scholarly journals Assembly of an FtsZ Mutant Deficient in GTPase Activity Has Implications for FtsZ Assembly and the Role of the Z Ring in Cell Division

2001 ◽  
Vol 183 (24) ◽  
pp. 7190-7197 ◽  
Author(s):  
Amit Mukherjee ◽  
Cristian Saez ◽  
Joe Lutkenhaus
Keyword(s):  
Cell Division ◽  
Critical Concentration ◽  
Gtpase Activity ◽  
Support Cell ◽  
Z Ring ◽  
Ftsz Assembly ◽  
The Stability

ABSTRACT FtsZ, the ancestral homologue of eukaryotic tubulins, assembles into the Z ring, which is required for cytokinesis in prokaryotic cells. Both FtsZ and tubulin have a GTPase activity associated with polymerization. Interestingly, the ftsZ2 mutant is viable, although the FtsZ2 mutant protein has dramatically reduced GTPase activity due to a glycine-for-aspartic acid substitution within the synergy loop. In this study, we have examined the properties of FtsZ2 and found that the reduced GTPase activity is not enhanced by DEAE-dextran-induced assembly, indicating it has a defective catalytic site. In the absence of DEAE-dextran, FtsZ2 fails to assemble unless supplemented with wild-type FtsZ. FtsZ has to be at or above the critical concentration for copolymerization to occur, indicating that FtsZ is nucleating the copolymers. The copolymers formed are relatively stable and appear to be stabilized by a GTP-cap. These results indicate that FtsZ2 cannot nucleate assembly in vitro, although it must in vivo. Furthermore, the stability of FtsZ-FtsZ2 copolymers argues that FtsZ2 polymers would be stable, suggesting that stable FtsZ polymers are able to support cell division.

scholarly journals Investigation of Regulation of FtsZ Assembly by SulA and Development of a Model for FtsZ Polymerization

2008 ◽  
Vol 190 (7) ◽  
pp. 2513-2526 ◽  
Author(s):  
Alex Dajkovic ◽  
Amit Mukherjee ◽  
Joe Lutkenhaus
Keyword(s):  
Kinetic Scheme ◽  
Sos Response ◽  
Ring Structure ◽  
Linear Polymers ◽  
Gtpase Activity ◽  
Gtp Hydrolysis ◽  
Z Ring ◽  
Ftsz Assembly

ABSTRACT In Escherichia coli FtsZ organizes into a cytoskeletal ring structure, the Z ring, which effects cell division. FtsZ is a GTPase, but the free energy of GTP hydrolysis does not appear to be used for generation of the constriction force, leaving open the question of the function of the GTPase activity of FtsZ. Here we study the mechanism by which SulA, an inhibitor of FtsZ induced during the SOS response, inhibits FtsZ function. We studied the effects of SulA on the in vitro activities of FtsZ, on Z rings in vivo, and on a kinetic model for FtsZ polymerization in silico. We found that the binding of SulA to FtsZ is necessary but not sufficient for inhibition of polymerization, since the assembly of FtsZ polymers in the absence of the GTPase activity was not inhibited by SulA. We developed a new model for FtsZ polymerization that accounts for the cooperativity of FtsZ and could account for cooperativity observed in other linear polymers. When SulA was included in the kinetic scheme, simulations revealed that SulA with strong affinity for FtsZ delayed, but did not prevent, the assembly of polymers when they were not hydrolyzing GTP. Furthermore, the simulations indicated that SulA controls the assembly of FtsZ by binding to a polymerization-competent form of the FtsZ molecule and preventing it from participating in assembly. In vivo stoichiometry of the disruption of Z rings by SulA suggests that FtsZ may undergo two cooperative transitions in forming the Z ring.

scholarly journals HNRNPA2B1 promotes multiple myeloma progression by increasing AKT3 expression via m6A-dependent stabilization of ILF3 mRNA

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Fengjie Jiang ◽  
Xiaozhu Tang ◽  
Chao Tang ◽  
Zhen Hua ◽  
Mengying Ke ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Cell Proliferation ◽  
Cellular Proliferation ◽  
Aberrant Expression ◽  
The Stability ◽  
Induced Apoptosis ◽  
Proliferation In Vitro

AbstractN6-methyladenosine (m6A) modification is the most prevalent modification in eukaryotic RNAs while accumulating studies suggest that m6A aberrant expression plays an important role in cancer. HNRNPA2B1 is a m6A reader which binds to nascent RNA and thus affects a perplexing array of RNA metabolism exquisitely. Despite unveiled facets that HNRNPA2B1 is deregulated in several tumors and facilitates tumor growth, a clear role of HNRNPA2B1 in multiple myeloma (MM) remains elusive. Herein, we analyzed the function and the regulatory mechanism of HNRNPA2B1 in MM. We found that HNRNPA2B1 was elevated in MM patients and negatively correlated with favorable prognosis. The depletion of HNRNPA2B1 in MM cells inhibited cell proliferation and induced apoptosis. On the contrary, the overexpression of HNRNPA2B1 promoted cell proliferation in vitro and in vivo. Mechanistic studies revealed that HNRNPA2B1 recognized the m6A sites of ILF3 and enhanced the stability of ILF3 mRNA transcripts, while AKT3 downregulation by siRNA abrogated the cellular proliferation induced by HNRNPA2B1 overexpression. Additionally, the expression of HNRNPA2B1, ILF3 and AKT3 was positively associated with each other in MM tissues tested by immunohistochemistry. In summary, our study highlights that HNRNPA2B1 potentially acts as a therapeutic target of MM through regulating AKT3 expression mediated by ILF3-dependent pattern.

scholarly journals Analysis of MinC Reveals Two Independent Domains Involved in Interaction with MinD and FtsZ

2000 ◽  
Vol 182 (14) ◽  
pp. 3965-3971 ◽  
Author(s):  
Zonglin Hu ◽  
Joe Lutkenhaus
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Functional Domains ◽  
Wild Type ◽  
Terminal Domain ◽  
Ring Assembly ◽  
Z Ring ◽  
Ftsz Assembly

ABSTRACT In Escherichia coli FtsZ assembles into a Z ring at midcell while assembly at polar sites is prevented by themin system. MinC, a component of this system, is an inhibitor of FtsZ assembly that is positioned within the cell by interaction with MinDE. In this study we found that MinC consists of two functional domains connected by a short linker. When fused to MalE the N-terminal domain is able to inhibit cell division and prevent FtsZ assembly in vitro. The C-terminal domain interacts with MinD, and expression in wild-type cells as a MalE fusion disrupts minfunction, resulting in a minicell phenotype. We also find that MinC is an oligomer, probably a dimer. Although the C-terminal domain is clearly sufficient for oligomerization, the N-terminal domain also promotes oligomerization. These results demonstrate that MinC consists of two independently functioning domains: an N-terminal domain capable of inhibiting FtsZ assembly and a C-terminal domain responsible for localization of MinC through interaction with MinD. The fusion of these two independent domains is required to achieve topological regulation of Z ring assembly.

scholarly journals Probing for Binding Regions of the FtsZ Protein Surface through Site-Directed Insertions: Discovery of Fully Functional FtsZ-Fluorescent Proteins

2016 ◽  
Vol 199 (1) ◽  
Author(s):  
Desmond A. Moore ◽  
Zakiya N. Whatley ◽  
Chandra P. Joshi ◽  
Masaki Osawa ◽  
Harold P. Erickson
Keyword(s):  
Cell Division ◽  
Fluorescent Proteins ◽  
Sole Source ◽  
Ring Structure ◽  
Content Type ◽  
Z Ring ◽  
Complete Cell ◽  
The Right

ABSTRACT FtsZ, a bacterial tubulin homologue, is a cytoskeletal protein that assembles into protofilaments that are one subunit thick. These protofilaments assemble further to form a “Z ring” at the center of prokaryotic cells. The Z ring generates a constriction force on the inner membrane and also serves as a scaffold to recruit cell wall remodeling proteins for complete cell division in vivo. One model of the Z ring proposes that protofilaments associate via lateral bonds to form ribbons; however, lateral bonds are still only hypothetical. To explore potential lateral bonding sites, we probed the surface of Escherichia coli FtsZ by inserting either small peptides or whole fluorescent proteins (FPs). Among the four lateral surfaces on FtsZ protofilaments, we obtained inserts on the front and back surfaces that were functional for cell division. We concluded that these faces are not sites of essential interactions. Inserts at two sites, G124 and R174, located on the left and right surfaces, completely blocked function, and these sites were identified as possible sites for essential lateral interactions. However, the insert at R174 did not interfere with association of protofilaments into sheets and bundles in vitro. Another goal was to find a location within FtsZ that supported insertion of FP reporter proteins while allowing the FtsZ-FPs to function as the sole source of FtsZ. We discovered one internal site, G55-Q56, where several different FPs could be inserted without impairing function. These FtsZ-FPs may provide advances for imaging Z-ring structure by superresolution techniques. IMPORTANCE One model for the Z-ring structure proposes that protofilaments are assembled into ribbons by lateral bonds between FtsZ subunits. Our study excluded the involvement of the front and back faces of the protofilament in essential interactions in vivo but pointed to two potential lateral bond sites, on the right and left sides. We also identified an FtsZ loop where various fluorescent proteins could be inserted without blocking function; these FtsZ-FPs functioned as the sole source of FtsZ. This advance provides improved tools for all fluorescence imaging of the Z ring and may be especially important for superresolution imaging.

scholarly journals Transcription Regulation of ezrA and Its Effect on Cell Division of Bacillus subtilis

2004 ◽  
Vol 186 (17) ◽  
pp. 5926-5932 ◽  
Author(s):  
Kuei-Min Chung ◽  
Hsin-Hsien Hsu ◽  
Suresh Govindan ◽  
Ban-Yang Chang
Keyword(s):  
Bacillus Subtilis ◽  
Cell Division ◽  
Critical Concentration ◽  
In Vitro Transcription ◽  
Ring Formation ◽  
Negative Regulator ◽  
Cell Length ◽  
Intracellular Level ◽  
Z Ring

ABSTRACT The EzrA protein of Bacillus subtilis is a negative regulator for FtsZ (Z)-ring formation. It is able to modulate the frequency and position of Z-ring formation during cell division. The loss of this protein results in cells with multiple Z rings located at polar as well as medial sites; it also lowers the critical concentration of FtsZ required for ring formation (P. A. Levin, I. G. Kurster, and A. D. Grossman, Proc. Natl. Acad. Sci. USA 96:9642-9647, 1999). We have studied the regulation of ezrA expression during the growth of B. subtilis and its effects on the intracellular level of EzrA as well as the cell length of B. subtilis. With the aid of promoter probing, primer extension, in vitro transcription, and Western blotting analyses, two overlapping σA-type promoters, P1 and P2, located about 100 bp upstream of the initiation codon of ezrA, have been identified. P1, supposed to be an extended −10 promoter, was responsible for most of the ezrA expression during the growth of B. subtilis. Disruption of this promoter reduced the intracellular level of EzrA very significantly compared with disruption of P2. Moreover, deletion of both promoters completely abolished EzrA in B. subtilis. More importantly, the cell length and percentage of filamentous cells of B. subtilis were significantly increased by disruption of the promoter(s). Thus, EzrA is required for efficient cell division during the growth of B. subtilis, despite serving as a negative regulator for Z-ring formation.

scholarly journals Architecture of the ring formed by the tubulin homologue FtsZ in bacterial cell division

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Piotr Szwedziak ◽  
Qing Wang ◽  
Tanmay A M Bharat ◽  
Matthew Tsim ◽  
Jan Löwe
Keyword(s):  
Cell Division ◽  
Molecular Model ◽  
Three Dimensional ◽  
Bacterial Cell Division ◽  
Electron Cryomicroscopy ◽  
E Coli ◽  
Ftsz Ring ◽  
Z Ring

Membrane constriction is a prerequisite for cell division. The most common membrane constriction system in prokaryotes is based on the tubulin homologue FtsZ, whose filaments in E. coli are anchored to the membrane by FtsA and enable the formation of the Z-ring and divisome. The precise architecture of the FtsZ ring has remained enigmatic. In this study, we report three-dimensional arrangements of FtsZ and FtsA filaments in C. crescentus and E. coli cells and inside constricting liposomes by means of electron cryomicroscopy and cryotomography. In vivo and in vitro, the Z-ring is composed of a small, single-layered band of filaments parallel to the membrane, creating a continuous ring through lateral filament contacts. Visualisation of the in vitro reconstituted constrictions as well as a complete tracing of the helical paths of the filaments with a molecular model favour a mechanism of FtsZ-based membrane constriction that is likely to be accompanied by filament sliding.

scholarly journals Volumetric Properties of Four-Stranded DNA Structures

Biology ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 813
Author(s):  
Tigran V. Chalikian ◽  
Robert B. Macgregor
Keyword(s):  
Regulation Of Transcription ◽  
Solvent Interactions ◽  
Dna Structures ◽  
Cellular Processes ◽  
Molecular Determinants ◽  
Telomere Biology ◽  
The Stability

Four-stranded non-canonical DNA structures including G-quadruplexes and i-motifs have been found in the genome and are thought to be involved in regulation of biological function. These structures have been implicated in telomere biology, genomic instability, and regulation of transcription and translation events. To gain an understanding of the molecular determinants underlying the biological role of four-stranded DNA structures, their biophysical properties have been extensively studied. The limited libraries on volume, expansibility, and compressibility accumulated to date have begun to provide insights into the molecular origins of helix-to-coil and helix-to-helix conformational transitions involving four-stranded DNA structures. In this article, we review the recent progress in volumetric investigations of G-quadruplexes and i-motifs, emphasizing how such data can be used to characterize intra-and intermolecular interactions, including solvation. We describe how volumetric data can be interpreted at the molecular level to yield a better understanding of the role that solute–solvent interactions play in modulating the stability and recognition events of nucleic acids. Taken together, volumetric studies facilitate unveiling the molecular determinants of biological events involving biopolymers, including G-quadruplexes and i-motifs, by providing one more piece to the thermodynamic puzzle describing the energetics of cellular processes in vitro and, by extension, in vivo.

Expanding the role of HsEg5 within the mitotic and post-mitotic phases of the cell cycle

1998 ◽  
Vol 111 (17) ◽  
pp. 2551-2561 ◽  
Author(s):  
C.M. Whitehead ◽  
J.B. Rattner
Keyword(s):  
Cell Division ◽  
Microtubule Bundle ◽  
Spindle Poles ◽  
Centrosome Separation ◽  
Spindle Dynamics ◽  
Spindle Microtubules ◽  
Protein Components ◽  
The Stability

The BimC family of kinesin like proteins are involved in spindle dynamics in a wide variety of organisms. The human member of this family, HsEg5, has been implicated in centrosome separation during prophase/prometaphase and in the organization of in vitro mitotic asters. HsEg5 displays a complex distribution during mitosis, associating with the centrosomes, spindle microtubules, specific regions of the intracellular bridge and a microtubule bundle that forms in association with the post-mitotic migration of the centrosome. In an effort to determine the function of HsEg5 during late mitotic events and refine its proposed function during early mitotic centrosome separation, we microinjected antibodies specific to HsEg5 into HeLa cells during various stages of mitosis. In the presence of HsEg5 antibodies we find that the microtubule arrays responsible for both pre- and post-mitotic centrosome movement never form. Similarly, the microtubule bundle within the intracellular bridge becomes prematurely altered following karyokinesis resulting in the loss of the microtubule array at either end of the bridge. In addition, some peri-centrosomal material at the spindle poles becomes fragmented and the distribution of the spindle protein NuMA becomes more concentrated at the minus ends of the spindle microtubules. Our study also provides direct evidence that there is a link between post-mitotic centrosome migration and Golgi complex positioning and reformation following mitosis. We conclude that HsEg5 plays a recurrent role in establishing and/or determining the stability of specific microtubule arrays that form during cell division and that this role may encompass the ability of HsEg5 to influence the distribution of other protein components associated with cell division

scholarly journals Dual Roles of Bradyrhizobium japonicumNickelin Protein in Nickel Storage and GTP-Dependent Ni Mobilization

2000 ◽  
Vol 182 (6) ◽  
pp. 1702-1705 ◽  
Author(s):  
Jonathan W. Olson ◽  
Robert J. Maier
Keyword(s):  
Gtpase Activity ◽  
Dual Roles ◽  
Nickel Ion ◽  
Nickel Ions ◽  
Hydrogenase Maturation ◽  
Nickel Binding ◽  
And Storage

ABSTRACT The hydrogenase accessory protein HypB, or nickelin, has two functions in the N2-fixing, H2-oxidizing bacterium Bradyrhizobium japonicum. One function of HypB involves the mobilization of nickel into hydrogenase. HypB also carries out a nickel storage/sequestering function in B. japonicum, binding nine nickel ions per monomer. Here we report that the two roles (nickel mobilization and storage) of HypB can be separated in vitro and in vivo using molecular and biochemical approaches. The role of HypB in hydrogenase maturation is completely dependent on its intrinsic GTPase activity; strains which produce a HypB protein that is severely deficient in GTPase activity but that fully retains nickel-sequestering ability cannot produce active hydrogenase even upon prolonged nickel supplementation. A HypB protein that lacks the nickel-binding polyhistidine region near the N terminus lacks only the nickel storage capacity function; it is still able to bind a single nickel ion and also retains complete GTPase activity.

scholarly journals HuR Protein in Hepatocellular Carcinoma: Implications in Development, Prognosis and Treatment

Biomedicines ◽  
2021 ◽  
Vol 9 (2) ◽  
pp. 119
Author(s):  
Vasiliki Papatheofani ◽  
Georgia Levidou ◽  
Panagiotis Sarantis ◽  
Evangelos Koustas ◽  
Michalis V. Karamouzis ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Liver Tumors ◽  
Normal Liver ◽  
Primary Liver Tumors ◽  
The Stability ◽  
Hu Antigen R ◽  
Post Transcriptional Regulation

Hu-antigen R (HuR) is a post-transcriptional regulator that belongs to the embryonic lethal abnormal vision Drosophila-like family (ELAV). HuR regulates the stability, translation, subcellular localization, and degradation of several target mRNAs, which are implicated in carcinogenesis and could affect therapeutic options. HuR protein is consistently highly expressed in hepatocellular carcinoma (HCC) compared to the adjacent normal liver tissue and is involved in the post-transcriptional regulation of various genes implicated in liver malignant transformation. Additionally, HuR protein seems to be a putative prognosticator in HCC, predicting worse survival. This review summarizes the recent evidence regarding the role of HuR in primary liver tumors, as presented in clinical studies, in vitro experiments and in vivo animal models. In conclusion, our review supports the consistent role of HuR protein in the development, prognosis, and treatment of HCC. Additional studies are expected to expand current information and exploit its putative employment as a future candidate for more personalized treatment in these tumors.

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