Biogenesis; Genetics; Molecular Biology; Ribonucleoproteins; Radiation Oncology; RNA Helicases; Genes, rRNA; Biochemical Processes
Therapeutic Radiology: Radiobiology
Of Yeast and Ribosome Biogenesis: Ribosome biogenesis is a complex process requiring the coordinated expression of rRNA and protein moieties and their assembly in the eukaryotic nucleolus. In order to better understand each aspect of this process, we are using an array of genetic, biochemical, and cell biological techniques in the yeast Saccharomyces cerevisiae. My laboratory focuses on the role of the ribonucleoprotein and protein complexes involved in generating the mature rRNAs.
Specialized Terms: Ribosome biogenesis; RRNA processing; U3 RNP structure and function; RNA helicases; Polymerase I transcription and processing
Extensive Research Description
Study of RNA helicases required for ribosome biogenesis and their cofactors
investigations into the role of ribosome biogenesis in cell cycle regulation
discovery of a subset of SSU processome proteins that are associated with the rDNA and are required for rDNA transcription
subcomplexes of the SSU processome and deciphering the direct
protein-protein and protein-RNA interactions that mediate their assembly
purification and electron microscopy to visualize pre-ribosomes
characterization of an essential new protein-protein interaction motif found in RNA processing RNPs
developing a method to identify individual proteins in chromatin spreads.
Using innovative proteomics techniques, my laboratory has recently identified the protein components of a large nucleolar ribonucleoprotein that is required for processing of the 18S small subunit rRNA. This RNP, which we termed the SSU processome, is composed of the U3 snoRNA and 40 proteins. Currently, projects in the lab are aimed at determining the architecture of this RNP and the functions of individual proteins in 18S processing. We approach this question from several perspectives, using genetic and biochemical methods to identify direct interactions between components, and cryo electron microscopy to visualize the complex in three dimensions.
Through these studies we have discovered and characterized several unique protein motifs and their specific roles in rRNA processing. We have recently discovered that a subset of the SSU processome proteins are associated with the rDNA and are required for rDNA transcription. Stemming from this idea, we are interested in studying the proteins which regulate transcription of the rDNA by Pol I and initiate the processing of the rRNA. We have learned that these steps are intimately linked, and endeavor to describe this complex process in detail. Seventeen putative RNA helicases have been shown to be required for processing of the small and large ribosomal subunit RNAs, perhaps by remodeling the rRNA to allow access to cleavage sites. Ongoing genetic and biochemical studies in the lab examine the roles of each putative RNA helicase and test its ability to unwind RNA. Through these projects, we strive to ascertain how and why the helicases are required at each step in ribosome biogenesis. Because ribosomes are essential to cell growth via the production of new proteins, we are studying the role of ribosome biogenesis in cell cycle regulation.
We have previously shown that rRNA maturation by the SSU processome is required for cell cycle progression, indicating that the production of ribosomes has a distinct influence on the cell cycle. Specifically, we seek to find the ribosome-regulated trigger that allows the cell to progress through the cell cycle, grow in size, and divide. Transcription of the rDNA and processing of the rRNA can be visualized in Miller chromatin spreads, as shown here. In a, the SSU processome corresponds to the terminal knobs at the end of each rRNA branching off the rDNA. When components of the SSU processome are depleted (the U3 snoRNA in b, or the Utp7 protein in c), the knobs are no longer present, due to incomplete formation of the SSU processome.
- McCann, K.L., Charette, J.M., Vincent, N.G., and Baserga, S.J. A protein interaction map of the LSU processome. Genes Dev. (2015) 29: 862-875. PMC4403261
- McCann, K.L., Teramoto, T., Zhang, J., Hall, T.M.T.*, Baserga, S.J.* The molecular basis for ANE syndrome revealed by the large ribosomal subunit processome interactome. eLife (2016) 5:e16381. *co-corresponding authors. PMCID: PMC4859800
- Yip, W.S.V., Shigematsu, H., Taylor, D.W. and Baserga, S.J. Box C/D sRNA stem ends act as stabilizing anchors for box C/D di-sRNPs. Nucleic Acids Research, in press.
- Zhang, J., McCann, K.L., Qiu, C., Gonzalez, L.E., Baserga, S.J.*, and Hall, T.M.T.* Nop9 is a PUF-like protein that prevents premature cleavage to correctly process pre-18S rRNA. Nature Communications, in press. *co-corresponding authors.
- Tao, T.*, Sondalle, S.B.*, Shi, H.*, Zhu, S., Perez-Atayde, A.R., Peng, J., Baserga, S.J#., and Look, A.T#. The pre-rRNA processing factor DEF is rate limiting for the pathogenesis of MYCN-driven neuroblastoma. submitted to Oncogene. *These authors contributed equally to this work. #co-corresponding authors.
- Robson, A., Owens, N.D.L., Baserga, S.J., Khokha, M.K., and Griffin, J.N. Expression of ribosomopathy genes during Xenopus tropicalis embryogenesis. In press, BMC Developmental Biology
- Farley, K.I., Surovtseva, Y., Merkel, J. and Baserga, S.J. Determinants of mammalian nucleolar architecture. (2015) Chromosoma 124: 323-331, PMCID: PMC4534358. Review
- Sondalle, S.B., Baserga, S.J.* and Yelick, P.C.* The contributions of the ribosome biogenesis protein, Utp5/WDR43, to craniofacial development. Journal of Dental Research, in press. *co-corresponding authors. Review
- Farley, K.I. and Baserga, S.J. Probing the mechanisms underlying human diseases in making ribosomes. Biochemical Society Transactions, in press. Review