Annexin A2 is required for the early steps of cytokinesis

Cytokinesis requires the formation of an actomyosin contractile ring between the two sets of sister chromatids. Annexin A2 is a calcium- and phospholipid-binding protein implicated in cortical actin remodeling. We report that annexin A2 accumulates at the equatorial cortex at the onset of cytokinesis and depletion of annexin A2 results in cytokinetic failure, due to a defective cleavage furrow assembly. In the absence of annexin A2, the small GTPase RhoA—which regulates cortical cytoskeletal rearrangement—fails to form a compact ring at the equatorial plane. Furthermore, annexin A2 is required for cortical localization of the RhoGEF Ect2 and to maintain the association between the equatorial cortex and the central spindle. Our results demonstrate that annexin A2 is necessary in the early phase of cytokinesis. We propose that annexin A2 participates in central spindle–equatorial plasma membrane communication.     

A NAD-dependent glutamate dehydrogenase coordinates metabolism with cell division in Caulobacter crescentus

Coupling cell cycle with nutrient availability is a crucial process for all living cells. But how bacteria control cell division according to metabolic supplies remains poorly understood. Here, we describe a molecular mechanism that coordinates central metabolism with cell division in the α-proteobacterium Caulobacter crescentus. This mechanism involves the NAD-dependent glutamate dehydrogenase GdhZ and the oxidoreductase-like KidO. While enzymatically active GdhZ directly interferes with FtsZ polymerization by stimulating its GTPase activity, KidO bound to NADH destabilizes lateral interactions between FtsZ protofilaments. Both GdhZ and KidO share the same regulatory network to concomitantly stimulate the rapid disassembly of the Z-ring, necessary for the subsequent release of progeny cells. Thus, this mechanism illustrates how proteins initially dedicated to metabolism coordinate cell cycle progression with nutrient availability.     

Dynein LIC1 localizes to the mitotic spindle and midbody and LIC2 localizes to spindle poles during cell division

CD-1 (cytoplasmic dynein-1) is a multisubunit motor protein complex involved in intracellular trafficking and mitosis. The dynein LIC (light intermediate chain) subunits, LIC1 (DLIC-1, gene symbol DYNC1LI1) and LIC2 (DLIC-2, gene symbol DYNC1LI2), associate with the dynein HC (heavy chain) in a mutually exclusive manner and thus define distinct functional CD-1 complexes. Here, we analysed the mitotic distribution of LIC1 and LIC2. We found that from metaphase through anaphase, LIC1 localizes to the mitotic spindle and concentrates within the midbody during the abscission step of cytokinesis. Conversely, LIC2 strongly localizes to the spindle poles from prophase through telophase. These data suggest distinct functions for LIC1 and LIC2-containing CD-1 complexes during cell division.     

Novel chromosomal translocation t(11;9)(p15;p23) involving deletion and duplication of 9p in a girl associated with autism and mental retardation

We describe a 5-year-old girl presented with autism and mental retardation features. Conventional karyotyping revealed a novel unidirectional translocation t(11;9)(p15;p23). HumanCytoSNP-12 Chip analysis identified a 13 Mb deletion from 9p24.3 to 9p23 and a 12.5Mb duplication from 9p23 to 9p21.2. The karyotype was described as 45,XX,psu dic(11; 9)(p15;p23), which was reported for the first time here. The deleted region, extending from 9p24.3 to 9p23, overlaps with the candidate region for monosomy 9p syndrome and contains a potential autism spectrum disorders (ASD) locus. The duplication region extending from 9p23 to 9p21.2 was previously identified as a critical region for the 9p duplication syndrome. These results suggested that the apparently balanced de novo translocations could produce cryptic deletions or duplications, and the precise mapping of the abnormal area may improve clinical management.     

ARF6 GTPase protects the post-mitotic midbody from 14-3-3-mediated disintegration

In cytokinesis, there is a lengthy interval between cleavage furrow ingression and abscission, during which the midbody microtubule bundle provides both structural support for a narrow intercellular bridge and a platform that orchestrates the biochemical preparations for abscission. It is currently unclear how the midbody structure is stably maintained during this period. Here, we report a novel role for the ADP-ribosylation factor 6 (ARF6) GTPase in the post-mitotic stabilisation of midbody. Centralspindlin kinesin-6/RhoGAP complex, a midbody component critical for both the formation and function of the midbody, assembles in a sharp band at the centre of the structure in a manner antagonised by 14-3-3 protein. We show that ARF6 competes with 14-3-3 for binding to centralspindlin such that midbodies formed by centralspindlin mutants that can bind 14-3-3 but not ARF6 frequently collapse before abscission. These data indicate a novel mechanism for the regulation of midbody dynamics in which ARF6 protects the compacted centralspindlin assembly from dissipation by 14-3-3.     

N-Terminus-Mediated Solution Structure of Dimerization Domain of PRC1

Simple SummaryThe solution structure of the N-terminal domain of Protein Regulator of Cytokinesis 1 (PRC1) was determined, and compared with the previously published crystal structure, significant differences were found. Extensive analyses were carried out to find the true reason for the differences between the solution and crystal structures, we discovered that this might be related to the conformation of residue M1, which is buried in the protein core of the solution structure, while situated outside of the hydrophobic core in the crystal structure. In this study, we have carried out a series of examinations using various methods and confirmed that the N terminal conformation is the key point in better describing the structure of PRC1 dimerization domain under solution conditions.AbstractMicrotubule-associated proteins (MAPs) are essential for the accurate division of a cell into two daughter cells. These proteins target specific microtubules to be incorporated into the spindle midzone, which comprises a special array of microtubules that initiate cytokinesis during anaphase. A representative member of the MAPs is Protein Regulator of Cytokinesis 1 (PRC1), which self-multimerizes to cross-link microtubules, the malfunction of which might result in cancerous cells. The importance of PRC1 multimerization makes it a popular target for structural studies. The available crystal structure of PRC1 has low resolution (>3 Å) and accuracy, limiting a better understanding of the structure-related functions of PRC1. Therefore, we used NMR spectroscopy to better determine the structure of the dimerization domain of PRC1. The NMR structure shows that the PRC1 N terminus is crucial to the overall structure integrity, but the crystal structure bespeaks otherwise. We systematically addressed the role of the N terminus by generating a series of mutants in which N-terminal residues methionine (Met1) and arginine (Arg2) were either deleted, extended or substituted with other rationally selected amino acids. Each mutant was subsequently analyzed by NMR spectroscopy or fluorescence thermal shift assays for its structural or thermal stability; we found that N-terminal perturbations indeed affected the overall protein structure and that the solution structure better reflects the conformation of PRC1 under solution conditions. These results reveal that the structure of PRC1 is governed by its N terminus through hydrophobic interactions with other core residues, such hitherto unidentified N-terminal conformations might shed light on the structure–function relationships of PRC1 or other proteins. Therefore, our study is of major importance in terms of identifying a novel structural feature and can further the progress of protein folding and protein engineering.     

KLIF-associated cytoskeletal proteins in Trypanosoma brucei regulate cytokinesis by promoting cleavage furrow positioning and ingression

Cytokinesis in the early divergent protozoan Trypanosoma brucei occurs from the anterior cell tip of the new-flagellum daughter toward the nascent posterior end of the old-flagellum daughter of a dividing biflagellated cell. The cleavage furrow ingresses unidirectionally along the preformed cell division fold and is regulated by an orphan kinesin named kinesin localized to the ingressing furrow (KLIF) that localizes to the leading edge of the ingressing furrow. Little is known about how furrow ingression is controlled by KLIF and whether KLIF interacts with and cooperates with other cytokinesis regulatory proteins to promote furrow ingression. Here, we investigated the roles of KLIF in cleavage furrow ingression and identified a cohort of KLIF-associated cytoskeletal proteins as essential cytokinesis regulators. By genetic complementation, we demonstrated the requirement of the kinesin motor activity, but not the putative tropomyosin domain, of KLIF in promoting furrow ingression. We further showed that depletion of KLIF impaired the resolution of the nascent posterior of the old-flagellar daughter cell, thereby stalking cleavage furrow ingression at late stages of cytokinesis. Through proximity biotinylation, we identified a subset of cytoskeleton-associated proteins (CAPs) as KLIF-proximal proteins, and functional characterization of these cytoskeletal proteins revealed the essential roles of CAP46 and CAP52 in positioning the cleavage furrow and the crucial roles of CAP42 and CAP50 in promoting cleavage furrow ingression. Together, these results identified multiple cytoskeletal proteins as cytokinesis regulators and uncovered their essential and distinct roles in cytokinesis.     

Deregulated ERK1/2 MAP kinase signaling promotes aneuploidy by a Fbxw7β-Aurora A pathway

Aneuploidy is a common feature of human solid tumors and is often associated with poor prognosis. There is growing evidence that oncogenic signaling pathways, which are universally dysregulated in cancer, contribute to the promotion of aneuploidy. However, the mechanisms connecting signaling pathways to the execution of mitosis and cytokinesis are not well understood. Here, we show that hyperactivation of the ERK1/2 MAP kinase pathway in epithelial cells impairs cytokinesis, leading to polyploidization and aneuploidy. Mechanistically, deregulated ERK1/2 signaling specifically downregulates expression of the F-box protein Fbxw7β, a substrate-binding subunit of the SCF^Fbxw7 ubiquitin ligase, resulting in the accumulation of the mitotic kinase Aurora A. Reduction of Aurora A levels by RNA interference or pharmacological inhibition of MEK1/2 reverts the defect in cytokinesis and decreases the frequency of abnormal cell divisions induced by oncogenic H-Ras^V12. Reciprocally, overexpression of Aurora A or silencing of Fbxw7β phenocopies the effect of H-Ras^V12 on cell division. In vivo, conditional activation of MEK2 in the mouse intestine lowers Fbxw7β expression, resulting in the accumulation of cells with enlarged nuclei. We propose that the ERK1/2/ Fbxw7β/Aurora A axis identified in this study contributes to genomic instability and tumor progression.     

RhoA-mediated MLC2 regulates actin dynamics for cytokinesis in meiosis

During oocyte meiosis, the bipolar spindle forms in the central cytoplasm and then migrates to the cortex. Subsequently, the oocyte extrudes the polar body through two successive asymmetric divisions, which are regulated primarily by actin filaments. Myosin light chain2 (MLC2) phosphorylation plays pivotal roles in smooth muscle contraction, stress fiber formation, cell motility and cytokinesis. However, whether MLC2 phosphorylation participates in the oocyte polarization and asymmetric division has not been clarified. The present study investigated the expression and functions of MLC2 during mouse oocyte meiosis. Our result showed that p-MLC2 was localized in the oocyte cortex, with a thickened cap above the chromosomes. Meanwhile, p-MLC2 was also localized in the poles of spindle. Disruption of MLC2 activity by MLC2 knock down (K_D) caused the failure of polar body extrusion. Immunofluorescent staining showed that a large proportion of oocytes arrested in telophase stage and failed to undergo cytokinesis after culturing for 12 hours. In the meantime, actin filament staining at oocyte membrane and cytoplasm were reduced in MLC2 K_D oocytes. Finally, we found that the phosphorylation of MLC2 protein levels was decreased after disruption of RhoA activity. Above all, our data indicated that the RhoA-mediated MLC2 regulates the actin organization for cytokinesis during mouse oocyte maturation.