DREAM interrupted: Establishing a CRISPR/Cas9 functional genomics pipeline in Caenorhabditis elegans (2021-02-19)

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DREAM interrupted: Establishing a CRISPR/Cas9 functional genomics pipeline in Caenorhabditis elegans (2021-02-19)

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In multicellular organisms, how do multiple cell types originate from one cell and one genome? A critical early cell fate decision is whether to develop as germline (reproductive) or soma (non-reproductive). My research focuses on assessing how the highly conserved DREAM transcriptional repressor complex maintains this germ vs. soma cell fate decisions. Loss of the Caenorhabditis elegans DREAM complex results in misexpression of germline genes in somaticcells, a phenomenon called a "soma-to-germline transition" that is commonly observed in cancer cells. To tackle how DREAM protects somatic cell identity, the Goetsch lab utilizes a functional genomic pipeline that integrates new CRISPR/Cas9 genome editing tools, high-throughput sequencing analysis, genetics, biochemistry, and cell biology. We are currently investigating how DREAM complex formation on chromatin mediates target gene repression. The 8-subunit DREAM complex contains 3 subcomponenets: an E2F-DP transcription factor heterodimer, a Retinoblastoma(Rb)-like pocket protein, and a 5-submit complex called MuvB. We hypothesize that MuvB chromatin occupancy, aided and stabilized by its association with E2F-DP and the Rb-like pocket protein, establishes and maintains target gene repression. Using CRISPR/Cas9 targeted mutagensis, we disrupted the interaction between MuvB and the sole C. elegans Rb-like pocket protein LIN-35. We expected that severing MuvB from the complex would destablize DREAM component assembly on chromatin and its repression of target genes. Instead, we observed that disrupting LIN-35-MuvB association did not affect DREAM chromatin occupancy, but we did observe upregulation of some DREAM target genes. Our current aim is to establish how the molecular events that drive DREAM complex formation contribute to target gene repression using our CRISPR/Cas9 functional genomics pipeline. In the future, we intend to use similar methods to illuminate how DREAM loss unlocks a network of downstream transcriptional processes that ultimately destabilizes somatic cell identity.


Department of Biology Seminar this Friday, February 19, 2021 at 3:00 pm on Zoom; Speaker: Paul Goetsch, Assistant Professor at Michigan Technological University; Hosted by: Dr. Huai Deng; Friday February 19, 2021; We encourage you to join us at https://z.umn.edu/scsefriday at 3:00 PM

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Goetsch, Paul; University of Minnesota Duluth. Department of Biology. (2021). DREAM interrupted: Establishing a CRISPR/Cas9 functional genomics pipeline in Caenorhabditis elegans (2021-02-19). Retrieved from the University Digital Conservancy, https://hdl.handle.net/11299/220689.

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