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The advent of high-throughput sequencing technology facilitates the exploration of a variety of reference species outside the few established molecular genetic model systems. Bioinformatic and gene expression analyses provide new ways for comparative analyses between species, for instance, in the field of evolution and development. Despite these advances, a critical bottleneck for the exploration of new model species remains the establishment of functional tools, such as the ability to experimentally express genes in specific cells of an organism. We recently established a first transgenic strain of the annelid Platynereis, using a Tc1/mariner-type Mos1 transposon vector. Here, we compare Mos1 with Tol2, a member of the hAT family of transposons. In Platynereis, Tol2-based constructs showed a higher frequency of nuclear genome insertion and sustained gene expression in the G0 generation. However, in contrast to Mos1-mediated transgenes, Tol2-mediated insertions failed to retain fluorescence in the G1 generation, suggesting a germ line-based silencing mechanism. Furthermore, we present three novel expression constructs that were generated by a simple fusion-PCR approach and allow either ubiquitous or cell-specific expression of a reporter gene. Our study indicates the versatility of Tol2 for transient transgenesis, and provides a template for transgenesis work in other emerging reference species.

Research in eye evolution has mostly focused on eyes residing in the head. In contrast, noncephalic light sensors are far less understood and rather regarded as evolutionary innovations. We established stable transgenesis in the annelid Platynereis , a reference species for evolutionary and developmental comparisons. EGFP controlled by cis -regulatory elements of r-opsin , a characteristic marker for rhabdomeric photoreceptors, faithfully recapitulates known r-opsin expression in the adult eyes, and marks a pair of pigment-associated frontolateral eyelets in the brain. Unexpectedly, transgenic animals revealed an additional series of photoreceptors in the ventral nerve cord as well as photoreceptors that are located in each pair of the segmental dorsal appendages (notopodia) and project into the ventral nerve cord. Consistent with a photosensory function of these noncephalic cells, decapitated animals display a clear photoavoidance response. Molecular analysis of the receptors suggests that they differentiate independent of pax6 , a gene involved in early eye development of many metazoans, and that the ventral cells may share origins with the Hesse organs in the amphioxus neural tube. Finally, expression analysis of opn4×-2 and opn4m-2 , two zebrafish orthologs of Platynereis r-opsin , reveals that these genes share expression in the neuromasts, known mechanoreceptors of the lateral line peripheral nervous system. Together, this establishes that noncephalic photoreceptors are more widespread than assumed, and may even reflect more ancient aspects of sensory systems. Our study marks significant advance for the understanding of photoreceptor cell (PRC) evolution and development and for Platynereis as a functional lophotrochozoan model system.

The advent of high-throughput sequencing technology facilitates the exploration of a variety of reference species outside the few established molecular genetic model systems. Bioinformatic and gene expression analyses provide new ways for comparative analyses between species, for instance, in the field of evolution and development. Despite these advances, a critical bottleneck for the exploration of new model species remains the establishment of functional tools, such as the ability to experimentally express genes in specific cells of an organism. We recently established a first transgenic strain of the annelid Platynereis, using a Tc1/mariner-type Mos1 transposon vector. Here, we compare Mos1 with Tol2, a member of the hAT family of transposons. In Platynereis, Tol2-based constructs showed a higher frequency of nuclear genome insertion and sustained gene expression in the G0 generation. However, in contrast to Mos1-mediated transgenes, Tol2-mediated insertions failed to retain fluorescence in the G1 generation, suggesting a germ line-based silencing mechanism. Furthermore, we present three novel expression constructs that were generated by a simple fusion-PCR approach and allow either ubiquitous or cell-specific expression of a reporter gene. Our study indicates the versatility of Tol2 for transient transgenesis, and provides a template for transgenesis work in other emerging reference species.

Research in eye evolution has mostly focused on eyes residing in the head. In contrast, noncephalic light sensors are far less understood and rather regarded as evolutionary innovations. We established stable transgenesis in the annelid Platynereis , a reference species for evolutionary and developmental comparisons. EGFP controlled by cis -regulatory elements of r-opsin , a characteristic marker for rhabdomeric photoreceptors, faithfully recapitulates known r-opsin expression in the adult eyes, and marks a pair of pigment-associated frontolateral eyelets in the brain. Unexpectedly, transgenic animals revealed an additional series of photoreceptors in the ventral nerve cord as well as photoreceptors that are located in each pair of the segmental dorsal appendages (notopodia) and project into the ventral nerve cord. Consistent with a photosensory function of these noncephalic cells, decapitated animals display a clear photoavoidance response. Molecular analysis of the receptors suggests that they differentiate independent of pax6 , a gene involved in early eye development of many metazoans, and that the ventral cells may share origins with the Hesse organs in the amphioxus neural tube. Finally, expression analysis of opn4×-2 and opn4m-2 , two zebrafish orthologs of Platynereis r-opsin , reveals that these genes share expression in the neuromasts, known mechanoreceptors of the lateral line peripheral nervous system. Together, this establishes that noncephalic photoreceptors are more widespread than assumed, and may even reflect more ancient aspects of sensory systems. Our study marks significant advance for the understanding of photoreceptor cell (PRC) evolution and development and for Platynereis as a functional lophotrochozoan model system.