Ribosomes catalyze protein synthesis in all cells. Consequently, mature and assembling ribosomes are the target for many antibiotics. Ribosome assembly defects also underlie many serious human diseases. Thus, regulation and ensuing quality control in ribosome assembly, as in other steps of translation, is of key importance. The goal of this proposal is to uncover the basis for ordered assembly steps, which likely form the basis for quality control, and to dissect the function of three regulatory proteins during the final assembly step of the 40S ribosomal subunit in yeast. The step under investigation is relatively simple and well-defined, involving two changes in the primary sequence of the RNA. Yet, it requires at least nine accessory factors, which are essential in yeast. Some of these factors have activities typically considered regulatory, suggesting that this step is simple enough for mechanistic dissection, yet complex enough to yield important insight into RNA-protein complex (RNP) assembly. Since ribosomes are the most conserved and ancient RNPs, these insights are expected to be of fundamental importance. This proposal addresses the role of Fap7, an ATPase, whose energy-requiring function positions it uniquely for a regulatory role, the putative endonuclease Nob1, and the helicase Rok1, in catalyzing successive pre-ribosome remodeling steps. Based on current data we hypothesize that energy-requiring conformational switches built into these steps are used to order and spatially and temporally regulate ribosome assembly, in a manner that is responsive to changes in the cellular growth environment. We will use a unique combination of yeast genetics, biochemical and biophysical experiments, and mechanistic enzymology to address Specific Aims: (1) Characterize a Conformational Change that Regulates Nob1. (2) Determine the Role of Rok1 in Promoting a Conformational Change. (3) Establish the Role of Fap7 In Ribosome Assembly. This work will lead to partial reconstitution of facilitated ribosome assembly, the discovery of novel intermediates and molecular-level information about the function of key assembly factors in orchestrating a conformational change that spatially and temporally regulates 40S assembly. In addition, knowledge of slow, regulated steps and information about the proteins catalyzing these steps will allow the ribosome assembly pathway to be targeted for drug purposes.
Ribosomes catalyze protein synthesis in all cells. Failure to correctly assembly ribosomes or to correctly regulate assembly can result in serious human disease, suggesting the ribosome assembly pathway as an important drug target. However, knowledge of individual processing steps and their regulation is required and will be garnered in this proposal.
| Collins, Jason C; Ghalei, Homa; Doherty, Joanne R et al. (2018) Ribosome biogenesis factor Ltv1 chaperones the assembly of the small subunit head. J Cell Biol 217:4141-4154 |
| Ferretti, Max Berman; Barre, Jennifer Louise; Karbstein, Katrin (2018) Translational Reprogramming Provides a Blueprint for Cellular Adaptation. Cell Chem Biol 25:1372-1379.e3 |
| Ghalei, Homa; Trepreau, Juliette; Collins, Jason C et al. (2017) The ATPase Fap7 Tests the Ability to Carry Out Translocation-like Conformational Changes and Releases Dim1 during 40S Ribosome Maturation. Mol Cell 67:990-1000.e3 |
| Johnson, Matthew C; Ghalei, Homa; Doxtader, Katelyn A et al. (2017) Structural Heterogeneity in Pre-40S Ribosomes. Structure 25:329-340 |
| Ferretti, Max B; Ghalei, Homa; Ward, Ethan A et al. (2017) Rps26 directs mRNA-specific translation by recognition of Kozak sequence elements. Nat Struct Mol Biol 24:700-707 |
| de la Cruz, Jesus; Karbstein, Katrin; Woolford Jr, John L (2015) Functions of ribosomal proteins in assembly of eukaryotic ribosomes in vivo. Annu Rev Biochem 84:93-129 |
| Karbstein, Katrin (2015) What will the future hold: RNP quality control and degradation. RNA 21:657-8 |
| Tyagi, Richa; Shahani, Neelam; Gorgen, Lindsay et al. (2015) Rheb Inhibits Protein Synthesis by Activating the PERK-eIF2? Signaling Cascade. Cell Rep : |
| Ghalei, Homa; Schaub, Franz X; Doherty, Joanne R et al. (2015) Hrr25/CK1?-directed release of Ltv1 from pre-40S ribosomes is necessary for ribosome assembly and cell growth. J Cell Biol 208:745-59 |
| Young, Crystal L; Khoshnevis, Sohail; Karbstein, Katrin (2013) Cofactor-dependent specificity of a DEAD-box protein. Proc Natl Acad Sci U S A 110:E2668-76 |
Showing the most recent 10 out of 20 publications
