scholarly journals The coordinated localization of mRNA to centrosomes facilitates error-free mitosis

2020 ◽  
Author(s):  
Pearl V. Ryder ◽  
Junnan Fang ◽  
Dorothy A. Lerit
Keyword(s):  
Cell Cycle ◽  
Genome Stability ◽  
Fragile X ◽  
Protein Localization ◽  
Cell Types ◽  
Rna Granules ◽  
Mental Retardation Protein ◽  
Pericentriolar Material ◽  
Cycle Dependent ◽  
Post Transcriptional Regulation

AbstractCentrosomes are microtubule-organizing centers required for error-free mitosis and embryonic development. The microtubule-nucleating activity of centrosomes is conferred by the pericentriolar material (PCM), a composite of numerous proteins subject to cell cycle-dependent oscillations in levels and organization. In diverse cell types, mRNAs localize to centrosomes and may contribute to changes in PCM abundance. Here, we investigate the regulation of mRNA localization to centrosomes in the rapidly cycling Drosophila melanogaster embryo. We find that RNA localization to centrosomes is regulated during the cell cycle and developmentally. We identify a novel role for the fragile-X mental retardation protein (FMRP), which localizes to pericentrosomal RNA granules, in the post-transcriptional regulation of centrosomal RNA. Further, the mis-targeting of a model centrosomal mRNA, centrocortin (cen), is sufficient to alter cognate protein localization to centrosomes and impair spindle morphogenesis and genome stability.

scholarly journals c entrocortin RNA localization to centrosomes is regulated by FMRP and facilitates error-free mitosis

2020 ◽  
Vol 219 (12) ◽  
Author(s):  
Pearl V. Ryder ◽  
Junnan Fang ◽  
Dorothy A. Lerit
Keyword(s):  
Cell Cycle ◽  
Posttranscriptional Regulation ◽  
Genome Stability ◽  
Fragile X ◽  
Protein Localization ◽  
Cell Types ◽  
Rna Localization ◽  
Mental Retardation Protein ◽  
Pericentriolar Material ◽  
Cycle Dependent

Centrosomes are microtubule-organizing centers required for error-free mitosis and embryonic development. The microtubule-nucleating activity of centrosomes is conferred by the pericentriolar material (PCM), a composite of numerous proteins subject to cell cycle–dependent oscillations in levels and organization. In diverse cell types, mRNAs localize to centrosomes and may contribute to changes in PCM abundance. Here, we investigate the regulation of mRNA localization to centrosomes in the rapidly cycling Drosophila melanogaster embryo. We find that RNA localization to centrosomes is regulated during the cell cycle and developmentally. We identify a novel role for the fragile-X mental retardation protein in the posttranscriptional regulation of a model centrosomal mRNA, centrocortin (cen). Further, mistargeting cen mRNA is sufficient to alter cognate protein localization to centrosomes and impair spindle morphogenesis and genome stability.

A monoclonal antibody, raised against mammalian centrosomes and screened by recognition of plant microtubule organizing centers, identifies a pericentriolar component in different cell types

1992 ◽  
Vol 101 (4) ◽  
pp. 823-835 ◽  
Author(s):  
V. Chevrier ◽  
S. Komesli ◽  
A.C. Schmit ◽  
M. Vantard ◽  
A.M. Lambert ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Monoclonal Antibody ◽  
Mammalian Cells ◽  
Cell Types ◽  
Immunoblot Analysis ◽  
Dependent Manner ◽  
Specific Staining ◽  
Microtubule Organizing Centers ◽  
Pericentriolar Material ◽  
Cycle Dependent

We have used monoclonal antibodies raised against isolated native calf thymus centrosomes to probe the structure and composition of the pericentriolar material. To distinguish prospective antibodies as specific to conserved elements of this material, we screened clones by their identification of microtubule organizing centers (MTOCs) in different animal and plant cells. Among the clonal antibodies that reacted with MTOCs in both plant and mammalian cells, we describe one (mAb 6C6) that was found to immunostain centrosomes in a variety of bovine and human cells. In cycling cells this signal persisted through the entire cell cycle. Microscopy showed that the mAb 6C6 antigen was a component of the pericentriolar material and this was confirmed by biochemical analysis of centrosomes. Using immunoblot analysis of protein fractions derived from purified components of centrosomes, we have characterized the mAb 6C6 antigen as a 180 kDa polypeptide. We conclude that we have identified a protein component permanently associated with the pericentriolar material. Surprisingly, monoclonal antibody 6C6 also stained other mitotic organelles in mammalian cells, in a cell-cycle-dependent manner. During prometaphase and metaphase the antibody stained both centrosomes and kinetochores. At the onset of anaphase the kinetochore-specific staining dissociated from chromosomes and was subsequently redistributed onto a newly characterized organelle, the telophase disc while the centrosomal stain remained intact. It is not known if the 180 kDa centrosomal protein itself redistributes during mitosis, or if the pattern observed represents other antigens with shared epitopes. The pericentriolar material is thought to be composed of conserved elements, which appeared very early during the evolution of eukaryotes. Our results strongly suggest that mAb 6C6 identifies one of these elements.

The Xenopus protein kinase pEg2 associates with the centrosome in a cell cycle-dependent manner, binds to the spindle microtubules and is involved in bipolar mitotic spindle assembly

1998 ◽  
Vol 111 (5) ◽  
pp. 557-572 ◽  
Author(s):  
C. Roghi ◽  
R. Giet ◽  
R. Uzbekov ◽  
N. Morin ◽  
I. Chartrain ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Protein Kinase ◽  
Mitotic Spindle ◽  
Cultured Cells ◽  
Dependent Manner ◽  
Egg Extracts ◽  
Pericentriolar Material ◽  
Spindle Microtubules ◽  
Cycle Dependent

By differential screening of a Xenopus laevis egg cDNA library, we have isolated a 2,111 bp cDNA which corresponds to a maternal mRNA specifically deadenylated after fertilisation. This cDNA, called Eg2, encodes a 407 amino acid protein kinase. The pEg2 sequence shows significant identity with members of a new protein kinase sub-family which includes Aurora from Drosophila and Ipl1 (increase in ploidy-1) from budding yeast, enzymes involved in centrosome migration and chromosome segregation, respectively. A single 46 kDa polypeptide, which corresponds to the deduced molecular mass of pEg2, is immunodetected in Xenopus oocyte and egg extracts, as well as in lysates of Xenopus XL2 cultured cells. In XL2 cells, pEg2 is immunodetected only in S, G2 and M phases of the cell cycle, where it always localises to the centrosomal region of the cell. In addition, pEg2 ‘invades’ the microtubules at the poles of the mitotic spindle in metaphase and anaphase. Immunoelectron microscopy experiments show that pEg2 is located precisely around the pericentriolar material in prophase and on the spindle microtubules in anaphase. We also demonstrate that pEg2 binds directly to taxol stabilised microtubules in vitro. In addition, we show that the presence of microtubules during mitosis is not necessary for an association between pEg2 and the centrosome. Finally we show that a catalytically inactive pEg2 kinase stops the assembly of bipolar mitotic spindles in Xenopus egg extracts.

scholarly journals CA Mutation N57A Has Distinct Strain-Specific HIV-1 Capsid Uncoating and Infectivity Phenotypes

2019 ◽  
Vol 93 (9) ◽  
Author(s):  
Douglas K. Fischer ◽  
Akatsuki Saito ◽  
Christopher Kline ◽  
Romy Cohen ◽  
Simon C. Watkins ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Amino Acid ◽  
Cell Types ◽  
Careful Consideration ◽  
Immunodeficiency Virus ◽  
Cell Cycle Dependence ◽  
Cycle Dependent ◽  
Multiple Strains ◽  
Different Cell Types ◽  
Hiv 1

ABSTRACTThe ability of human immunodeficiency virus type 1 (HIV-1) to transduce nondividing cells is key to infecting terminally differentiated macrophages, which can serve as a long-term reservoir of HIV-1 infection. The mutation N57A in the viral CA protein renders HIV-1 cell cycle dependent, allowing examination of HIV-1 infection of nondividing cells. Here, we show that the N57A mutation confers a postentry infectivity defect that significantly differs in magnitude between the common lab-adapted molecular clones HIV-1NL4-3(>10-fold) and HIV-1LAI(2- to 5-fold) in multiple human cell lines and primary CD4+T cells. Capsid permeabilization and reverse transcription are altered when N57A is incorporated into HIV-1NL4-3but not HIV-1LAI. The N57A infectivity defect is significantly exacerbated in both virus strains in the presence of cyclosporine (CsA), indicating that N57A infectivity is dependent upon CA interacting with host factor cyclophilin A (CypA). Adaptation of N57A HIV-1LAIselected for a second CA mutation, G94D, which rescued the N57A infectivity defect in HIV-1LAIbut not HIV-1NL4-3. The rescue of N57A by G94D in HIV-1LAIis abrogated by CsA treatment in some cell types, demonstrating that this rescue is CypA dependent. An examination of over 40,000 HIV-1 CA sequences revealed that the four amino acids that differ between HIV-1NL4-3and HIV-1LAICA are polymorphic, and the residues at these positions in the two strains are widely prevalent in clinical isolates. Overall, a few polymorphic amino acid differences between two closely related HIV-1 molecular clones affect the phenotype of capsid mutants in different cell types.IMPORTANCEThe specific mechanisms by which HIV-1 infects nondividing cells are unclear. A mutation in the HIV-1 capsid protein abolishes the ability of the virus to infect nondividing cells, serving as a tool to examine cell cycle dependence of HIV-1 infection. We have shown that two widely used HIV-1 molecular clones exhibit significantly different N57A infectivity phenotypes due to fewer than a handful of CA amino acid differences and that these clones are both represented in HIV-infected individuals. As such minor differences in closely related HIV-1 strains may impart significant infectivity differences, careful consideration should be given to drawing conclusions from one particular HIV-1 clone. This study highlights the potential for significant variation in results with the use of multiple strains and possible unanticipated effects of natural polymorphisms.

scholarly journals Single-molecule imaging of translational output from individual RNA granules in neurons

2012 ◽  
Vol 23 (5) ◽  
pp. 918-929 ◽  
Author(s):  
Vedakumar Tatavarty ◽  
Marius F. Ifrim ◽  
Mikhail Levin ◽  
George Korza ◽  
Elisa Barbarese ◽  
...  
Keyword(s):  
Single Molecule ◽  
Fluorescent Protein ◽  
Fragile X ◽  
Single Molecule Imaging ◽  
Wild Type ◽  
Rna Molecules ◽  
Temporal Regulation ◽  
Rna Granules ◽  
Mental Retardation Protein ◽  
Insight Into

Dendritic RNAs are localized and translated in RNA granules. Here we use single-molecule imaging to count the number of RNA molecules in each granule and to record translation output from each granule using Venus fluorescent protein as a reporter. For RNAs encoding activity-regulated cytoskeletal-associated protein (ARC) or fragile X mental retardation protein (FMRP), translation events are spatially clustered near individual granules, and translational output from individual granules is either sporadic or bursty. The probability of bursty translation is greater for Venus-FMRP RNA than for Venus-ARC RNA and is increased in Fmr1-knockout neurons compared to wild-type neurons. Dihydroxyphenylglycine (DHPG) increases the rate of sporadic translation and decreases bursty translation for Venus-FMRP and Venus-ARC RNAs. Single-molecule imaging of translation in individual granules provides new insight into molecular, spatial, and temporal regulation of translation in granules.

scholarly journals Plk4 triggers autonomous de novo centriole biogenesis and maturation

2021 ◽  
Vol 220 (5) ◽  
Author(s):  
Catarina Nabais ◽  
Delphine Pessoa ◽  
Jorge de-Carvalho ◽  
Thomas van Zanten ◽  
Paulo Duarte ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
De Novo ◽  
Cell Types ◽  
Proliferating Cells ◽  
Cycle Progression ◽  
Controlled System ◽  
Pericentriolar Material ◽  
Centriole Assembly ◽  
Critical Components

Centrioles form centrosomes and cilia. In most proliferating cells, centrioles assemble through canonical duplication, which is spatially, temporally, and numerically regulated by the cell cycle and the presence of mature centrioles. However, in certain cell types, centrioles assemble de novo, yet by poorly understood mechanisms. Herein, we established a controlled system to investigate de novo centriole biogenesis, using Drosophila melanogaster egg explants overexpressing Polo-like kinase 4 (Plk4), a trigger for centriole biogenesis. We show that at a high Plk4 concentration, centrioles form de novo, mature, and duplicate, independently of cell cycle progression and of the presence of other centrioles. Plk4 concentration determines the temporal onset of centriole assembly. Moreover, our results suggest that distinct biochemical kinetics regulate de novo and canonical biogenesis. Finally, we investigated which other factors modulate de novo centriole assembly and found that proteins of the pericentriolar material (PCM), and in particular γ-tubulin, promote biogenesis, likely by locally concentrating critical components.

scholarly journals The fragile X mental retardation protein is a molecular adaptor between the neurospecific KIF3C kinesin and dendritic RNA granules

2007 ◽  
Vol 16 (24) ◽  
pp. 3047-3058 ◽  
Author(s):  
L. Davidovic ◽  
X. H. Jaglin ◽  
A.-M. Lepagnol-Bestel ◽  
S. Tremblay ◽  
M. Simonneau ◽  
...  
Keyword(s):  
Mental Retardation ◽  
Fragile X ◽  
Rna Granules ◽  
Fragile X Mental Retardation ◽  
Mental Retardation Protein ◽  
Dendritic Rna

scholarly journals Fragile X–Related Protein 1 Regulates Nucleoporin Localization in a Cell Cycle–Dependent Manner

2021 ◽  
Vol 9 ◽  
Author(s):  
Arantxa Agote-Arán ◽  
Junyan Lin ◽  
Izabela Sumara
Keyword(s):  
Cell Cycle ◽  
Mammalian Cells ◽  
Cell Cycle Progression ◽  
Fragile X ◽  
Protein Translation ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Dependent Manner ◽  
Independent Manner ◽  
Cycle Dependent

Nuclear pore complexes (NPCs) are embedded in the nuclear envelope (NE) where they ensure the transport of macromolecules between the nucleus and the cytoplasm. NPCs are built from nucleoporins (Nups) through a sequential assembly order taking place at two different stages during the cell cycle of mammalian cells: at the end of mitosis and during interphase. In addition, fragile X–related proteins (FXRPs) can interact with several cytoplasmic Nups and facilitate their localization to the NE during interphase likely through a microtubule-dependent mechanism. In the absence of FXRPs or microtubule-based transport, Nups aberrantly localize to the cytoplasm forming the so-called cytoplasmic nucleoporin granules (CNGs), compromising NPCs’ function on protein export. However, it remains unknown if Nup synthesis or degradation mechanisms are linked to the FXRP–Nup pathway and if and how the action of FXRPs on Nups is coordinated with the cell cycle progression. Here, we show that Nup localization defects observed in the absence of FXR1 are independent of active protein translation. CNGs are cleared in an autophagy- and proteasome-independent manner, and their presence is restricted to the early G1 phase of the cell cycle. Our results thus suggest that a pool of cytoplasmic Nups exists that contributes to the NPC assembly specifically during early G1 to ensure NPC homeostasis at a short transition from mitosis to the onset of interphase.

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