scholarly journals Miro-dependent mitochondrial pool of CENP-F and its farnesylated C-terminal domain are dispensable for normal development in mice

2018 ◽  
Author(s):  
Martin Peterka ◽  
Benoît Kornmann
Keyword(s):  
Chromosome Segregation ◽  
Normal Development ◽  
Cultured Cells ◽  
Cancer Development ◽  
Point Mutant ◽  
Cellular Processes ◽  
C Terminus ◽  
Cultured Human Cells ◽  
Mitochondrial Trafficking ◽  
Cellular Phenotypes

AbstractCENP-F is a large, microtubule-binding protein that regulates multiple cellular processes including chromosome segregation and mitochondrial trafficking at cytokinesis. This multiplicity of function is mediated through the binding of various partners, like Bub1 at the kinetochore and Miro at mito-chondria. Due to the multifunctionality of CENP-F, the cellular phenotypes observed upon its depletion are difficult to interpret and there is a need to genetically separate its different functions by preventing binding to selected partners. Here we engineer a CENP-F point-mutant that is deficient in Miro binding and thus is unable to localize to mitochondria, but retains other localizations. We introduced this mutation in cultured human cells using CRISPR/Cas9 and show it causes a defect in mitochondrial spreading similar to that observed upon Miro depletion. We further create a mouse model carrying this CENP-F variant, as well as truncated CENP-F mutants lacking the farnesylated C-terminus of the protein. Importantly, one of these truncations leads to ∼80% downregulation of CENP-F expression. We observe that, despite the phenotypes apparent in cultured cells, mutant mice develop normally. Taken together, these mice will serve as important models to study CENP-F biology at organismal level. In addition, because truncations of CENP-F in humans cause a lethal disease termed Strømme syndrome and because CENP-F is involved in cancer development, they might also be relevant disease models.

scholarly journals The Bifunctional FtsK Protein Mediates Chromosome Partitioning and Cell Division in Caulobacter

2006 ◽  
Vol 188 (4) ◽  
pp. 1497-1508 ◽  
Author(s):  
Sherry C. E. Wang ◽  
Lisandra West ◽  
Lucy Shapiro
Keyword(s):  
Cell Division ◽  
Chromosome Segregation ◽  
Caulobacter Crescentus ◽  
Topoisomerase Iv ◽  
Division Plane ◽  
Cellular Processes ◽  
Chromosome Partitioning ◽  
C Terminus ◽  
N Terminus ◽  
Necessary And Sufficient

ABSTRACT Bacterial chromosome partitioning and cell division are tightly connected cellular processes. We show here that the Caulobacter crescentus FtsK protein localizes to the division plane, where it mediates multiple functions involved in chromosome segregation and cytokinesis. The first 258 amino acids of the N terminus are necessary and sufficient for targeting the protein to the division plane. Furthermore, the FtsK N terminus is required to either assemble or maintain FtsZ rings at the division plane. The FtsK C terminus is essential in Caulobacter and is involved in maintaining accurate chromosome partitioning. In addition, the C-terminal region of FtsK is required for the localization of the topoisomerase IV ParC subunit to the replisome to facilitate chromosomal decatenation prior to cell division. These results suggest that the interdependence between chromosome partitioning and cell division in Caulobacter is mediated, in part, by the FtsK protein.

E3 ubiquitin ligases

2005 ◽  
Vol 41 ◽  
pp. 15-30 ◽  
Author(s):  
Helen C. Ardley ◽  
Philip A. Robinson
Keyword(s):  
Protein Complexes ◽  
Loss Of Function ◽  
Direct Role ◽  
Cellular Processes ◽  
Substrate Protein ◽  
Protein Ubiquitination ◽  
C Terminus ◽  
Full Complement ◽  
Spatio Temporal ◽  
Eukaryotic Organisms

The selectivity of the ubiquitin–26 S proteasome system (UPS) for a particular substrate protein relies on the interaction between a ubiquitin-conjugating enzyme (E2, of which a cell contains relatively few) and a ubiquitin–protein ligase (E3, of which there are possibly hundreds). Post-translational modifications of the protein substrate, such as phosphorylation or hydroxylation, are often required prior to its selection. In this way, the precise spatio-temporal targeting and degradation of a given substrate can be achieved. The E3s are a large, diverse group of proteins, characterized by one of several defining motifs. These include a HECT (homologous to E6-associated protein C-terminus), RING (really interesting new gene) or U-box (a modified RING motif without the full complement of Zn2+-binding ligands) domain. Whereas HECT E3s have a direct role in catalysis during ubiquitination, RING and U-box E3s facilitate protein ubiquitination. These latter two E3 types act as adaptor-like molecules. They bring an E2 and a substrate into sufficiently close proximity to promote the substrate's ubiquitination. Although many RING-type E3s, such as MDM2 (murine double minute clone 2 oncoprotein) and c-Cbl, can apparently act alone, others are found as components of much larger multi-protein complexes, such as the anaphase-promoting complex. Taken together, these multifaceted properties and interactions enable E3s to provide a powerful, and specific, mechanism for protein clearance within all cells of eukaryotic organisms. The importance of E3s is highlighted by the number of normal cellular processes they regulate, and the number of diseases associated with their loss of function or inappropriate targeting.

scholarly journals Reggie/flotillin proteins are organized into stable tetramers in membrane microdomains

2007 ◽  
Vol 403 (2) ◽  
pp. 313-322 ◽  
Author(s):  
Gonzalo P. Solis ◽  
Maja Hoegg ◽  
Christina Munderloh ◽  
Yvonne Schrock ◽  
Edward Malaga-Trillo ◽  
...  
Keyword(s):  
T Cell Activation ◽  
Cell Activation ◽  
Coiled Coil ◽  
Cellular Prion Protein ◽  
Membrane Microdomains ◽  
Protein Assemblies ◽  
Cellular Processes ◽  
C Terminus ◽  
Functional Relevance ◽  
Interfering Rna

Reggie-1 and -2 proteins (flotillin-2 and -1 respectively) form their own type of non-caveolar membrane microdomains, which are involved in important cellular processes such as T-cell activation, phagocytosis and signalling mediated by the cellular prion protein and insulin; this is consistent with the notion that reggie microdomains promote protein assemblies and signalling. While it is generally known that membrane microdomains contain large multiprotein assemblies, the exact organization of reggie microdomains remains elusive. Using chemical cross-linking approaches, we have demonstrated that reggie complexes are composed of homo- and hetero-tetramers of reggie-1 and -2. Moreover, native reggie oligomers are indeed quite stable, since non-cross-linked tetramers are resistant to 8 M urea treatment. We also show that oligomerization requires the C-terminal but not the N-terminal halves of reggie-1 and -2. Using deletion constructs, we analysed the functional relevance of the three predicted coiled-coil stretches present in the C-terminus of reggie-1. We confirmed experimentally that reggie-1 tetramerization is dependent on the presence of coiled-coil 2 and, partially, of coiled-coil 1. Furthermore, since depletion of reggie-1 by siRNA (small interfering RNA) silencing induces proteasomal degradation of reggie-2, we conclude that the protein stability of reggie-2 depends on the presence of reggie-1. Our data indicate that the basic structural units of reggie microdomains are reggie homo- and hetero-tetramers, which are dependent on the presence of reggie-1.

scholarly journals Immunocytochemical localization of glucocorticoid receptors in midgestation murine embryos and human embryonic cultured cells.

1984 ◽  
Vol 32 (11) ◽  
pp. 1234-1237 ◽  
Author(s):  
C S Kim ◽  
J M Lauder ◽  
T H Joh ◽  
R M Pratt
Keyword(s):  
Cleft Palate ◽  
Glucocorticoid Receptors ◽  
Normal Development ◽  
Cultured Cells ◽  
Mesenchymal Cells ◽  
Immunocytochemical Localization ◽  
Secondary Palate ◽  
Murine Embryos ◽  
Mesenchyme Cells

Glucocorticoid receptors have been localized immunocytochemically in the developing mouse secondary palatal shelves and in cultured human embryonic palatal mesenchyme cells. In the midgestation embryo, receptors are found in the highest concentration in the palatal mesenchymal cells, suggesting that they play a major role in normal development as well as in glucocorticoid-induced cleft palate. The presence of these receptors in cultured human embryonic palatal cells also suggests that development of the human secondary palate may be dependent on glucocorticoids.

scholarly journals Proteotoxicity from aberrant ribosome biogenesis compromises cell fitness

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Blake W Tye ◽  
Nicoletta Commins ◽  
Lillia V Ryazanova ◽  
Martin Wühr ◽  
Michael Springer ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ribosomal Proteins ◽  
Ribosome Biogenesis ◽  
Ribosome Assembly ◽  
Maximal Growth ◽  
Proliferating Cells ◽  
Cellular Processes ◽  
Direct Contribution ◽  
The Face ◽  
Cellular Phenotypes

To achieve maximal growth, cells must manage a massive economy of ribosomal proteins (r-proteins) and RNAs (rRNAs) to produce thousands of ribosomes every minute. Although ribosomes are essential in all cells, natural disruptions to ribosome biogenesis lead to heterogeneous phenotypes. Here, we model these perturbations in Saccharomyces cerevisiae and show that challenges to ribosome biogenesis result in acute loss of proteostasis. Imbalances in the synthesis of r-proteins and rRNAs lead to the rapid aggregation of newly synthesized orphan r-proteins and compromise essential cellular processes, which cells alleviate by activating proteostasis genes. Exogenously bolstering the proteostasis network increases cellular fitness in the face of challenges to ribosome assembly, demonstrating the direct contribution of orphan r-proteins to cellular phenotypes. We propose that ribosome assembly is a key vulnerability of proteostasis maintenance in proliferating cells that may be compromised by diverse genetic, environmental, and xenobiotic perturbations that generate orphan r-proteins.

scholarly journals Bax Inhibitor 1 (BI-1) as a conservative regulator of Programmed Cell Death

2019 ◽  
Vol 73 ◽  
pp. 681-702
Author(s):  
Mirosław Godlewski ◽  
Agnieszka Kobylińska
Keyword(s):  
Cell Death ◽  
Programmed Cell Death ◽  
Actin Polymerization ◽  
Functional Condition ◽  
Sphingolipid Metabolism ◽  
Antiapoptotic Proteins ◽  
Crop Tolerance ◽  
Cellular Processes ◽  
C Terminus ◽  
Suicidal Death

Programmed cell death (PCD) is a physiological process in which infected or unnecessary cells due to their suicidal death capability can be selectively eliminated. Pro- and antiapoptotic proteins play an important role in the induction or inhibition of this process. Presented article shows property of Bax-1 (BI-1) inhibitor which is one of the conservative protein associated with the endoplasmic reticulum (ER) as well as its cytoprotective role in the regulation of cellular processes. It was shown that: 1) BI-1 is a small protein consisting of 237 amino acids (human protein - 36 kDa) and has 6 (in animals) and 7 (in plants) α-helical transmembrane domains, 2) BI-1 is expressed in all organisms and in most tissues, moreover its level depends on the functional condition of cells and it is involved in the development or reaction to biotic and abiotic stresses, 3) BI-1 forms a pH-dependent Ca2+ channel enabling release of these ions from the ER, 4) cytoprotective effects of BI-1 requires a whole, unchanged C-terminus, 5) BI-1 can interact directly with numerous other proteins, BI-1 protein affects numerous cellular processes, including: counteracting ER stress, oxidative stress, loss of cellular Ca2+ homeostasis as well as this protein influences on sphingolipid metabolism, autophagy, actin polymerization, lysosomal activity and cell proliferation. Studies of BI-1 functions will allow understanding the mechanisms of anticancer therapy or increases the knowledge of crop tolerance to environmental stresses.

scholarly journals Structural basis for UFM1 transfer from UBA5 to UFC1

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Manoj Kumar ◽  
Prasanth Padala ◽  
Jamal Fahoum ◽  
Fouad Hassouna ◽  
Tomer Tsaban ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Active Site ◽  
Structural Basis ◽  
Adenylation Domain ◽  
Post Translational Modification ◽  
Target Proteins ◽  
Cellular Processes ◽  
Enzyme Cascade ◽  
Linear Sequence ◽  
C Terminus

AbstractUfmylation is a post-translational modification essential for regulating key cellular processes. A three-enzyme cascade involving E1, E2 and E3 is required for UFM1 attachment to target proteins. How UBA5 (E1) and UFC1 (E2) cooperatively activate and transfer UFM1 is still unclear. Here, we present the crystal structure of UFC1 bound to the C-terminus of UBA5, revealing how UBA5 interacts with UFC1 via a short linear sequence, not observed in other E1-E2 complexes. We find that UBA5 has a region outside the adenylation domain that is dispensable for UFC1 binding but critical for UFM1 transfer. This region moves next to UFC1’s active site Cys and compensates for a missing loop in UFC1, which exists in other E2s and is needed for the transfer. Overall, our findings advance the understanding of UFM1’s conjugation machinery and may serve as a basis for the development of ufmylation inhibitors.

scholarly journals Heterochromatin: Guardian of the Genome

2018 ◽  
Vol 34 (1) ◽  
pp. 265-288 ◽  
Author(s):  
Aniek Janssen ◽  
Serafin U. Colmenares ◽  
Gary H. Karpen
Keyword(s):  
Chromosome Segregation ◽  
Genome Stability ◽  
Constitutive Heterochromatin ◽  
Repetitive Sequences ◽  
Genome Integrity ◽  
Chromatin Domain ◽  
Cellular Processes ◽  
Eukaryotic Nucleus ◽  
And Function ◽  
Heterochromatin Structure

Constitutive heterochromatin is a major component of the eukaryotic nucleus and is essential for the maintenance of genome stability. Highly concentrated at pericentromeric and telomeric domains, heterochromatin is riddled with repetitive sequences and has evolved specific ways to compartmentalize, silence, and repair repeats. The delicate balance between heterochromatin epigenetic maintenance and cellular processes such as mitosis and DNA repair and replication reveals a highly dynamic and plastic chromatin domain that can be perturbed by multiple mechanisms, with far-reaching consequences for genome integrity. Indeed, heterochromatin dysfunction provokes genetic turmoil by inducing aberrant repeat repair, chromosome segregation errors, transposon activation, and replication stress and is strongly implicated in aging and tumorigenesis. Here, we summarize the general principles of heterochromatin structure and function, discuss the importance of its maintenance for genome integrity, and propose that more comprehensive analyses of heterochromatin roles in tumorigenesis will be integral to future innovations in cancer treatment.

scholarly journals Temperature-Regulated Microcolony Formation by Burkholderia pseudomallei Requires pilA and Enhances Association with Cultured Human Cells

2006 ◽  
Vol 74 (9) ◽  
pp. 5374-5381 ◽  
Author(s):  
Justin A. Boddey ◽  
Cameron P. Flegg ◽  
Chris J. Day ◽  
Ifor R. Beacham ◽  
Ian R. Peak
Keyword(s):  
Biofilm Formation ◽  
Burkholderia Pseudomallei ◽  
Cultured Cells ◽  
Genomic Variation ◽  
Eukaryotic Cells ◽  
Northern Australia ◽  
Fatal Disease ◽  
Cell Association ◽  
Cultured Human Cells ◽  
Encoding Gene

ABSTRACT Burkholderia pseudomallei is the causative agent of melioidosis, a potentially fatal disease that is endemic to Northern Australia and Southeast Asia and is acquired from soil or water. Adherence of B. pseudomallei 08 to cultured cells increases dramatically following prior growth at 30°C or less compared to that following prior growth at 37°C. Here, we show that this occurs almost entirely as the result of microcolony formation (bacterium-bacterium interactions) following growth at 27°C but not at 37°C, which considerably enhances bacterial association with eukaryotic cells. Further, we demonstrate that the type IVA pilin-encoding gene, pilA, is essential for microcolony development by B. pseudomallei 08, and thus optimum association with eukaryotic cells, but is not required for direct adherence (bacterium-cell interactions). In contrast, although the B. pseudomallei genome sequence strain, K96243, also contains transcriptionally active pilA, microcolony formation rarely occurs following growth at either 27°C or 37°C and cell association occurs significantly less than with strain 08. Analysis of pilA transcription in 08 identified that pilA is dramatically upregulated under microcolony-forming conditions, viz., growth at low temperature, and association with eukaryotic cells; the pattern of transcription of pilA in K96243 differed from that in 08. Our study also suggests that biofilm formation by B. pseudomallei 08 and K96243 on polyvinylchloride is not mediated by pilA. Adherence and microcolony formation, and pilA transcription, vary between strains, consistent with known genomic variation in B. pseudomallei, and these phenotypes may be relevant to colonization from the environment.

Role of the C Terminus of FtsK in Escherichia coli Chromosome Segregation

1998 ◽  
Vol 180 (23) ◽  
pp. 6424-6428 ◽  
Author(s):  
Xuan-Chuan Yu ◽  
Elizabeth K. Weihe ◽  
William Margolin
Keyword(s):  
Escherichia Coli ◽  
Chromosome Segregation ◽  
C Terminus
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