Prime movers : mechanochemistry of mitotic kinesins

Mitotic spindles are self-organizing protein machines that harness teams of multiple force generators to drive chromosome segregation. Kinesins are key members of these force-generating teams. Different kinesins walk directionally along dynamic microtubules, anchor, crosslink, align and sort microtubules into polarized bundles, and influence microtubule dynamics by interacting with microtubule tips. The mechanochemical mechanisms of these kinesins are specialized to enable each type to make a specific contribution to spindle self-organization and chromosome segregation

Mammalian Kinesin-3 Motors Are Dimeric In Vivo and Move by Processive Motility upon Release of Autoinhibition

Kinesin-3 motors drive the transport of synaptic vesicles and other membrane-bound organelles in neuronal cells. In the absence of cargo, kinesin motors are kept inactive to prevent motility and ATP hydrolysis. Current models state that the Kinesin-3 motor KIF1A is monomeric in the inactive state and that activation results from concentration-driven dimerization on the cargo membrane. To test this model, we have examined the activity and dimerization state of KIF1A. Unexpectedly, we found that both native and expressed proteins are dimeric in the inactive state. Thus, KIF1A motors are not activated by cargo-induced dimerization. Rather, we show that KIF1A motors are autoinhibited by two distinct inhibitory mechanisms, suggesting a simple model for activation of dimeric KIF1A motors by cargo binding. Successive truncations result in monomeric and dimeric motors that can undergo one-dimensional diffusion along the microtubule lattice. However, only dimeric motors undergo ATP-dependent processive motility. Thus, KIF1A may be uniquely suited to use both diffuse and processive motility to drive long-distance transport in neuronal cells

B-Cyclin/CDKs Regulate Mitotic Spindle Assembly by Phosphorylating Kinesins-5 in Budding Yeast

Although it has been known for many years that B-cyclin/CDK complexes regulate the assembly of the mitotic spindle and entry into mitosis, the full complement of relevant CDK targets has not been identified. It has previously been shown in a variety of model systems that B-type cyclin/CDK complexes, kinesin-5 motors, and the SCFCdc4 ubiquitin ligase are required for the separation of spindle poles and assembly of a bipolar spindle. It has been suggested that, in budding yeast, B-type cyclin/CDK (Clb/Cdc28) complexes promote spindle pole separation by inhibiting the degradation of the kinesins-5 Kip1 and Cin8 by the anaphase-promoting complex (APCCdh1). We have determined, however, that the Kip1 and Cin8 proteins are present at wild-type levels in the absence of Clb/Cdc28 kinase activity. Here, we show that Kip1 and Cin8 are in vitro targets of Clb2/Cdc28 and that the mutation of conserved CDK phosphorylation sites on Kip1 inhibits spindle pole separation without affecting the protein's in vivo localization or abundance. Mass spectrometry analysis confirms that two CDK sites in the tail domain of Kip1 are phosphorylated in vivo. In addition, we have determined that Sic1, a Clb/Cdc28-specific inhibitor, is the SCFCdc4 target that inhibits spindle pole separation in cells lacking functional Cdc4. Based on these findings, we propose that Clb/Cdc28 drives spindle pole separation by direct phosphorylation of kinesin-5 motors

Polo kinase recruitment via the constitutive centromere-associated network at the kinetochore elevates centromeric RNA

The kinetochore, a multi-protein complex assembled on centromeres, is essential to segregate chromosomes during cell division. Deficiencies in kinetochore function can lead to chromosomal instability and aneuploidy-a hallmark of cancer cells. Kinetochore function is controlled by recruitment of regulatory proteins, many of which have been documented, however their function often remains uncharacterized and many are yet to be identified. To identify candidates of kinetochore regulation we used a proteome-wide protein association strategy in budding yeast and detected many proteins that are involved in post-translational modifications such as kinases, phosphatases and histone modifiers. We focused on the Polo-like kinase, Cdc5, and interrogated which cellular components were sensitive to constitutive Cdc5 localization. The kinetochore is particularly sensitive to constitutive Cdc5 kinase activity. Targeting Cdc5 to different kinetochore subcomplexes produced diverse phenotypes, consistent with multiple distinct functions at the kinetochore. We show that targeting Cdc5 to the inner kinetochore, the constitutive centromere-associated network (CCAN), increases the levels of centromeric RNA via an SPT4 dependent mechanism

Etude des sites de fixation des métaux dans les sédiments. Apport des méthodes d'attaques sélectives

L'utilisation d'un schéma d'attaques sélectives permet de déterminer les sites de fixation du chrome, et plomb sur un sédiment d'étang saumâtre utilisé en thérapeutique thermale. Ce matériau est surchargé en chrome, cuivre et plomb. La comparaison des teneurs métalliques du sédiment avant et après surcharge conduit à une discussion sur sites de fixation et sur l'origine de ces trois métaux

Phosphorylation relieves autoinhibition of the kinetochore motor Cenp-E

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