scholarly journals A Proteomics Analysis of Calmodulin-Binding Proteins in Dictyostelium discoideum during the Transition from Unicellular Growth to Multicellular Development

2021 ◽  
Vol 22 (4) ◽  
pp. 1722
Author(s):  
William D. Kim ◽  
Shyong Q. Yap ◽  
Robert J. Huber
Keyword(s):  
Dictyostelium Discoideum ◽  
Calcium Binding ◽  
Mammalian Cells ◽  
Binding Proteins ◽  
Model Organism ◽  
Developmental Processes ◽  
Multicellular Development ◽  
Calmodulin Binding ◽  
Search Tool

Calmodulin (CaM) is an essential calcium-binding protein within eukaryotes. CaM binds to calmodulin-binding proteins (CaMBPs) and influences a variety of cellular and developmental processes. In this study, we used immunoprecipitation coupled with mass spectrometry (LC-MS/MS) to reveal over 500 putative CaM interactors in the model organism Dictyostelium discoideum. Our analysis revealed several known CaMBPs in Dictyostelium and mammalian cells (e.g., myosin, calcineurin), as well as many novel interactors (e.g., cathepsin D). Gene ontology (GO) term enrichment and Search Tool for the Retrieval of Interacting proteins (STRING) analyses linked the CaM interactors to several cellular and developmental processes in Dictyostelium including cytokinesis, gene expression, endocytosis, and metabolism. The primary localizations of the CaM interactors include the nucleus, ribosomes, vesicles, mitochondria, cytoskeleton, and extracellular space. These findings are not only consistent with previous work on CaM and CaMBPs in Dictyostelium, but they also provide new insight on their diverse cellular and developmental roles in this model organism. In total, this study provides the first in vivo catalogue of putative CaM interactors in Dictyostelium and sheds additional light on the essential roles of CaM and CaMBPs in eukaryotes.

scholarly journals Calmodulin and Calmodulin Binding Proteins in Dictyostelium: A Primer

2020 ◽  
Vol 21 (4) ◽  
pp. 1210
Author(s):  
Danton H. O’Day ◽  
Ryan J. Taylor ◽  
Michael A. Myre
Keyword(s):  
Conformational Changes ◽  
Calcium Binding ◽  
Binding Proteins ◽  
Regulatory Protein ◽  
Model Organism ◽  
Ion Binding ◽  
Calcium Dependent ◽  
Calmodulin Binding ◽  
Human Counterpart ◽  
Ef Hands

Dictyostelium discoideum is gaining increasing attention as a model organism for the study of calcium binding and calmodulin function in basic biological events as well as human diseases. After a short overview of calcium-binding proteins, the structure of Dictyostelium calmodulin and the conformational changes effected by calcium ion binding to its four EF hands are compared to its human counterpart, emphasizing the highly conserved nature of this central regulatory protein. The calcium-dependent and -independent motifs involved in calmodulin binding to target proteins are discussed with examples of the diversity of calmodulin binding proteins that have been studied in this amoebozoan. The methods used to identify and characterize calmodulin binding proteins is covered followed by the ways Dictyostelium is currently being used as a system to study several neurodegenerative diseases and how it could serve as a model for studying calmodulinopathies such as those associated with specific types of heart arrythmia. Because of its rapid developmental cycles, its genetic tractability, and a richly endowed stock center, Dictyostelium is in a position to become a leader in the field of calmodulin research.

Calcium binding to complexes of calmodulin and calmodulin binding proteins

Biochemistry ◽  
1985 ◽  
Vol 24 (27) ◽  
pp. 8081-8086 ◽  
Author(s):  
Bradley B. Olwin ◽  
Daniel R. Storm
Keyword(s):  
Calcium Binding ◽  
Binding Proteins ◽  
Calmodulin Binding

scholarly journals Identification of Novel Human Cdt1-binding Proteins by a Proteomics Approach: Proteolytic Regulation by APC/CCdh1

2008 ◽  
Vol 19 (3) ◽  
pp. 1007-1021 ◽  
Author(s):  
Nozomi Sugimoto ◽  
Issay Kitabayashi ◽  
Satoko Osano ◽  
Yasutoshi Tatsumi ◽  
Takashi Yugawa ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Ubiquitin Ligase ◽  
Mammalian Cells ◽  
Topoisomerase I ◽  
Binding Proteins ◽  
Mass Spectrometry Analysis ◽  
Anaphase Promoting Complex ◽  
Chromosomal Damage ◽  
Spectrometry Analysis

In mammalian cells, Cdt1 activity is strictly controlled by multiple independent mechanisms, implying that it is central to the regulation of DNA replication during the cell cycle. In fact, unscheduled Cdt1 hyperfunction results in rereplication and/or chromosomal damage. Thus, it is important to understand its function and regulations precisely. We sought to comprehensively identify human Cdt1-binding proteins by a combination of Cdt1 affinity chromatography and liquid chromatography and tandem mass spectrometry analysis. Through this approach, we could newly identify 11 proteins, including subunits of anaphase-promoting complex/cyclosome (APC/C), SNF2H and WSTF, topoisomerase I and IIα, GRWD1/WDR28, nucleophosmin/nucleoplasmin, and importins. In vivo interactions of Cdt1 with APC/CCdh1, SNF2H, topoisomerase I and IIα, and GRWD1/WDR28 were confirmed by coimmunoprecipitation assays. A further focus on APC/CCdh1 indicated that this ubiquitin ligase controls the levels of Cdt1 during the cell cycle via three destruction boxes in the Cdt1 N-terminus. Notably, elimination of these destruction boxes resulted in induction of strong rereplication and chromosomal damage. Thus, in addition to SCFSkp2 and cullin4-based ubiquitin ligases, APC/CCdh1 is a third ubiquitin ligase that plays a crucial role in proteolytic regulation of Cdt1 in mammalian cells.

Tissue distribution of rat S100 alpha and S100 beta and S100-binding proteins

1987 ◽  
Vol 252 (3) ◽  
pp. C285-C289 ◽  
Author(s):  
D. B. Zimmer ◽  
L. J. Van Eldik
Keyword(s):  
Calcium Binding ◽  
Binding Proteins ◽  
Physiological Role ◽  
Rat Tissues ◽  
Differential Distribution ◽  
Binding Profile ◽  
Calmodulin Binding ◽  
The Individual ◽  
Intracellular Targets

To understand the physiological role of the calcium-binding proteins S100 alpha and S100 beta, it is necessary to determine the distribution of these proteins and detect their intracellular targets in various tissues. The distribution of immunoreactive S100 alpha and S100 beta in various rat tissues was examined by radioimmunoassay. All tissues examined contained detectable S100, but the S100 beta/S100 alpha ratio in each tissue differed. Brain, adipose, and testes contained 18- to 40-fold more S100 beta than S100 alpha; skin and liver contained approximately equivalent amounts and kidney, spleen, and heart contained 8- to 75-fold more S100 alpha than S100 beta. Analysis of S100-binding proteins by gel overlay showed that each tissue possessed its own complement of binding proteins. The S100 beta-binding profile was indistinguishable from the S100 alpha-binding profile and both of these profiles were distinct from the calmodulin-binding profile. These observations suggest that the differential distribution and quantity of the individual S100 polypeptides and their binding proteins in various tissues may be important factors in determining S100 function.

Surprising roles for phospholipid binding proteins revealed by high throughput geneticsThis paper is one of a selection of papers published in this special issue entitled “Second International Symposium on Recent Advances in Basic, Clinical, and Social Medicine” and has undergone the Journal's usual peer review process.

2010 ◽  
Vol 88 (4) ◽  
pp. 565-574 ◽  
Author(s):  
Marissa A. LeBlanc ◽  
Christopher R. McMaster
Keyword(s):  
High Throughput ◽  
Mammalian Cells ◽  
Binding Proteins ◽  
Essential Gene ◽  
Peer Review Process ◽  
Vesicular Transport ◽  
Lipid Binding ◽  
And Function ◽  
Lipid Binding Proteins

Saccharomyces cerevisiae remains an ideal organism for studying the cell biological roles of lipids in vivo, as yeast has phospholipid metabolic pathways similar to mammalian cells, is easy and economical to manipulate, and is genetically tractable. The availability of isogenic strains containing specific genetic inactivation of each non-essential gene allowed for the development of a high-throughput method, called synthetic genetic analysis (SGA), to identify and describe precise pathways or functions associated with specific genes. This review describes the use of SGA to aid in elucidating the function of two lipid-binding proteins that regulate vesicular transport, Sec14 and Kes1. Sec14 was first identified as a phosphatidylcholine (PC) – phosphatidylinositol (PI) transfer protein required for viability, with reduced Sec14 function resulting in diminished vesicular transport out of the trans-Golgi. Although Sec14 is required for cell viability, inactivating the KES1 gene that encodes for a member of the oxysterol binding protein family in cells lacking Sec14 function results in restoration of vesicular transport and cell growth. SGA analysis identified a role for Kes1 and Sec14 in regulating the level and function of Golgi PI-4-phosphate (PI-4-P). SGA also determined that Sec14 not only regulates vesicular transport out of the trans-Golgi, but also transport from endosomes to the trans-Golgi. Comparing SGA screens in databases, coupled with genetic and cell biological analyses, further determined that the PI-4-P pool affected by Kes1 is generated by the PI 4-kinase Pik1. An important biological role for Sec14 and Kes1 revealed by SGA is coordinate regulation of the Pik1-generated Golgi PI-4-P pool that in turn is essential for vesicular transport into and out of the trans-Golgi.

scholarly journals A Hairpin-like Structure within an AU-rich mRNA-destabilizing Element Regulates trans-Factor Binding Selectivity and mRNA Decay Kinetics

2005 ◽  
Vol 280 (23) ◽  
pp. 22406-22417 ◽  
Author(s):  
Elizabeth J. Fialcowitz ◽  
Brandy Y. Brewer ◽  
Bridget P. Keenan ◽  
Gerald M. Wilson
Keyword(s):  
Mammalian Cells ◽  
Binding Proteins ◽  
Phylogenetic Analyses ◽  
Higher Order ◽  
Mrna Turnover ◽  
Decay Kinetics ◽  
Binding Selectivity ◽  
Factor Binding ◽  
Factor Α

In mammals, rapid mRNA turnover directed by AU-rich elements (AREs) is mediated by selective association of cellular ARE-binding proteins. These trans-acting factors display overlapping RNA substrate specificities and may act to either stabilize or destabilize targeted transcripts; however, the mechanistic features of AREs that promote preferential binding of one trans-factor over another are not well understood. Here, we describe a hairpin-like structure adopted by the ARE from tumor necrosis factor α (TNFα) mRNA that modulates its affinity for selected ARE-binding proteins. In particular, association of the mRNA-destabilizing factor p37AUF1 was strongly inhibited by adoption of the higher order ARE structure, whereas binding of the inducible heat shock protein Hsp70 was less severely compromised. By contrast, association of the mRNA-stabilizing protein HuR was only minimally affected by changes in ARE folding. Consistent with the inverse relationship between p37AUF1 binding affinity and the stability of ARE folding, mutations that stabilized the ARE hairpin also inhibited its ability to direct rapid mRNA turnover in transfected cells. Finally, phylogenetic analyses and structural modeling indicate that TNFα mRNA sequences flanking the ARE are highly conserved and may stabilize the hairpin fold in vivo. Taken together, these data suggest that local higher order structures involving AREs may function as potent regulators of mRNA turnover in mammalian cells by modulating trans-factor binding selectivity.

Ca2+/calmodulin-binding proteins in Dictyostelium discoideum

1991 ◽  
Vol 142 (5) ◽  
pp. 509-519 ◽  
Author(s):  
T Winckler ◽  
H Dammann ◽  
R Mutzel
Keyword(s):  
Dictyostelium Discoideum ◽  
Binding Proteins ◽  
Calmodulin Binding

scholarly journals Mitochondrial regulation in Dictyostelium discoideum

2018 ◽  
Author(s):  
Mehak Rafiq
Keyword(s):  
Dictyostelium Discoideum ◽  
Serine Proteases ◽  
Model Organism ◽  
Complex Life Cycle ◽  
Transcriptional Networks ◽  
Multicellular Development ◽  
Mitochondrial Regulation ◽  
Rhomboid Proteases ◽  
The Social ◽  
Membrane Bound

Proteolysis is increasingly documented as a method of regulation of mitochondrial function. Our studies of rhomboidfamily proteins’ roles in organelles show that this is also the case in the social amoeba Dictyostelium discoideum, in which four of these membrane-bound, evolutionarily ubiquitous, serine proteases are found. Rhomboid proteases act on disparate substrates in different organisms so far studied, but their mode of action is conserved: their location in the membrane means that their membrane-tethered substrates can act in signalling upon release, or be activated, by rhomboid-mediated cleavage. Among eukaryotic rhomboids is the mitochondrial protease ‘PARL’, which ensures the maintenance of the structural and functional integrity of mitochondria and plastids, but we have found that other Dictyostelium rhomboids also affect the organelle. Studying the development and behaviour of Dictyostelium, a microbial model organism with a complex life cycle that includes uni- and multicellular stages, allowed investigation of the role of rhomboids in unicellular vegetative growth, multicellular development and sporulation, phagocytosis, and response to the environment. We found that two rhomboid-null mutants gave rise to changes in development, rhmA altering the response to chemoattractants and demonstrating decreased motility in general, whereas rhmB null cells had slower growth rates with decreased response to folic acid. RhmA, although located in the contractile vacuole, affects the ultrastructure of mitochondria, and RhmB-GFP fusions protein was localised to the mitochondrion. qPCR analysis revealed RhmA and RhmB transcript levels peaking during the multicellular growth phase and transcriptional networks suggest the Dictyostelium rhmA is regulated along with the orthologues of Saccharomyces cerevisiae mitochondrial rhomboid substrates.

1,25-Dihydroxyvitamin D-stimulated calmodulin binding proteins: a sustained effect on distal tubules

2002 ◽  
Vol 282 (1) ◽  
pp. F77-F84 ◽  
Author(s):  
Emmanuel K. O. Siaw ◽  
Marian R. Walters
Keyword(s):  
Vitamin D ◽  
Time Course ◽  
Binding Proteins ◽  
Day Treatment ◽  
Rat Kidney ◽  
Vitamin D Receptors ◽  
Dihydroxyvitamin D ◽  
Calmodulin Binding ◽  
1,25 Dihydroxyvitamin D

The tubular localization of 1,25-dihydroxyvitamin D[1,25(OH)2D3]-stimulated calmodulin binding proteins (CaMBP-Ds) in the rat kidney and the specificity of their induction were characterized to better understand renal responses to protracted 1,25(OH)2D3 treatment in vivo. None of the other hormones tested (parathyroid hormone, calcitonin, estradiol-17β, testosterone, progesterone, hydrocortisone, or dexamethasone) stimulated the CaMBP-Ds, whereas maximal 1,25(OH)2D3 stimulation occurred after a 5- to 7-day treatment with 100 ng/day 1,25(OH)2D3. With the exception of the more ubiquitously distributed CaMBP-D150, the CaMBP-Ds were localized in distal, but not proximal, tubule preparations. 1,25(OH)2D3 induction of vitamin D receptors and the CaMBP-Ds was similar with respect to dose-response and time course. Finally, the CaMBP-Ds remained elevated for at least 4 wk after 1,25(OH)2D3 withdrawal. Because the vitamin D-stimulated renal CaMBP-Ds are principally proteins of the distal tubule, they may be associated with renal regulation of Ca2+ homeostasis. The sustained induction of CaMBP-Ds is important in addressing the question of whether their induction is a function of normal Ca2+ homeostasis or a pathophysiological consequence of hypervitaminosis D and hypercalcemia.

scholarly journals CP110 Cooperates with Two Calcium-binding Proteins to Regulate Cytokinesis and Genome Stability

2006 ◽  
Vol 17 (8) ◽  
pp. 3423-3434 ◽  
Author(s):  
William Y. Tsang ◽  
Alexander Spektor ◽  
Daniel J. Luciano ◽  
Vahan B. Indjeian ◽  
Zhihong Chen ◽  
...  
Keyword(s):  
Calcium Binding ◽  
Genome Stability ◽  
Binding Proteins ◽  
Cell Formation ◽  
Eukaryotic Cell ◽  
Binucleate Cell ◽  
Binding Domains ◽  
Biochemical Analyses

The centrosome is an integral component of the eukaryotic cell cycle machinery, yet very few centrosomal proteins have been fully characterized to date. We have undertaken a series of biochemical and RNA interference (RNAi) studies to elucidate a role for CP110 in the centrosome cycle. Using a combination of yeast two-hybrid screens and biochemical analyses, we report that CP110 interacts with two different Ca2+-binding proteins, calmodulin (CaM) and centrin, in vivo. In vitro binding experiments reveal a direct, robust interaction between CP110 and CaM and the existence of multiple high-affinity CaM-binding domains in CP110. Native CP110 exists in large (∼300 kDa to 3 MDa) complexes that contain both centrin and CaM. We investigated a role for CP110 in CaM-mediated events using RNAi and show that its depletion leads to a failure at a late stage of cytokinesis and the formation of binucleate cells, mirroring the defects resulting from ablation of either CaM or centrin function. Importantly, expression of a CP110 mutant unable to bind CaM also promotes cytokinesis failure and binucleate cell formation. Taken together, our data demonstrate a functional role for CaM binding to CP110 and suggest that CP110 cooperates with CaM and centrin to regulate progression through cytokinesis.

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