My lab is using functional genomics, molecular biology and model organism genetics to study fundamental mechanisms of genome maintenance and stability. Failure to maintain genome integrity leads to mutations that can lead to tumour formation. Individuals can therefore be disposed to cancer if they are exposed to carcinogens that overwhelm normal genome maintenance mechanisms, or if they harbour germline or somatic variants that create genomic instability. Our work is aimed at determining the causes of genomic instability as an enabling characteristic of tumour formation and exploring the potential of these early events to suggest novel therapeutic targets. Currently we are working on several projects, briefly outlined below.
Roles of RNA processing in genome maintenance
Functional genomic screening of the yeast model, Saccharomyces cerevisiae, and of mammalian cells has revealed that most steps in RNA metabolism can cause genome instability when disrupted. In some cases it is clear that mutations in RNA processing factors lead to transcription-coupled DNA:RNA hybrids that form R-loops. R-loop structures expose damage-prone single-stranded DNA and can also damage DNA by impeding DNA replication fork progression. Importantly, not all genome destabilizing RNA processing mutants create R-loops and we are interested in defining these other mechanisms as well as continuing to investigate causes and consequences of R-loop formation.
Most importantly, RNA processing mutations are now recognized to occur in various tumours. These include widely distributed and frequent mutations of the spliceosome (e.g. SF3B1 in hematopoietic malignancies), mutations in the RNA degrading exosome (e.g. DIS3) and in mRNA processing genes (e.g. FIP1L1). The importance of these cancer-associated mutations for genome maintenance and the potential for R-loop formation to contribute to cancer cell phenotypes is also under investigation.
Finally, at least in yeast, some RNA processing mutants exhibit synthetic lethality with cancer-associated genome maintenance genes, and therefore represent candidate therapeutic targets. We are exploring the conservation of such interactions and the potential for inhibiting aspects of RNA metabolism for therapeutic benefit.
Mechanistic basis of tumour mutation signatures
Mutational processes leave a characteristic pattern of mutations on tumour genomes based on the underlying biology that is disrupted or the causative environmental agent. While elegant computational methods can extract such mutation ‘signatures’ from tumour genomes, the mechanisms are usually unknown. We have established a yeast model for linking specific genetic changes in conserved genome integrity pathways to patterns of accumulated mutations using whole genome sequencing. Yeast provides a clean and tractable genetic system whose genome can be sequenced in an efficient, cost-effective manner. We are working to refine and expand our efforts to describe the mutation patterns associated with new genetic backgrounds, genotoxic environments and gene-environment interactions. Ultimately this work will intersect with tumour genome sequencing efforts to create testable hypotheses for defining the critical mutational processes operative in cancers.
Biogenesis of tumour-suppressor complexes
We have a long-standing interest in how molecular chaperones promote the biogenesis of multiprotein complexes. Our recent work has focused on the assembly and nuclear import of RNA polymerase and on the biogenesis and stability of PI3-kinase related kinases (PIKKs). Since human PIKKs are important tumour suppressors (e.g. ATM, ATR, mTOR, DNAPKcs), we are continuing to investigate how conserved chaperones such as the TTT (Tel2-Tti1-Tti2) complex and R2TP (Rvb1-Rvb2-Tah1-Pih1) complex regulate PIKK signaling. This will lead to studies investigating the interplay between chaperones and known disease-associated alleles of PIKKs.
Sorgjerd KM, Zako T, Sakono M, Stirling PC, Leroux MR, Saito T, Nilsson P, Sekimoto M, Saido TC and Maeda M. Human prefoldin inhibits Amyloid β (Aβ) fibrillation and contributes to formation of non-toxic Aβ aggregates. Biochemistry. 52, 3532-3542, 2013. View Abstract
van Pel DM*, Stirling PC*, Minaker SW, Sipahimalani P and Hieter P. Saccharomyces cerevisiae genetics predicts candidate therapeutic genetic interactions at the mammalian replication fork. G3 (Bethesda) 3, 273-282, 2013 *Authors contributed equally. View Abstract
Minaker SW, Filiatrault MC, Ben-Aroya S, Hieter P and Stirling PC. Biogenesis of RNA polymerases II and III requires the conserved GPN small GTPases in S. cerevisiae. Genetics 193, 853-864, 2013. View Abstract
Stirling PC*, Chan YA*, Minaker SW*, Aristizabal MJ, Barrett I, Sipahimalani P, Kobor, MS and Hieter P. R-loop mediated genome instability in mRNA cleavage and polyadenylation mutants. Genes Dev. 26, 163-175, 2012 *Authors contributed equally. View Abstract
Stirling PC, Crisp MJ, Basrai MA, Tucker CM, Dunham MJ, Spencer FA and Hieter P. Mutability and mutational spectrum of chromosome transmission fidelity (Ctf) genes. Chromosoma 121, 263-275, 2012. View Abstract
Stirling PC, Bloom MS, Solanki-Patil T, Smith S, Sipahimalani P, Li Z, Kofoed M, Ben-Aroya S, Myung K and Hieter P. The complete spectrum of yeast chromosome instability genes identifies candidate CIN cancer genes and functional roles for ASTRA complex components. PLoS Genet. 7, e1002057, 2011. *Faculty of 1000 rated. View Abstract
Lundin VF, Leroux MR* and Stirling PC*. Quality control of cytoskeletal proteins and human disease. Trends Biochem. Sci. Review. 35, 288-297, 2010. *Corresponding authors. View Abstract
Carroll SY*, Stirling PC*, Stimpson HE, Giesselman E, Schmitt MJ and Drubin DG. A yeast killer toxin screen provides insights into A/B toxin entry, trafficking and killing mechanisms. Dev. Cell 17, 552-560, 2009. *Authors contributed equally. *Faculty of 1000 rated. View Abstract
Dekker C*, Stirling PC*, Filmore H, McCormack EA, Pappenberger G, Paul A, Brost RL, Costanzo M, Boone C, Leroux MR and Willison KR. The interaction network of the chaperonin CCT. EMBO J. 27, 1837-1839, 2008.*Authors contributed equally. *Faculty of 1000 rated. View Abstract
Stirling PC, Srayko M, Takhar KS, Pozniakovsky A, Hyman AA and Leroux MR. Functional interaction between phosducin-like protein 2 and cytosolic chaperonin is essential for cytoskeletal protein function and cell cycle progression. Mol. Biol. Cell 18, 2336-2345, 2007. View Abstract
Martín-Benito J, Gómez-Reino J, Stirling PC, Lundin VF, Gómez-Puertas P, Boskovic J, Chacón P, Fernández JJ, Berenguer J, Leroux MR and Valpuesta JM. Divergent substrate-binding mechanisms reveal an evolutionary specialization of eukaryotic prefoldin compared to its archaeal counterpart. Structure 15, 101-110, 2007. View Abstract
Stirling PC, Bakhoum SF, Feigl AB and Leroux MR. Convergent evolution of clamp-like binding sites in diverse chaperones. Nat. Struct. Mol. Biol. 13, 865-870. 2006. Review. View Abstract
Stirling PC, Cuellar J, Alfaro GA, El Khadali F, Beh CT, Valpuesta JM, Melki R and Leroux MR. PhLP3 modulates CCT-mediated actin and tubulin folding via ternary complexes with substrates. J. Biol. Chem. 281, 7012-7021, 2006. View Abstract
Lundin VF*, Stirling PC*, Gomez-Reino J, Mwenifumbo JC, Obst JM, Valpuesta JM and Leroux MR. Molecular clamp mechanism of substrate binding by hydrophobic coiled coil residues in the archaeal chaperone prefoldin. Proc. Natl. Acad. Sci. USA 101, 4367-4372, 2004. *Authors contributed equally. *Faculty of 1000 rated. View Abstract
Stirling PC*, Lundin VF* and Leroux MR. Getting a grip on non-native proteins. EMBO Reports 4, 565-570, 2003. *Authors contributed equally. Review. View Abstract