsion, and the maturation, size, and exocytosis of PLs.

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Copine A is expressed in prestalk cells and regulates slug phototaxis and thermotaxis in developing Dictyostelium

Cited by 10 publications

References 44 publications

Cyclic AMP signaling in Dictyostelium promotes the translocation of the copine family of calcium-binding proteins to the plasma membrane

Cyclic AMP signaling in Dictyostelium promotes the translocation of the copine family of calcium-binding proteins to the plasma membrane

Copine A Interacts with Actin Filaments and Plays a Role in Chemotaxis and Adhesion

Copine A regulates the size and exocytosis of contractile vacuoles and postlysosomes in Dictyostelium

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