Welcome to the Paddison Lab website. General lab focus areas include the study of molecular circuitry fueling cancer stem cells, the investigation of regulation of developmental transitions, and the use of cutting-edge genomic techniques, such as CRISPR-Cas9 and single cell techniques.
Our cancer-related research focuses on experimentation in patient-derived tumor stem-like cells using function genomics and single cell technologies. We seek to understand cancer-specific requirement for gene activities and, also, how tumors regulate cellular heterogeneity and developmental gene expression programs. Of late, we have pursued identification of genes impacting quiescent-like states in glioblastoma brain tumors. Because quiescent tumor cells are more resistant to standard of care therapies (e.g., chemotherapy and radiation), uncovering key regulatory features of quiescence ingress and egress may reveal unexpected therapeutic opportunities. Interestingly, recent studies have revealed that inducing quiescence in brain tumor cells can trigger gene expression programs observed in adult and fetal neural stem cells.
Our basic biological research has focused on understanding developmental programs and lineage transitions in embryonic and somatic stem and progenitor cells. We have uncovered multiple pathways and gene activities required for lineages transitions, including epigenetic and epitranscriptomic regulators. Regarding the later, the term "epitranscriptomics" refers to the dynamic chemical marking of RNAs which can affect their turnover, regulation, and function. From functional genomic screening, we uncovered N6-adenosine methylation of mRNA as a key molecular event regulating human hematopoietic lineage specification. Our studies have revealed thousands of mRNAs targeted for N6-adenosine methylation, the regulatory outcomes of methylation, and the developmental windows during which methylation is required. A recent screen identifed N6-adenosine methylation reader proteins required for regulatory outcomes.
We have also used CRISPR-Cas9 techniques to help better annotate human genes and reveal hidden gene domains in protein coding genes. Our strategy employs tiling mutagenesis to in living cells to mutate specific amino acid codons to determine their phenotypic requirement and their presence in predicted and novel gene domains. Most protein domains in the human genome are predicted based on homology to other sequenced organisms rather than through functional validation. However, these predictions make up only about 55% of the human proteome; the remaining proteome exists in a domain "void". Our approach has already identified 100s of previously unrecognized protein domains in genes with critical roles in chromosome segregation (in collaboration with the Biggins Lab).
An underlying theme uniting each project in lab is the use of functional genomics. While in graduate school, Dr. Paddison helped develop functional genomic techniques and genome-scale libraries to inhibit gene function in mammalian cells. We use these same strategies with updated gene inhibition technologies, such as CRISPR-Cas9, to perform functional genomic screens. We have successfully designed and performed dozens of genome scale screens in diverse developmental contexts, ranging from embryonic stem cells to neural stem cells to human brain tumor avatars.
Paddison PJ et al., 2002. Stable suppression of gene expression by RNAi in mammalian cells. PNAS 99:1443-1448.
Paddison PJ et al., 2002. Short hairpin RNAs (shRNAs) induce sequence-specific silencing in mammalian cells. Genes and Dev 16: 948-958.
Paddison PJ et al., 2004. A resource for large-scale RNAi based screens in mammals. Nature 428: 427-431.
Paddison PJ, 2008. RNA interference in mammalian cell systems. In: RNA interference. Current Topics in Microbiology and Immunology. Paddison PJ, Vogt PK (eds.). Springer Press.
Betschinger J et al., 2013. Exit from pluripotency is gated by intracellular redistribution of the bHLH transcription factor Tfe3. Cell 153(2):335-47.
Ding Y et al., 2013. Cancer-specific requirement for BUB1B/BUBR1 in human brain tumor isolates and genetically transformed cells. Cancer Discovery 3(2):198-211.
Hubert CG et al., 2013. Genome-wide RNAi screens in human brain tumor isolates reveal a novel viability requirement for PHF5A. Genes and Development 27:1032-45, 2013.
Schones DE et al., 2014. G9a/GLP-dependent H3K9me2 patterning alters chromatin structure at CpG islands in hematopoietic progenitors. Epigenetics and Chromatin 7:23. doi: 10.1186/1756-8935-7-23, 2014.
Toledo CM et al., 2015. Genome-wide CRISPR-Cas9 screens reveal loss of redundancy between PKMYT1 and WEE1 in Glioblastoma stem-like cells. Cell Reports 13:2425-39.
Ding Y et al., 2017. ZNF131 suppresses centrosome fragmentation in glioblastoma stem-like cells through regulation of HAUS5. Oncotarget 8:48545-48562.
Kuppers DA et al., 2019. N6-methyladenosine mRNA marking promotes selective translation of regulons required for human erythropoiesis. Nature Communications 10:4596, 2019.
Hoellerbauer P et al., 2020. A simple and highly efficient method for multi-allelic CRISPR-Cas9 editing in primary cell cultures. Cancer Reports, doi: 10.1002/cnr2.1269, 2020.
O'Connor SA et al., 2021. Neural G0: a quiescent-like state found in neuroepithelial-derived cells and glioma. Molecular Systems Biology 17:9522.
Herman JA et al., 2021. Functional dissection of human mitotic proteins using CRISPR-Cas9 tiling screens. BioRxiv, doi: https://doi.org/10.1101/2021.05.20.445000