Visualization and Orchestration of the Dynamic Molecular Society in Cells

Visualization and orchestration of the dynamic molecular society in cells

Xuebiao Yao1, Guowei Fang2

1Division of Cellular Dynamics, Hefei National Laboratory for Physical Sciences at Nano-scale, and the University of Science & Technology of China, Hefei 230027, China; 2Department of Biological Sciences, Stanford University, Stanford, CA 94305, USA Cell Research (2009) 19:152-155. doi: 10.1038/cr.2009.7; published online 20 January 2009

Visualization of specific molecules Lippincott-Schwartz (National In- underlying mitotic checkpoint. Andrea and their interactions in real space stitutes of Health, USA) opened the Musacchio of European Institute of and time is essential to delineate how meeting with a report on the latest supra- Oncology, Italy, has recently succeeded cellular plasticity and dynamics are resolution imaging technique named in determining the crystal structure of a achieved and orchestrated as perturba- PALM (photo-activated localization truncated “bonsai” version of Ndc80, a tion of cellular plasticity and dynamics microscopy) [1]. PALM enables opti- tetrameric complex essential for accu- is detrimental to health. Elucidation of cally imaging intracellular proteins at rate spindle microtubule-chromosome cellular dynamics requires molecular nanometer (2~25 nm) spatial resolution attachment [3]. Xin Cai from Xuebiao imaging at nanometer scale at milli- by isolation of single molecules at high Yao’s laboratory (USTC, China) pre- second resolution. The 1st International densities (up to ~105/mm2) based on the sented supporting evidence for the Conference on Cellular Dynamics and serial photo-activation and subsequent function of Ndc80 in accurate spindle- Chemical Biology held in Hefei, China bleaching of numerous sparse subsets microtubule attachment regulated by (from 12 September to 15 September, of photo-activatable fluorescent protein Nek2A-mediated phosphorylation 2008) launched the quest by bringing molecules within a sample. By combin- [4]. Andrea Musacchio also presented synergism among photonics, chemistry ing PALM with single-particle track- his working model accounting for the and biology. ing, the same group developed a new Mad2-dependent spindle checkpoint, The conference was held in conjunc- method termed sptPALM [2], which in which he proposed that a confor- tion with the 50th anniversary of the enables the recording of several orders mational switch of Mad2 mediates its University of Science & Technology of magnitude more trajectories per cell binding and inhibitory activities towards of China (USTC). About 100 scientists than traditional single-particle tracking the Cdc20 activator of the anaphase and students gathered in Hefei to dis- allows. By probing distinct subsets of promoting complex/cyclosome [5]. cuss the recent advances in molecular molecules, sptPALM can resolve the Cytoplasmic dynein is a minus-end imaging and chemical manipulation of dynamics and origins of spatial and directed microtubule-based molecular cell biology. temporal molecular heterogeneities in motor and plays multiple functions in cells. mitosis. Xueliang Zhu (Shanghai In- Seeing is believing stitutes for Biological Sciences, CAS, Cell division and growth signal- China) presented his recent finding in The inherent resolution limit of the ling which NudE/Nudel (NudE-like) and light microscope has been a hurdle Lis1 might serve as novel positive for visualization of molecular nano- During cell division, a super-protein regulators of dynein in eukaryotes machineries in cell biology. Jennifer structure called kinetochore orches- [6]. Both the Nudel-Lis1 and Nudel- trates the equal segregation of parental dynein interactions are crucial for dy- genome into two daughter cells. One nein activity. He proposed that Nudel of the biggest challenges in the mitosis functions not only in dynein-mediated Correspondence: Xuebiao Yaoa, Guowei Fangb aE-mail: [email protected] field is to illustrate the kinetochore as- protein transport, but also in stabilizing bE-mail: [email protected] sembly dynamics and signaling cascade kinetochore association with dynein

Cell Research | Vol 19 No 2 | February npg 153 to facilitate dynein-mediated chromo- molecular interaction [12]. Plk1 inter- principles underlying the networking some poleward movement via tethering acts with and phosphorylates Cdc14A, of such interactions. This requires an to mitosin/CENP-F. To delineate the which releases the intra-molecular inter- accurate understanding of their points molecular composition and interaction action of Cdc14A. Using a combination of contact. Yunyu Shi (USTC, China) networks of the mammalian kineto- of Plk1 activity reporter and a sensor presented her studies on human PPIL1 chore, Xuebiao Yao’s laboratory carried to monitor Cdc14A intra-molecular (peptidyl prolyl isomerase-like protein out a combination of proteomics and interaction, Yuan is profiling the spa- 1), a component in 45S U5 snRNP systemic yeast 2-hybrid analyses. They tiotemporal dynamics of Cdc14A-Plk1 in activated spliceosome B* and 35S revealed the role of Nek2A kinase in interaction in real-time chromosome snRNP. It was believed to participate orchestrating kinetochore-microtubule segregation. in the activation of spliceosome. Us- attachments and mitotic checkpoint [7, TGF-b signalling plays pivotal ing NMR spectroscopic analysis, the 8]. Recent studies from Yao and col- roles in embryogenesis and cancer solution structure of PPIL1 was de- leagues discovered how Nuf2 tethers development. This signalling path- termined and shown to resemble that CENP-E for the kinetochore localiza- way is spatiotemporally regulated to of other members of the cyclophilin tion [9], and how septin 7 filaments orchestrate cellular dynamics and ho- family [17]. Chemical shift perturbation enable the accurate translocation of meostasis. Ye-Guang Chen (Tsinghua experiments were employed to study CENP-E to the kinetochore [10]. Yao University, China) presented studies the interaction between PPIL1 and also presented a molecular dissection on several newly identified proteins SKIP (Ski-interaction protein), another of the CENP-E interaction network in implicated in TGF-b signalling [13, essential component of 45S activated mitotic checkpoint signalling. 14]. He showed that Dapper2 negatively spliceosome. Their studies illustrated One central question in the cell cycle regulates TGF-b/Nodal signaling by that a novel cyclophilin-protein contact field is what controls the phase transi- influencing lysosomal degradation of module exists in the PPIL1-SKIP com- tions? Although the accumulation of the endocytosed TGF-b/Nodal recep- plex, suggesting that protein-protein mitotic cyclins drives the transition tors while endofin, a member of the contact-mediated conformation changes from the G2 phase to the M phase in FYVE domain protein family, functions may function in activation of the spli- embryonic cells, the trigger for mitotic in endosomal trafficking. Preliminary ceosome. entry in somatic cells remains unknown. evidence suggests that membrane lipid- Reversible protein tyrosine phos- Guowei Fang (Stanford University, rafts participate in TGF-b signalling phorylation plays a major role in cel- USA) reported his recent finding that dynamics by determining the activation lular signalling by changing protein’s Bora synergizes with the Aurora A ki- of MAPK by TGF-b [15]. Systemic plasticity. However, delineation of nase in controlling the G2-M transition acquired resistance (SAR) is a plant the functional significance of PTPs [11]. Bora accumulates in the G2 phase immune response that can be induced in normal physiology and in diseases and promotes Aurora A-mediated acti- after a local infection. Using SAR as remains a major challenge in cell sig- vation of Plk1, leading to the activation a model system, Xinnian Dong (Duke naling as genetic manipulations such of cyclin-dependent kinase 1 and mitot- University, USA) showed that Arabi- as gene knockout and RNAi can not ic entry. Mechanistically, Bora interacts dopsis NPR1 protein is an essential resolve the involvement of a protein with Plk1 and controls the accessibility signaling component of SAR. NPR1 is in two temporally separated events. of its activation loop for phosphoryla- sequestered in the cytoplasm as an oli- Zhong-yin Zhang (Indiana University, tion and activation by Aurora A. While gomer through intermolecular disulfide USA) described how chemical biology chromosome segregation in mitosis bonds and S-nitrosylation of NPR1 by approaches with potent and selective is orchestrated by protein kinase- and S-nitrosoglutathione at cysteine-156 small molecule inhibitors can be used to phosphatase-signaling cascades, early facilitates its oligomerization, suggest- delineate PTP function and to develop studies showed that overexpression of ing a link between pathogen-triggered PTP-based therapeutics [e.g., 18]. An- human phosphatase Cdc14A, an an- redox changes and gene regulation in notation of the budding yeast Saccharo- tagonist of cyclin-dependent kinase 1, plant immunity [16]. myces cerevisiae genome revealed that affects several aspects of cell division. there are 80 genes involved in oxidative However, the molecular mechanism Structure and activity stress response. Using a structural ge- underlying HsCdc14A regulation in nomics approach, Congzhao Zhou and mitosis has remained elusive. Kai Yuan One of our current challenges in colleagues (USTC, China) revealed of Xuebiao Yao’s laboratory showed understanding cellular dynamics is to the structural basis of protein-mediated that Cdc14A activity is regulated by an delineate the structural modules medi- electron transfer in response to oxidative auto-inhibitory mechanism via intra- ating protein-protein interactions and stress [19]. The 2b proteins encoded by www.cell-research.com | Cell Research npg 154 cucumovirus act as post-transcriptional elles and long-lived proteins. Reduc- key molecules in health and diseases, gene silencing suppressors to counter tion of autophagy activity has been he stressed the importance of allosteric host defence during infection. Adma shown to lead to the accumulation of modulators in harnessing disease-asso- Yuan (Singapore National University, misfolded proteins in disease settings. ciated target activity. Singapore) reported the crystal struc- Dawei Ma and colleagues (Shanghai ture of 2b protein complexed with a 19 Institute of Organic Chemistry, CAS, Cellular plasticity and manipula- bp small interfering RNA duplex [20]. China) recently discovered some small tion The 2b protein oligomerizes to form a molecule regulators of autophagy us- Glioblastomas display a hierarchy dimer of dimers through the conserved ing an image-based screening strategy of neoplastic cells with a restricted leucine-zipper-like motif at its amino- [22]. Interestingly, these compounds can population of cells that exhibit ex- terminal alpha-helix. promote degradation of long-lived pro- tensive self-renewal and the capac- teins and reduce the levels of expanded ity to generate complex tumors that Chemical biology of enzymes polyglutamine repeats in cultured cells, recapitulate parental tumours upon raising the possibility that some of these xenotransplantation. Xiao-Fan Wang The family of 14-3-3 proteins func- drugs may be useful for the treatment (Duke University, USA) presented an tions as critical regulators underlying of human diseases associated with the unexpected excitement in which N-myc diverse signaling events in health and accumulation of misfolded proteins. downstream-regulated gene 4 (Ndrg4) is diseases. Given the fact that 14-3-3- During cell division, chromosome critical for the survival of glioblastoma binding often switches the plastic- segregation is orchestrated by the in- cells. Knockdown of Ndrg4 induces ity of its binding partners, Haian Fu teraction of spindle microtubules with an increase in apoptosis, indicating (Emory University, USA) devised a the centromere. A dramatic remodel- that Ndrg4 diverges in function from high-throughput screen strategy for ing of interpolar microtubules into an its closely related tumour suppressive identifying chemicals that disrupt the organized central spindle between the family members in the context of glio- association of 14-3-3 with its binding separating chromatids is required for the blastoma. partners and carried out a large scale initiation and execution of cytokinesis. Cell-based phenotypic and pathway- screening campaign [21]. Since 14-3-3 It becomes increasingly clear that a co- specific screens of synthetic small is up-regulated in certain type of lung hort of kinesin molecules cooperate to molecules and natural products have cancers, these 14-3-3 inhibitors are of orchestrate the spindle dynamics in mi- historically provided useful chemical substantial significance in dissecting tosis. To dissect the spatiotemporal dy- ligands to modulate and/or study com- spatiotemporal dynamics of 14-3-3 and namics, Xuebiao Yao and colleagues set plex cellular processes. Sheng Ding and are also expected to be of great benefit up a combination of phenotype-based colleagues (Scripps Research Institute, in leading to therapeutic strategies for screen for mis-aligned chromosomes USA) discovered small molecules that treating diseases such as cancer. in mitosis and inhibition of CENP-E can be used to selectively regulate stem Polo and Aurora family kinases activity in vitro. Treatment of these cell fate and developmental signalling regulate important events during mitosis novel CENP-E inhibitors resulted in pathways [23, 24]. Such molecules will including centrosome maturation and chromosome alignment errors, which is provide useful tools to dissect stem cell separation, mitotic spindle assembly, reminiscent of siRNA-mediated knock- plasticity, and may ultimately contrib- and chromosome segregation. Mis- down of CENP-E. The excitement ute to regenerative medicine for tissue regulation of Aurora and Polo kinases ahead is to use these valuable chemical repair. Reverberatory activity within due to genetic amplification and protein tools to dissect the temporal function of neuronal cell assembly is thought to overexpression results in aneuploidy CENP-E in mitosis. represent the online memory trace in and may contribute to tumorigenesis. Kaixian Chen (Shanghai Institutes the brain and to serve as an elementary Therefore, chemical inhibitors of Au- for Biological Sciences, CAS, China) unit in thought process. Using cultured rora and Polo are currently undergoing exemplified the case study of applica- hippocampal neurons as a model phase 2 clinical trials. Jiancun Zhang tion of chemical biology strategies for system, Guoqiang Bi and colleagues (Guangzhou Institute of Biomedicine pharmacological lead identification (USTC, China) revealed that activity- and Health, CAS, China) provided his and development of traditional Chinese induced synaptic plasticity could drive rationale design of allosteric inhibitors medicine. He elaborated how molecu- the emergence of reverberatory activity for mitotic regulators such as Mps1. lar dynamics and network information in small networks [25]. It would be of Autophagy is a lysosome-dependent can be used for intervening disease great interest to illuminate the molecu- cellular catabolic mechanism underly- development and progression. Given lar regulatory network underlying the ing the turnover of intracellular organ- the differential involvement of certain reverberatory activity.

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