Michael Blower

Department of Genetics
Department of Molecular Biology
Massachusetts General Hospital
6622 Simches Research Building
185 Cambridge Street
Boston, MA 02115
Tel: 617-643-0777
Fax: 617-726-6893
Email: mblower@molbio.mgh.harvard.edu

We are interested in understanding both how and why RNA is targeted to microtubules during mitosis. We primarily use cell free extracts from the African clawed frog Xenopus laevis to study how RNA influences the assembly of microtubules into a dynamic bipolar spindle. Previous work has demonstrated that several proteins involved in RNA metabolism (Rae1, Maskin, and CPEB) are localized to the mitotic spindle and contribute to spindle assembly. Furthermore, we have shown that RNA is localized to mitotic microtubules in Xenopus egg extracts, and contributes to spindle assembly in a translation independent manner.

Our first approach is to identify specific mRNAs that are targeted to microtubules during mitosis. We have used Affymetrix gene chip microarrays to determine which mRNAs are specifically enriched on the mitotic spindle in both Xenopus and human cell extracts. We have found that a conserved set of mRNAs are localized to the microtubules during mitosis and that a subset of these mRNAs are actively translated on microtubules. Another set of MT-mRNAs appears to be present in inactive storage granules, and these mRNAs appear likely to regulate various aspects of development. While translation of RNA is not required for spindle assembly in egg extracts, our preliminary results suggest that local, mitotic translation of MT-mRNAs is likely to regulate various aspects of mitosis and cell cycle progression.

Our second approach is to identify all noncoding RNAs that associate with the mitotic spindle. We are using Affymetrix tilling microarrays to obtain an unbiased view of all RNAs associated with microtubules in Human cells. We are also generating miRNA libraries from both Xenopus and Human cells to determine if small noncoding RNAs influence spindle assembly or local gene translation.

Finally, we are using in vitro reconstitution to understand how RNA can influence microtubule assembly. We are using purified Rae1 and CPEB protein complexes in various in vitro assays to determine how the binding of RNA to these protein complexes influences their ability to nucleate, stabilize and bundle microtubules.

References:

• Blower, M.D., Nachury, M., Heald, R., and Weis, K.. A Rae1 containing ribonucleoprotein complex is required for mitotic spindle assembly. Cell. 2005 Apr 22;121(2):223-34.
• Brown, K.*, Blower, M.D.*, Maresca, T.*, Harland, R., Heald, R. (In Preparation) Use of Xenopus tropicalis egg extracts as a model system for the study of mitosis in vitro.
• Blower, M.D., Sullivan, B. A., and Karpen, G. H.. Conserved organization of centromeric chromatin in flies and humans. Dev Cell 2, 319-330.