Proteomics of isolated mitotic chromosomes identifies the kinetochore protein Ska3/Rama1.

Despite many decades of study, mitotic chromosomes remain poorly characterized with respect to their structure and composition. Here, we have purified mitotic chromosomes from nocodazole-treated chicken DT40 cells. These chromosomes have a 0.7:1:1 ratio of nonhistone proteins to histones to DNA. They also contain a significant content of RNAs that have yet to be characterized. Overall, the isolated chromosomes contained >4000 polypeptides, >500 of which are either novel or uncharacterized. Elsewhere, we have developed an approach for comparing the results of multiple proteomics experiments. As a validation of this approach, one of 13 novel centromere proteins identified was found to occur in a complex with the previously described proteins Ska1 and Ska2. This novel protein, now known as Ska3/Rama1, occupies a unique domain in the outer kinetochore and was revealed by RNA interference (RNAi) experiments to be essential for cell cycle progression in human cells. The approach presented here offers a powerful way to define the functional proteome of complex organelles and structures whose composition is not simple or fixed.

Posttraining effects of amphetamine, chlorpromazine, ketamine, and scopolamine on the acquisition and extinction of the rabbit's conditioned nictitating membrane response.

Four experiments examined whether posttraining deliveries of drugs modified the performance of the rabbit's conditioned nictitating membrane response (NMR) during acquisition and extinction. The results show that ketamine accelerated, but that amphetamine, chlorpromazine, and scopolamine retarded, conditioning when the drugs were injected immediately after the completion of daily training sessions. However, all conditioning effects of the drugs were lost when their deliveries were delayed by 2 hr after the end of training. During extinction, the only drug that altered conditioned performance was ketamine. Specifically, ketamine retarded the rate of extinction when the drug was given immediately after training. However, delaying ketamine by 2 hr after training neutralized the drug's influence. These findings indicate that the NMR preparation should be useful in examining how the posttraining delivery of drugs influences associative processes in conditioning.