Wnt-regulated lncRNA discovery enhanced by<i>in vivo</i>identification and CRISPRi functional validation

Liu, Shiyang; Harmston, Nathan; Glaser, Trudy Lee; Wong, Yunka; Zhong, Zheng; Madan, Babita; Virshup, David M.; Petretto, Enrico

doi:10.1101/2020.02.09.941005

Cited by 3 publications

(3 citation statements)

References 89 publications

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“…Loci containing let‐7 family microRNAs are frequently within 9q22.3 deletions and have been implicated in aspects of the phenotype (Yamada et al, 2020) but these sequences, at around 9:94,220,000, are outside the deletion in the present patient. A recent article (Liu et al, 2020) showed that a sequence within the deletion (designated RP11‐43505.2 in the article, which maps close to MT1P1 and likely overlaps with FANCC‐203 ) is targeted by WNT signaling. Several WNT genes are targets of hedgehog signaling (Onodera et al, 2017), and their expression would likely be increased when one copy of PTCH1 is deleted.…”

Section: Discussionmentioning

confidence: 99%

Microdeletion of 9q22.3: A patient with minimal deletion size associated with a severe phenotype

Ewing

Cheetham

McGill

et al. 2021

American J of Med Genetics Pt A

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Basal cell nevus syndrome (also known as Gorlin Syndrome; MIM109400) is an autosomal dominant disorder characterized by recurrent pathological features such as basal cell carcinomas and odontogenic keratocysts as well as skeletal abnormalities. Most affected individuals have point mutations or small insertions or deletions within the PTCH1 gene on human chromosome 9, but there are some cases with more extensive deletion of the region, usually including the neighboring FANCC and/or ERCC6L2 genes. We report a 16‐year‐old patient with a deletion of approximately 400,000 bases which removes only PTCH1 and some non‐coding RNA genes but leaves FANCC and ERCC6L2 intact. In spite of the small amount of DNA for which he is haploid, his phenotype is more extreme than many individuals with longer deletions in the region. This includes early presentation with a large number of basal cell nevi and other skin lesions, multiple jaw keratocysts, and macrosomia. We found that the deletion was in the paternal chromosome, in common with other macrosomia cases. Using public databases, we have examined possible interactions between sequences within and outside the deletion and speculate that a regulatory relationship exists with flanking genes, which is unbalanced by the deletion, resulting in abnormal activation or repression of the target genes and hence the severity of the phenotype.

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Section: Discussionmentioning

confidence: 99%

Microdeletion of 9q22.3: A patient with minimal deletion size associated with a severe phenotype

Ewing

Cheetham

McGill

et al. 2021

American J of Med Genetics Pt A

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“…They selected a total of 1503 Wnt-regulated lncRNAs for a CRISPRi screen of the HPAF-II cell line (a human pancreatic adenocarcinoma cell), which contained 3151 transcripts including different isoforms. The researchers found that the four Wnt-regulated lncRNA loci (RP11-481J2.3, LINC00910, LINC00263, CCAT1) that were hits in the in vivo and in vitro screenings were essential for the growth of HPAF-II cancer cells [43]. dCas9 can also be used to activate gene expression, in an approach termed CRISPR activation (CRISPRa).…”

Section: Application Of Crispr -Cas9 In Gene Screeningmentioning

confidence: 99%

Review of applications of CRISPR-Cas9 gene-editing technology in cancer research

Zhao

Tong

et al. 2021

Biol Proced Online

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Characterized by multiple complex mutations, including activation by oncogenes and inhibition by tumor suppressors, cancer is one of the leading causes of death. Application of CRISPR-Cas9 gene-editing technology in cancer research has aroused great interest, promoting the exploration of the molecular mechanism of cancer progression and development of precise therapy. CRISPR-Cas9 gene-editing technology provides a solid basis for identifying driver and passenger mutations in cancer genomes, which is of great value in genetic screening and for developing cancer models and treatments. This article reviews the current applications of CRISPR-Cas9 gene-editing technology in various cancer studies, the challenges faced, and the existing solutions, highlighting the potential of this technology for cancer treatment.

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“…The full reference transcriptome GTF file and time-course RNA-seq reads quantification are available in the figshare repository at https://doi.org/10.6084/m9.figshare. 13012535 [99]. Detailed results for the Wnt-regulated lncRNAs can be found in Additional file 2: Table S1.…”

Section: Supplementary Informationmentioning

confidence: 99%

Wnt-regulated lncRNA discovery enhanced by in vivo identification and CRISPRi functional validation

et al. 2020

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Background Wnt signaling is an evolutionarily conserved developmental pathway that is frequently hyperactivated in cancer. While multiple protein-coding genes regulated by Wnt signaling are known, the functional lncRNAs regulated by Wnt signaling have not been systematically characterized. Methods We comprehensively mapped Wnt-regulated lncRNAs from an orthotopic Wnt-addicted pancreatic cancer model and examined the response of lncRNAs to Wnt inhibition between in vivo and in vitro cancer models. We further annotated and characterized these Wnt-regulated lncRNAs using existing genomic classifications (using data from FANTOM5) in the context of Wnt signaling and inferred their role in cancer pathogenesis (using GWAS and expression data from the TCGA). To functionally validate Wnt-regulated lncRNAs, we performed CRISPRi screens to assess their role in cancer cell proliferation both in vivo and in vitro. Results We identified 3633 lncRNAs, of which 1503 were regulated by Wnt signaling in an orthotopic Wnt-addicted pancreatic cancer model. These lncRNAs were much more sensitive to changes in Wnt signaling in xenografts than in cultured cells. Our analysis suggested that Wnt signaling inhibition could influence the co-expression relationship of Wnt-regulated lncRNAs and their eQTL-linked protein-coding genes. Wnt-regulated lncRNAs were also implicated in specific gene networks involved in distinct biological processes that contribute to the pathogenesis of cancers. Consistent with previous genome-wide lncRNA CRISPRi screens, around 1% (13/1503) of the Wnt-regulated lncRNAs were found to modify cancer cell growth in vitro. This included CCAT1 and LINC00263, previously reported to regulate cancer growth. Using an in vivo CRISPRi screen, we doubled the discovery rate, identifying twice as many Wnt-regulated lncRNAs (25/1503) that had a functional effect on cancer cell growth. Conclusions Our study demonstrates the value of studying lncRNA functions in vivo, provides a valuable resource of lncRNAs regulated by Wnt signaling, and establishes a framework for systematic discovery of functional lncRNAs.

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Wnt-regulated lncRNA discovery enhanced byin vivoidentification and CRISPRi functional validation

Cited by 3 publications

References 89 publications

Microdeletion of 9q22.3: A patient with minimal deletion size associated with a severe phenotype

Microdeletion of 9q22.3: A patient with minimal deletion size associated with a severe phenotype

Review of applications of CRISPR-Cas9 gene-editing technology in cancer research

Wnt-regulated lncRNA discovery enhanced by in vivo identification and CRISPRi functional validation

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