The abnormally high number of centrosomes found in many human tumor cells can lead directly to aneuploidy and genomic instability through the formation of multipolar mitotic spindles. To facilitate investigation of the mechanisms that control centrosome reproduction, a frog egg extract arrested in S phase of the cell cycle that supported repeated assembly of daughter centrosomes was developed. Multiple rounds of centrosome reproduction were blocked by selective inactivation of cyclin-dependent kinase 2-cyclin E (Cdk2-E) and were restored by addition of purified Cdk2-E. Confocal immunomicroscopy revealed that cyclin E was localized at the centrosome. These results demonstrate that Cdk2-E activity is required for centrosome duplication during S phase and suggest a mechanism that could coordinate centrosome reproduction with cycles of DNA synthesis and mitosis.

Phosphorylated sphingolipids [ceramide-1-phosphate (C1P) and sphingosine-1-phosphate (S1P)] have emerged as key regulators of cell growth, survival, migration, and inflammation1–5. C1P (Fig. 1a) produced by ceramide kinase is an activator of group IVA cytosolic phospholipase A2α (cPLA2α), the rate-limiting releaser of arachidonic acid used for pro-inflammatory eicosanoid production3,6–9, which contributes to disease pathogenesis in asthma/airway hyper-responsiveness, cancer, atherosclerosis, and thrombosis. To modulate eicosanoid action and avoid the damaging effects of chronic inflammation, cells require efficient targeting, trafficking, and presentation of C1P to specific cellular sites. Vesicular trafficking is likely10 but nonvesicular mechanisms for C1P sensing, transfer, and presentation remain unexplored11,12. Moreover, the molecular basis for selective recognition and binding among signaling lipids with phosphate headgroups, namely C1P, phosphatidic acid (PA) or their lyso-derivatives, remains unclear. Herein, an ubiquitously-expressed lipid transfer protein (CPTP) is shown to specifically transfer C1P between membranes. Crystal structures establish C1P binding via a novel surface-localized, phosphate headgroup recognition center connected to an interior hydrophobic pocket that adaptively expands to ensheath differing-length lipid chains using a cleft-like gating mechanism. The two-layer, α-helically-dominated ‘sandwich’ topology identifies CPTP as the prototype for a new GLTP-fold13 subfamily. CPTP resides in the cell cytosol but associates with the trans-Golgi/TGN, nucleus, and plasma membrane. RNAi-induced CPTP depletion elevates C1P steady-state levels and alters Golgi cisternae stack morphology. The resulting C1P decrease in plasma membranes and increase in the Golgi complex stimulates cPLA2α release of arachidonic acid, triggering pro-inflammatory eicosanoid generation.

Centrosomes were microsurgically removed from BSC-1 African green monkey kidney cells before the completion of S phase. Karyoplasts (acentrosomal cells) entered and completed mitosis. However, postmitotic karyoplasts arrested before S phase, whereas adjacent control cells divided repeatedly. Postmitotic karyoplasts assembled a microtubule-organizing center containing gamma-tubulin and pericentrin, but did not regenerate centrioles. These observations reveal the existence of an activity associated with core centrosomal structures-distinct from elements of the microtubule-organizing center-that is required for the somatic cell cycle to progress through G1 into S phase. Once the cell is in S phase, these core structures are not needed for the G2-M phase transition.

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