Nanos-mediated repression of <i>hid</i> protects larval sensory neurons after a switch in sensitivity to apoptotic signals

Bhogal, Balpreet; Plaza-Jennings, Amara; Gavis, Elizabeth R.

doi:10.1242/dev.132415

Cited by 16 publications

(18 citation statements)

References 63 publications

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“…For the yg -HEG, only 17% of flies that inherited the HEG from the mother were fertile, again showing the negative effect on fitness due to maternal carryover. Nanos plays roles in somatic tissues in the nervous system of Drosophila ( 52 ). However, it is unlikely that somatic expression in the zygote contributes to the yellow-g sterile phenotype since no fitness cost was observed in HEG/TM3, Sb progeny when the HEG came from a father.…”

Section: Discussionmentioning

confidence: 99%

Behavior of homing endonuclease gene drives targeting genes required for viability or female fertility with multiplexed guide RNAs

Oberhofer

Ivy

Hay

2018

Proc. Natl. Acad. Sci. U.S.A.

107

130

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SignificanceHoming endonuclease gene (HEG)-based gene drive can bring about population suppression when genes required for viability or fertility are targeted. However, these strategies are vulnerable to failure through mechanisms that create alleles resistant to cleavage but that retain wild-type gene function. We show that resistance allele creation can be prevented through the use of guide RNAs designed to cleave a gene at four target sites. However, homing rates were modest, and the HEGs were unstable during homing. In addition, use of a promoter active in the female germline resulted in levels of HEG carryover that compromised the viability or fertility of HEG-bearing heterozygotes, thereby preventing drive. We propose strategies that can help to overcome these problems in next-generation HEG systems.

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Section: Discussionmentioning

confidence: 99%

Behavior of homing endonuclease gene drives targeting genes required for viability or female fertility with multiplexed guide RNAs

Oberhofer

Ivy

Hay

2018

Proc. Natl. Acad. Sci. U.S.A.

107

130

View full text Add to dashboard Cite

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“…The following reporters were used to visualize C4da neurons: ppk-GAL4, UAS-CD4:tdTom; ppk-GAL4, UAS-CD4:tdGFP; GAL4 477 , UAS-CD4:tdTom; GAL4 477 , UAS-CD4:td GFP (Bhogal et al, 2016), ppk-CD4:tdTom and ppk-CD4:tdGFP (Han et al, 2011). C1da neurons were visualized using GAL4 221 , UAS-mCD8:GFP (Grueber et al, 2003) and C3da neurons were visualized with UAS-mCD8:GFP; GAL4 1003.3 (Hughes et al, 2007).…”

Section: Methodsmentioning

confidence: 99%

“…Immunofluorescence was performed as previously described (Bhogal et al, 2016; Grueber et al, 2002). Unless otherwise noted, wandering third instar larvae were selected for analysis.…”

Section: Methodsmentioning

confidence: 99%

Enclosure of Dendrites by Epidermal Cells Restricts Branching and Permits Coordinated Development of Spatially Overlapping Sensory Neurons

et al. 2017

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SUMMARY Spatial arrangement of different neuron types within a territory is essential to neuronal development and function. How development of different neuron types is coordinated for spatial coexistence is poorly understood. In Drosophila, dendrites of four classes of dendritic arborization (C1-C4da) neurons innervate overlapping receptive fields within the larval epidermis. These dendrites are intermittently enclosed by epidermal cells, with different classes exhibiting varying degrees of enclosure. The role of enclosure in neuronal development and its underlying mechanism remain unknown. We show that the membrane-associated protein Coracle acts in C4da neurons and epidermal cells to locally restrict dendrite branching and outgrowth by promoting enclosure. Loss of C4da neuron enclosure results in excessive branching and growth of C4da neuron dendrites, and retraction of C1da neuron dendrites due to local inhibitory interactions between neurons. We propose that enclosure of dendrites by epidermal cells is a developmental mechanism for coordinated innervation of shared receptive fields.

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“…Thus, when the Dfd -HEG came from a female in the cross Dfd -HEG/+ x Df/TM3, Sb, the number of progeny carrying what should be a wildtype balancer chromosome (clv/TM3, Sb and HEG/TM3, Sb ) was only 8% of that obtained when the sexes were switched.. For the yg -HEG only 17% of flies that inherited the HEG from the mother, were fertile, again showing the negative effect on fitness due to maternal carry over. Nanos plays roles in somatic tissues in the nervous system of Drosophila (46) . However, it is unlikely that somatic expression in the zygote contributes to the yellow-g sterile phenotype, since no fitness cost was observed in HEG/TM3, Sb progeny when the HEG came from a father.…”

Section: Spreading Of the Heg Through The Female Germ Line Is Severelmentioning

confidence: 99%

Behavior of homing endonuclease gene drives targeting genes required for viability or female fertility with multiplexed guide RNAs

Oberhofer

Ivy

Hay

2018

Preprint

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A gene drive method of particular interest for population suppression utilizes homing endonuclease genes (HEGs), wherein a site-specific nuclease-encoding cassette is copied, in the germline, into a target gene whose loss of function results in loss of viability or fertility in homozygous, but not heterozygous progeny. Earlier work in Drosophila and mosquitoes utilized HEGs consisting of Cas9 and a single gRNA that together target a specific gene for cleavage. Homing was observed, but resistant alleles, immune to cleavage, while retaining wildtype gene function, were also created through non-homologous end joining. Such alleles prevent drive and population suppression. Targeting a gene for cleavage at multiple positions has been suggested as a strategy to prevent the appearance of resistant alleles. To test this hypothesis, we generated two suppression HEGs, targeting genes required for embryonic viability or fertility, using a HEG consisting of CRISPR/Cas9 and guide RNAs (gRNAs) designed to cleave each gene at four positions. Rates of target locus cleavage were very high, and multiplexing of gRNAs prevented resistant allele formation. However, germline homing rates were modest, and the HEG cassette was unstable during homing events, resulting in frequent partial copying of HEGs that lacked gRNAs, a dominant marker gene, or Cas9. Finally, in drive experiments the HEGs failed to spread, due to the high fitness load induced in offspring as a result of maternal carry over of Cas9/gRNA complex activity. Alternative design principles are proposed that may mitigate these problems in future gene drive engineering. Significance statementHEG-based gene drive can bring about population suppression when genes required for viability or fertility are targeted. However, these strategies are vulnerable to failure through mechanisms that create alleles resistant to cleavage, but that retain wildtype gene function. We show that resistance allele creation can be prevented through the use of gRNAs designed to cleave a gene at four target sites. However, homing rates were modest, and the HEGs were unstable during homing. In addition, use of a promoter active in the female germline resulted in levels of HEG carryover that compromised the viability or fertility of HEG-bearing heterozygotes, thereby preventing drive. We propose strategies that can help to overcome these problems in next generation HEG systems.

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Nanos-mediated repression of hid protects larval sensory neurons after a switch in sensitivity to apoptotic signals

Cited by 16 publications

References 63 publications

Behavior of homing endonuclease gene drives targeting genes required for viability or female fertility with multiplexed guide RNAs

Behavior of homing endonuclease gene drives targeting genes required for viability or female fertility with multiplexed guide RNAs

Enclosure of Dendrites by Epidermal Cells Restricts Branching and Permits Coordinated Development of Spatially Overlapping Sensory Neurons

Behavior of homing endonuclease gene drives targeting genes required for viability or female fertility with multiplexed guide RNAs

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