Insights into Mad2 Regulation in the Spindle Checkpoint Revealed by the Crystal Structure of the Symmetric Mad2 Dimer (original) (raw)
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The Mad2 spindle checkpoint protein has two distinct natively folded states
Nature Structural & Molecular Biology, 2004
The spindle checkpoint delays chromosome segregation in response to misaligned sister chromatids during mitosis, thus ensuring the fidelity of chromosome inheritance. Through binding to Cdc20, the Mad2 spindle checkpoint protein inhibits the target of this checkpoint, the ubiquitin protein ligase APC/C Cdc20 . We now show that without cofactor binding or covalent modification Mad2 adopts two distinct folded conformations at equilibrium (termed N1-Mad2 and N2-Mad2). The structure of N2-Mad2 has been determined by NMR spectroscopy. N2-Mad2 is much more potent in APC/C inhibition. Overexpression of a Mad2 mutant that specifically sequesters N2-Mad2 partially blocks checkpoint signaling in living cells. The two Mad2 conformers interconvert slowly in vitro, but interconversion is accelerated by a fragment of Mad1, an upstream regulator of Mad2. Our results suggest that the unusual two-state behavior of Mad2 is critical for spindle checkpoint signaling.
The Mad2 spindle checkpoint protein has two distinct natively folded statesNSMB
NATURE STRUCTURAL & MOLECULAR BIOLOGY, 2004
The spindle checkpoint delays chromosome segregation in response to misaligned sister chromatids during mitosis, thus ensuring the fidelity of chromosome inheritance. Through binding to Cdc20, the Mad2 spindle checkpoint protein inhibits the target of this checkpoint, the ubiquitin protein ligase APC/CCdc20. We now show that without cofactor binding or covalent modification Mad2 adopts two distinct folded conformations at equilibrium (termed N1-Mad2 and N2-Mad2). The structure of N2-Mad2 has been determined by NMR spectroscopy. N2-Mad2 is much more potent in APC/C inhibition. Overexpression of a Mad2 mutant that specifically sequesters N2-Mad2 partially blocks checkpoint signaling in living cells. The two Mad2 conformers interconvert slowly in vitro, but interconversion is accelerated by a fragment of Mad1, an upstream regulator of Mad2. Our results suggest that the unusual two-state behavior of Mad2 is critical for spindle checkpoint signaling.
Current Computer Aided-Drug Design, 2014
In normal cells, the accuracy of chromosome segregation which assures cells euploidy depends on mitosis mechanics and on proper functioning of a specific complex of proteins represented by the error-checking spindle assembly checkpoint (SAC). SAC proteins are deeply involved in correct cell divisions, but some of these, such as mitotic arrest-deficient proteins (Mad1 and Mad2), are critical. Mad1 and Mad2 are involved in preventing "wrong" cellular divisions which lead to cellular aneuploidy and are recognized as inductors of genetic disorders, as well as activators of oncoproteins. To clarify aneuploidy involvement in the evolution of cancer or other genetic disorders, structural and functional specificity of spindle checkpoint proteins have been analyzed, but the process is still poorly understood. In order to better understand SAC proteins involvement in initiation of cancer and other genetic disorders, here we review studies that conducted to relevant structural and functional information regarding these proteins. The results of these studies suggest that minor changes in structure and functionality of SAC proteins are able to generate aneuploidy. Therefore, a deeper understanding of Mad1 and Mad2 structural changes obtained by experimental and theoretical studies could open new perspectives of genetic medicine.
Checkpoint Signalling: Mad2 Conformers and Signal Propagation
Current Biology, 2005
The spindle checkpoint is one of the key self-monitoring systems of the eukaryotic cell cycle, acting to delay anaphase until all the sister chromatids are appropriately lined up so that the replicated genome is correctly segregated with one copy going to each daughter cell. Mad2 is a central player in the spindle checkpoint's regulation of anaphase onset [1,2]. Mad2 interacts with Mad1, which recruits it to unattached kinetochores [3], and with the spindle checkpoint effector Cdc20 [4,5]. The latter interaction can take place either in a Mad2-Cdc20 complex or in the mitotic checkpoint complex (MCC):Mad2-Mad3/BubR1-Bub3-Cdc20 [6,7]. There is much controversy over the site(s) of assembly of these complexes, their mode of action and their importance as in vivo 'anaphase inhibitors'. Mad2 multimers have been studied for some time, although their physiological relevance was uncertain [8,9]. NMR and crystal structures of Mad2 alone or with binding peptides from Mad1 or Cdc20 have shown how the respective complexes are formed [10-13]. A 'safety-belt' structure formed by the carboxy-terminal tail of Mad2 explains why they are so stable [13].
Cancer Letters, 2019
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The Mad1/Mad2 Complex as a Template for Mad2 Activation in the Spindle Assembly Checkpoint
Current Biology, 2005
The SAC monitors this process and delays anaphase until all chromosomes have attained bipolar attachment [1,. Spindle microtubules attach on kinetochores, prothe tension between sister chromatids building up during this process [1, 2]. European Institute of Oncology Via Ripamonti 435 The SAC is conserved in all eukaryotes and includes mitotic arrest deficient (MAD) and budding uninhibited 20141 Milano Italy by benzimidazole (BUB) genes [1, 2]. Their products temporarily sequester Cdc20, an activator of the ana-Hill phase-promoting complex/cyclosome, the E3 ubiquitin ligase targeting securin and cyclin B for proteasome-607 Fordham Hall Chapel Hill, North Carolina 27599 mediated degradation. Destruction of securin activates separase, which triggers anaphase by cleaving the complex linking the sister chromatids, named Cohesin [4, 5]. The sequestration of Cdc20 requires Mad2 and Summary BubR1 [1, 6]. Mad2 is a 002ف residue protein containing a Horma domain [7]. BubR1 consist of an N-terminal Background: The spindle assembly checkpoint (SAC) domain containing Bub3 and Cdc20 binding sites and a imparts fidelity to chromosome segregation by delaying C-terminal kinase domain (missing in the budding yeast anaphase until all sister chromatid pairs have become ortholog, Mad3) [1, 6]. Both Mad2 and BubR1 bind bipolarly attached. Mad2 is a component of the SAC Cdc20 tightly, and their effects are synergic [8-13]. Coneffector complex that sequesters Cdc20 to halt anasistently, Mad2, BubR1 (or Mad3), Bub3, and Cdc20 phase. In prometaphase, Mad2 is recruited to kinetoenter a single complex known as mitotic checkpoint chores with the help of Mad1, and it is activated to bind complex (MCC) [11, 14-16]. Cdc20. These events are linked to the existence of two Unattached kinetochores establish and maintain the distinct conformers of Mad2: a closed conformer bound SAC, and all SAC proteins show kinetochore localization to its kinetochore receptor Mad1 or its target in the in prometaphase [1, 3, 17]. Fluorescence recovery after checkpoint Cdc20 and an open conformer unbound to photobleaching (FRAP) revealed stable kinetochore resthese ligands.