On the molecular mechanisms of mitotic kinase activation

During mitosis, human cells exhibit a peak of protein phosphorylation that alters the behaviour of a significant proportion of proteins, driving a dramatic transformation in the cell's shape, intracellular structures and biochemistry. These mitotic phosphorylation events are catalysed by several families of protein kinases, including Auroras, Cdks, Plks, Neks, Bubs, Haspin and Mps1/TTK. The catalytic activities of these kinases are activated by phosphorylation and through protein–protein interactions. In this review, we summarize the current state of knowledge of the structural basis of mitotic kinase activation mechanisms. This review aims to provide a clear and comprehensive primer on these mechanisms to a broad community of researchers, bringing together the common themes, and highlighting specific differences. Along the way, we have uncovered some features of these proteins that have previously gone unreported, and identified unexplored questions for future work. The dysregulation of mitotic kinases is associated with proliferative disorders such as cancer, and structural biology will continue to play a critical role in the development of chemical probes used to interrogate disease biology and applied to the treatment of patients.

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The structural mechanisms that underpin mitotic kinase activation

Biochemical Society Transactions ◽

10.1042/bst20130066 ◽

2013 ◽

Vol 41 (4) ◽

pp. 1037-1041 ◽

Cited By ~ 12

Author(s):

Charlotte A. Dodson ◽

Tamanna Haq ◽

Sharon Yeoh ◽

Andrew M. Fry ◽

Richard Bayliss

Keyword(s):

Protein Phosphorylation ◽

Protein Kinases ◽

Rational Design ◽

Significant Proportion ◽

Small Subset ◽

Kinase Activation ◽

Mitotic Kinases ◽

Mitotic Kinase ◽

Almost All ◽

Structural Mechanisms

In eukaryotic cells, the peak of protein phosphorylation occurs during mitosis, switching the activities of a significant proportion of proteins and orchestrating a wholesale reorganization of cell shape and internal architecture. Most mitotic protein phosphorylation events are catalysed by a small subset of serine/threonine protein kinases. These include members of the Cdk (cyclin-dependent kinase), Plk (Polo-like kinase), Aurora, Nek (NimA-related kinase) and Bub families, as well as Haspin, Greatwall and Mps1/TTK. There has been steady progress in resolving the structural mechanisms that regulate the catalytic activities of these mitotic kinases. From structural and biochemical perspectives, kinase activation appears not as a binary process (from inactive to active), but as a series of states that exhibit varying degrees of activity. In its lowest activity state, a mitotic kinase may exhibit diverse autoinhibited or inactive conformations. Kinase activation proceeds via phosphorylation and/or association with a binding partner. These remodel the structure into an active conformation that is common to almost all protein kinases. However, all mitotic kinases of known structure have divergent features, many of which are key to understanding their specific regulatory mechanisms. Finally, mitotic kinases are an important class of drug target, and their structural characterization has facilitated the rational design of chemical inhibitors.

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Deciphering the Mounting Complexity of the p53 Regulatory Network in Correlation to Long Non-Coding RNAs (lncRNAs) in Ovarian Cancer

Cells ◽

10.3390/cells9030527 ◽

2020 ◽

Vol 9 (3) ◽

pp. 527 ◽

Cited By ~ 10

Author(s):

Sonali Pal ◽

Manoj Garg ◽

Amit Kumar Pandey

Keyword(s):

Ovarian Cancer ◽

Molecular Mechanisms ◽

Human Cancer ◽

Critical Role ◽

Functional Characterization ◽

Great Promise ◽

Altered Expression ◽

Disease Biology ◽

Non Coding Rnas ◽

Post Transcriptional Regulation

Amongst the various gynecological malignancies affecting female health globally, ovarian cancer is one of the predominant and lethal among all. The identification and functional characterization of long non-coding RNAs (lncRNAs) are made possible with the advent of RNA-seq and the advancement of computational logarithm in understanding human disease biology. LncRNAs can interact with deoxyribonucleic acid (DNA), ribonucleic acid (RNA), proteins and their combinations. Moreover, lncRNAs regulate orchestra of diverse functions including chromatin organization and transcriptional and post-transcriptional regulation. LncRNAs have conferred their critical role in key biological processes in human cancer including tumor initiation, proliferation, cell cycle, apoptosis, necroptosis, autophagy, and metastasis. The interwoven function of tumor-suppressor protein p53-linked lncRNAs in the ovarian cancer paradigm is of paramount importance. Several lncRNAs operate as p53 regulators or effectors and modulates a diverse array of functions either by participating in various signaling cascades or via interaction with different proteins. This review highlights the recent progress made in the identification of p53 associated lncRNAs while elucidating their molecular mechanisms behind the altered expression in ovarian cancer tumorigenesis. Moreover, the development of novel clinical and therapeutic strategies for targeting lncRNAs in human cancers harbors great promise.

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Determining rewiring effects of alternatively spliced isoforms on protein-protein interactions using a computational approach

10.1101/256834 ◽

2018 ◽

Author(s):

Oleksandr Narykov ◽

Nathan Johnson ◽

Dmitry Korkin

Keyword(s):

Protein Interactions ◽

Molecular Mechanisms ◽

Critical Role ◽

Computational Approach ◽

Superior Performance ◽

Large Set ◽

Protein Protein Interactions ◽

Specific Expression ◽

Alternatively Spliced ◽

Specific Regulation

AbstractThe critical role of alternative splicing (AS) in cell functioning has recently become apparent, whether in studying tissue-or cell-specific regulation, or understanding molecular mechanisms governing a complex disorder. Studying the rewiring, or edgetic, effects of alternatively spliced isoforms on protein interactome can provide system-wide insights into these questions. Unfortunately, high-throughput experiments for such studies are expensive and time-consuming, hence the need to develop an in-silico approach. Here, we formulated the problem of characterization the edgetic effects of AS on protein-protein interactions (PPIs) as a binary classification problem and introduced a first computational approach to solve it. We first developed a supervised feature-based classifier that benefited from the traditional features describing a PPI, the problem-specific features that characterized the difference between the reference and alternative isoforms, and a novel domain interaction potential that allowed pinpointing the domains employed during a specific PPI. We then expanded this approach by including a large set of unlabeled interactomics data and developing a semi-supervised learning method. Our method called AS-IN (Alternatively Splicing INteraction prediction) Tool was compared with the state-of-the-art PPI prediction tools and showed a superior performance, achieving 0.92 in precision and recall. We demonstrated the utility of AS-IN Tool by applying it to the transcriptomic data obtained from the brain and liver tissues of a healthy mouse and western diet fed mouse that developed type two diabetes. We showed that the edgetic effects of differentially expressed transcripts associated with the disease condition are system-wide and unlikely to be detected by looking only at the gene-specific expression levels.

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Structural basis of Atg conjugation systems essential for autophagy

Acta Crystallographica Section A Foundations and Advances ◽

10.1107/s2053273314096971 ◽

2014 ◽

Vol 70 (a1) ◽

pp. C302-C302

Author(s):

Nobuo Noda

Keyword(s):

Molecular Mechanisms ◽

Critical Role ◽

Amide Bond ◽

Structural Basis ◽

Autophagosome Formation ◽

X Ray Crystallography ◽

Intracellular Degradation ◽

Atg Proteins ◽

Evolutionarily Conserved ◽

Biochemical Analyses

Autophagy is an evolutionarily-conserved, intracellular degradation system for which two ubiquitin-like modifiers, Atg8 and Atg12, play essential roles. After processed by Atg4, the exposed C-terminal glycine of Atg8 is activated by Atg7 (E1) and is then transferred to Atg3 (E2), and is finally conjugated with a phospholipid, phosphatidylethanolamine (PE) through an amide bond. Whereas, Atg12 is activated by the same E1, Atg7, without processing, and is then transferred to Atg10 (E2), and is finally conjugated with Atg5 through an isopeptide bond. Atg12-Atg5 conjugates, together with Atg16, function as an E3-like enzyme to facilitate the conjugation reaction between Atg8 and PE. During autophagy, Atg8-PE conjugates play a critical role in selective cargo recognition in addition to autophagosome formation. We determined the structures of all these Atg proteins and their complexes mainly by X-ray crystallography, and performed structure-based biochemical analyses on them [1,2]. These studies established the molecular mechanisms of Atg8 and Atg12 modification reactions that have many unique features compared with canonical ubiquitin-like systems. Furthermore, we found a conserved motif named the Atg8-family interacting motif (AIM), through which Atg8 recognizes specific cargoes and selectively incorporates them into autophagosomes for degradation [3].

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Structural basis for the interaction of SARS-CoV-2 virulence factor nsp1 with Pol α - Primase

10.1101/2021.06.17.448816 ◽

2021 ◽

Author(s):

Mairi L Kilkenny ◽

Charlotte E Veale ◽

Amir Guppy ◽

Steven W Hardwick ◽

Dimitri Y Chirgadze ◽

...

Keyword(s):

Virulence Factor ◽

Protein Interactions ◽

Molecular Mechanisms ◽

Structural Basis ◽

Structural Protein ◽

Protein Protein Interactions ◽

Major Virulence Factor ◽

Biochemical Characterisation ◽

Human Proteins ◽

Primase Complex

The molecular mechanisms that drive the infection by the SARS-CoV-2 coronavirus, the causative agent of the COVID-19 (Coronavirus disease-2019) pandemic, are under intense current scrutiny, to understand how the virus operates and to uncover ways in which the disease can be prevented or alleviated. Recent cell-based analyses of SARS-CoV-2 protein - protein interactions have mapped the human proteins targeted by the virus. The DNA polymerase α - primase complex or primosome, responsible for initiating DNA synthesis in genomic duplication, was identified as a target of nsp1 (non structural protein 1), a major virulence factor in the SARS-CoV-2 infection. Here, we report the biochemical characterisation of the interaction between nsp1 and the primosome and the cryoEM structure of the primosome - nsp1 complex. Our data provide a structural basis for the reported interaction between the primosome and nsp1. They suggest that Pol α - primase plays a part in the immune response to the viral infection, and that its targeting by SARS-CoV-2 aims to interfere with such function.

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Cyclin A2 localises in the cytoplasm at the S/G2 transition to activate PLK1

10.1101/191437 ◽

2017 ◽

Cited By ~ 2

Author(s):

Helena Silva Cascales ◽

Kamila Burdova ◽

Anna Middleton ◽

Vladislav Kuzin ◽

Erik Müllers ◽

...

Keyword(s):

Dna Damage ◽

Dna Replication ◽

Feedback Loops ◽

List Type ◽

Kinase Activation ◽

Mitotic Kinases ◽

Mitotic Kinase ◽

Transition Functions ◽

G2 Phase ◽

Cyclin A2

AbstractCyclin A2 is a key regulator of the cell cycle, implicated both in DNA replication and mitotic entry. Cyclin A2 participates in feedback loops that activate mitotic kinases in G2-phase, but why active Cyclin A2-CDK2 during S phase does not trigger mitotic kinase activation remains unclear. Here we describe a change in localisation of Cyclin A2 from being only nuclear to both nuclear and cytoplasmic at the S/G2 border. We find that Cyclin A2-CDK2 can activate the mitotic kinase PLK1 through phosphorylation of Bora, and that only cytoplasmic Cyclin A2 interacts with Bora and PLK1. Expression of predominately cytoplasmic Cyclin A2 or phospho-mimicking PLK1 T210D can partially rescue a G2 arrest caused by Cyclin A2 depletion. Cytoplasmic presence of Cyclin A2 is restricted by p21, in particular after DNA damage. Cyclin A2 chromatin association during DNA replication and additional mechanisms contribute to Cyclin A2 localisation change in G2 phase. We find no evidence that such mechanisms involve G2 feedback loops and suggest that cytoplasmic appearance of Cyclin A2 at the S/G2 transition functions as a trigger for mitotic kinase activation.SynopsisMain mitotic kinases as PLK1 are activated at the S/G2 transition. A change in Cyclin A2 localisation at the S/G2 transition enables activation of PLK1.Main points-Cyclin A2 appears in the cytoplasm at the S/G2 transition-Association with replicating chromatin and p21 restricts Cyclin A2 to the nucleus-DNA damage ensures nuclear Cyclin A2 through p21-Cytoplasmic Cyclin A2 initiates PLK1 activationGraphical abstract

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Cyclin A2 localises in the cytoplasm at the S/G2 transition to activate PLK1

Life Science Alliance ◽

10.26508/lsa.202000980 ◽

2021 ◽

Vol 4 (3) ◽

pp. e202000980

Author(s):

Helena Silva Cascales ◽

Kamila Burdova ◽

Anna Middleton ◽

Vladislav Kuzin ◽

Erik Müllers ◽

...

Keyword(s):

Dna Replication ◽

Feedback Loops ◽

Kinase Activation ◽

Mitotic Entry ◽

Mitotic Kinases ◽

Mitotic Kinase ◽

Transition Functions ◽

G2 Phase ◽

Cyclin A2 ◽

Plk1 Expression

Cyclin A2 is a key regulator of the cell cycle, implicated both in DNA replication and mitotic entry. Cyclin A2 participates in feedback loops that activate mitotic kinases in G2 phase, but why active Cyclin A2-CDK2 during the S phase does not trigger mitotic kinase activation remains unclear. Here, we describe a change in localisation of Cyclin A2 from being only nuclear to both nuclear and cytoplasmic at the S/G2 border. We find that Cyclin A2-CDK2 can activate the mitotic kinase PLK1 through phosphorylation of Bora, and that only cytoplasmic Cyclin A2 interacts with Bora and PLK1. Expression of predominately cytoplasmic Cyclin A2 or phospho-mimicking PLK1 T210D can partially rescue a G2 arrest caused by Cyclin A2 depletion. Cytoplasmic presence of Cyclin A2 is restricted by p21, in particular after DNA damage. Cyclin A2 chromatin association during DNA replication and additional mechanisms contribute to Cyclin A2 localisation change in the G2 phase. We find no evidence that such mechanisms involve G2 feedback loops and suggest that cytoplasmic appearance of Cyclin A2 at the S/G2 transition functions as a trigger for mitotic kinase activation.

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Mitotic Kinases and p53 Signaling

Biochemistry Research International ◽

10.1155/2012/195903 ◽

2012 ◽

Vol 2012 ◽

pp. 1-14 ◽

Cited By ~ 7

Author(s):

Geun-Hyoung Ha ◽

Eun-Kyoung Yim Breuer

Keyword(s):

Molecular Mechanisms ◽

Kinase Inhibitors ◽

Kinase Signaling ◽

Tumor Suppressor P53 ◽

Mitotic Kinases ◽

Mitotic Kinase ◽

P53 Signaling ◽

Interactive Network ◽

Mutual Regulation ◽

And Function

Mitosis is tightly regulated and any errors in this process often lead to aneuploidy, genomic instability, and tumorigenesis. Deregulation of mitotic kinases is significantly associated with improper cell division and aneuploidy. Because of their importance during mitosis and the relevance to cancer, mitotic kinase signaling has been extensively studied over the past few decades and, as a result, several mitotic kinase inhibitors have been developed. Despite promising preclinical results, targeting mitotic kinases for cancer therapy faces numerous challenges, including safety and patient selection issues. Therefore, there is an urgent need to better understand the molecular mechanisms underlying mitotic kinase signaling and its interactive network. Increasing evidence suggests that tumor suppressor p53 functions at the center of the mitotic kinase signaling network. In response to mitotic spindle damage, multiple mitotic kinases phosphorylate p53 to either activate or deactivate p53-mediated signaling. p53 can also regulate the expression and function of mitotic kinases, suggesting the existence of a network of mutual regulation, which can be positive or negative, between mitotic kinases and p53 signaling. Therefore, deciphering this regulatory network will provide knowledge to overcome current limitations of targeting mitotic kinases and further improve the results of targeted therapy.

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Crystal structures of Magnaporthe oryzae trehalose-6-phosphate synthase (MoTps1) suggest a model for catalytic process of Tps1

Biochemical Journal ◽

10.1042/bcj20190289 ◽

2019 ◽

Vol 476 (21) ◽

pp. 3227-3240 ◽

Cited By ~ 1

Author(s):

Shanshan Wang ◽

Yanxiang Zhao ◽

Long Yi ◽

Minghe Shen ◽

Chao Wang ◽

...

Keyword(s):

Crystal Structures ◽

Conformational Changes ◽

Magnaporthe Oryzae ◽

Structural Information ◽

Complex Structure ◽

Critical Role ◽

Catalytic Process ◽

Structural Basis ◽

Salt Bridges ◽

Terminal Domain

Trehalose-6-phosphate (T6P) synthase (Tps1) catalyzes the formation of T6P from UDP-glucose (UDPG) (or GDPG, etc.) and glucose-6-phosphate (G6P), and structural basis of this process has not been well studied. MoTps1 (Magnaporthe oryzae Tps1) plays a critical role in carbon and nitrogen metabolism, but its structural information is unknown. Here we present the crystal structures of MoTps1 apo, binary (with UDPG) and ternary (with UDPG/G6P or UDP/T6P) complexes. MoTps1 consists of two modified Rossmann-fold domains and a catalytic center in-between. Unlike Escherichia coli OtsA (EcOtsA, the Tps1 of E. coli), MoTps1 exists as a mixture of monomer, dimer, and oligomer in solution. Inter-chain salt bridges, which are not fully conserved in EcOtsA, play primary roles in MoTps1 oligomerization. Binding of UDPG by MoTps1 C-terminal domain modifies the substrate pocket of MoTps1. In the MoTps1 ternary complex structure, UDP and T6P, the products of UDPG and G6P, are detected, and substantial conformational rearrangements of N-terminal domain, including structural reshuffling (β3–β4 loop to α0 helix) and movement of a ‘shift region' towards the catalytic centre, are observed. These conformational changes render MoTps1 to a ‘closed' state compared with its ‘open' state in apo or UDPG complex structures. By solving the EcOtsA apo structure, we confirmed that similar ligand binding induced conformational changes also exist in EcOtsA, although no structural reshuffling involved. Based on our research and previous studies, we present a model for the catalytic process of Tps1. Our research provides novel information on MoTps1, Tps1 family, and structure-based antifungal drug design.

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The Role of the Endothelium in Premature Atherosclerosis: Molecular Mechanisms

Current Medicinal Chemistry ◽

10.2174/0929867326666190911141951 ◽

2020 ◽

Vol 27 (7) ◽

pp. 1041-1051 ◽

Cited By ~ 1

Author(s):

Michael Spartalis ◽

Eleftherios Spartalis ◽

Antonios Athanasiou ◽

Stavroula A. Paschou ◽

Christos Kontogiannis ◽

...

Keyword(s):

Molecular Mechanisms ◽

Low Density Lipoprotein ◽

Critical Role ◽

Density Lipoprotein ◽

Number Of Genes ◽

Causes Of Mortality ◽

Artery Disease ◽

Genetic And Environmental Factors ◽

Made In

Atherosclerotic disease is still one of the leading causes of mortality. Atherosclerosis is a complex progressive and systematic artery disease that involves the intima of the large and middle artery vessels. The inflammation has a key role in the pathophysiological process of the disease and the infiltration of the intima from monocytes, macrophages and T-lymphocytes combined with endothelial dysfunction and accumulated oxidized low-density lipoprotein (LDL) are the main findings of atherogenesis. The development of atherosclerosis involves multiple genetic and environmental factors. Although a large number of genes, genetic polymorphisms, and susceptible loci have been identified in chromosomal regions associated with atherosclerosis, it is the epigenetic process that regulates the chromosomal organization and genetic expression that plays a critical role in the pathogenesis of atherosclerosis. Despite the positive progress made in understanding the pathogenesis of atherosclerosis, the knowledge about the disease remains scarce.

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