Biallelic editing of a lamprey genome using the CRISPR/Cas9 system

Zu, Yao; Zhang, Xushuai; Ren, Jianfeng; Dong, Xuehong; Zhu, Zhe; Jia, Longfei; Zhang, Qinghua; Li, Weiming

doi:10.1038/srep23496

Cited by 51 publications

(32 citation statements)

References 41 publications

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“…"F0" mutagenesis strategy. We used CRISPR/Cas9-mediated mutagenesis to induce deletions and insertions (indels) into the protein-coding exons of injected "F0" sea lamprey (Petromyzon marinus) and African clawed frog (Xenopus laevis) embryos as previously Also as previously reported, the severity of a CRISPR/Cas9-induced phenotype correlates well with the percentage mutant alleles; with most "severely affected" F0 mosaic mutants typically exhibiting 75%-100% mutant (indel) alleles 22,[53][54][55][56] . Consistent with this, the 74 severely affected phenotypic mutants selected for genotyping in this study had an average of 88% indel alleles at targeted loci (Supplemental Tab.…”

Section: Methodsmentioning

confidence: 99%

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Cooption and Specialization of Endothelin Signaling Pathways Drove Elaboration of the Neural Crest in Early Vertebrates

Square¹,

Jandzík

Massey³

et al. 2019

Preprint

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The neural crest (NC) is a vertebrate-specific embryonic tissue that forms an array of clade-defining adult features. A key step in the formation of these diverse derivatives is the partitioning of NC cells into subpopulations with distinct migration routes and potencies 1 . The evolution of these developmental modules is poorly understood. Endothelin (Edn) signaling is unique to vertebrates, and performs various functions in different NC subpopulations 2-5 . To better understand the evolution of NC patterning, we used CRISPR/Cas9-driven mutagenesis to disrupt Edn receptors, ligands, and Dlx transcription factors in the sea lamprey, Petromyzon marinus. Lampreys and modern gnathostomes last shared a common ancestor 500 million years ago 6 . Thus, comparisons between the two groups can identify deeply conserved and divergent features of vertebrate development. Using Xenopus laevis to facilitate side-by-side analyses, we show here that lamprey and gnathostomes display fundamental differences in Edn signaling function. Unlike gnathostomes, both lamprey Ednrs cooperate during oropharyngeal skeleton development. Furthermore, neither paralog regulates hand transcription factors, which are required for mandible development in gnathostomes. We also identify conserved roles for Edn signaling in dlx gene regulation, pigment cell, and heart development. Together our results illustrate the stepwise neofunctionalization and specialization of this vertebrate-specific signaling pathway, and suggest key intermediate stages in the early evolution of the NC.

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Section: Methodsmentioning

confidence: 99%

“…described22,[53][54][55][56][57] . Though CRISPR/Cas9 is highly efficient in sea lamprey, differences in the efficiency of individual sgRNAs results in different ratios of wildtype and mutant alleles in F0 mosaic mutants.…”

mentioning

confidence: 99%

Cooption and Specialization of Endothelin Signaling Pathways Drove Elaboration of the Neural Crest in Early Vertebrates

Square¹,

Jandzík

Massey³

et al. 2019

Preprint

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show abstract

“…When delivered into zygotes or individual cells, the Cas9 protein complexes with the sgRNA and, when bound to the targeted sequence, induces a double-stranded DNA break that may be repaired by non-homologous end joining (NHEJ) repair machinery. Because NEHJ repair is error prone, this can result in short insertion/deletions (indels) that generate embryonic mutant genotypes that can be screened for phenotypic effects (Guo et al, 2014;Hwang et al, 2013;Irion et al, 2014;Jao et al, 2013;Qi et al, 2013;Square et al, 2015;Wang et al, 2016;Zu et al, 2016).…”

Section: Functional Genetic Analysis Of Lamprey Development Early Appmentioning

confidence: 99%

“…The Medeiros group also found that perturbation of FGF signaling caused loss of HhA expression in the zona limitans intrathalamica and Engrailed expression at the midbrain-hindbrain boundary, both of which are key organizing centers in the embryonic brain (Sugahara et al, 2011). A second demonstration of CRISPR/Cas9 in sea lamprey was by the Li laboratory, which focused on evaluating the efficacy of CRISPR knockouts on several genomic loci, including Golden (Gol), Kctd10, Wee1, SoxE2 and Wnt7b (Zu et al, 2016). Overall, and similar to Square et al (2015), Zu et al (2016) found high rates of biallelic mutagenesis, albeit with modest variation in efficacy across genomic targets and with different sgRNAs.…”

Section: Crispr/cas9 Genome Editing In Sea Lampreyproof Of Principlementioning

confidence: 99%

Functional genetic analysis in a jawless vertebrate, the sea lamprey: insights into the developmental evolution of early vertebrates

York

McCauley

2020

Journal of Experimental Biology

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Lampreys and hagfishes are the only surviving relicts of an ancient but ecologically dominant group of jawless fishes that evolved in the seas of the Cambrian era over half a billion years ago. Because of their phylogenetic position as the sister group to all other vertebrates ( jawed vertebrates), comparisons of embryonic development between jawless and jawed vertebrates offers researchers in the field of evolutionary developmental biology the unique opportunity to address fundamental questions related to the nature of our earliest vertebrate ancestors. Here, we describe how genetic analysis of embryogenesis in the sea lamprey (Petromyzon marinus) has provided insight into the origin and evolution of developmentalgenetic programs in vertebrates. We focus on recent work involving CRISPR/Cas9-mediated genome editing to study gene regulatory mechanisms involved in the development and evolution of neural crest cells and new cell types in the vertebrate nervous system, and transient transgenic assays that have been instrumental in dissecting the evolution of cis-regulatory control of gene expression in vertebrates. Finally, we discuss the broad potential for these functional genomic tools to address previously unanswerable questions related to the evolution of genomic regulatory mechanisms as well as issues related to invasive sea lamprey population control.

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“…While these hypotheses are speculative, they make predictions about ancestral regulatory connections and are hence testable by dissecting gene regulation in multiple lineages. With the recent development of methods for manipulating genes and regulatory elements in amphioxus and lamprey, 100,[192][193][194][195][196][197] plus application of genome-wide methods to identify regulatory landscapes, 184 this is rapidly becoming tractable.…”

Section: Conclusion/future Perspectives: Evolutionary Change In Spmentioning

confidence: 99%

Evolution of vertebrate spinal cord patterning

Leung

Shimeld

2019

Developmental Dynamics

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The vertebrate spinal cord is organized across three developmental axes, anterior‐posterior (AP), dorsal‐ventral (DV), and medial‐lateral (ML). Patterning of these axes is regulated by canonical intercellular signaling pathways: the AP axis by Wnt, fibroblast growth factor, and retinoic acid (RA), the DV axis by Hedgehog, Tgfβ, and Wnt, and the ML axis where proliferation is controlled by Notch. Developmental time plays an important role in which signal does what and when. Patterning across the three axes is not independent, but linked by interactions between signaling pathway components and their transcriptional targets. Combined this builds a sophisticated organ with many different types of cell in specific AP, DV, and ML positions. Two living lineages share phylum Chordata with vertebrates, amphioxus, and tunicates, while the jawless fish such as lampreys, survive as the most basally divergent vertebrate lineage. Genes and mechanisms shared between lampreys and other vertebrates tell us what predated vertebrates, while those also shared with other chordates tell us what evolved early in chordate evolution. Between these lie vertebrate innovations: genetic and developmental changes linked to evolution of new morphology. These include gene duplications, differences in how signals are received, and new regulatory connections between signaling pathways and their target genes.

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Biallelic editing of a lamprey genome using the CRISPR/Cas9 system

Cited by 51 publications

References 41 publications

Cooption and Specialization of Endothelin Signaling Pathways Drove Elaboration of the Neural Crest in Early Vertebrates

Cooption and Specialization of Endothelin Signaling Pathways Drove Elaboration of the Neural Crest in Early Vertebrates

Functional genetic analysis in a jawless vertebrate, the sea lamprey: insights into the developmental evolution of early vertebrates

Evolution of vertebrate spinal cord patterning

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