The Embryonic Development of the Hawaiian Bobtail Squid <i>(Euprymna scolopes)</i>

Lee, Patricia N.; Callaerts, Patrick; Couet, H. Gert de

doi:10.1101/pdb.ip77

Cited by 22 publications

(28 citation statements)

References 6 publications

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“…The embryonic stages of development spanned eye and light organ morphogenesis (Naef stages 18, 22, 26, 29, Table 1; Arnold et al, 1972; Lee et al, 2009). To minimize variation in expression patterns due to genetic effects, we tested each gene using samples from a single clutch in this and all subsequent ISH experiments.…”

Section: Methodsmentioning

confidence: 99%

Eye-specification genes in the bacterial light organ of the bobtail squid Euprymna scolopes, and their expression in response to symbiont cues

Peyer¹,

Pankey²,

Oakley³

et al. 2014

Mechanisms of Development

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The squid Euprymna scolopes has evolved independent sets of tissues capable of light detection, including a complex eye and a photophore or ‘light organ’, which houses the luminous bacterial symbiont Vibrio fischeri. As the eye and light organ originate from different embryonic tissues, we examined whether the eye-specification genes, pax6, eya, six, and dac, are shared by these two organs, and if so, whether they are regulated in the light organ by symbiosis. We obtained sequences of the four genes with PCR, confirmed orthology with phylogenetic analysis, and determined that each was expressed in the eye and light organ. With in situ hybridization (ISH), we localized the gene transcripts in developing embryos, comparing the patterns of expression in the two organs. The four transcripts localized to similar tissues, including those associated with the visual system ~1/4 into embryogenesis (Naef stage 18) and the light organ ~3/4 into embryogenesis (Naef stage 26). We used ISH and quantitative real-time PCR to examine transcript expression and differential regulation in postembryonic light organs in response to the following colonization conditions: wild-type, luminescent V. fischeri; a mutant strain defective in light production; and as a control, no symbiont. In ISH experiments light organs showed down regulation of the pax6, eya, and six transcripts in response to wild-type V. fischeri. Mutant strains also induced down regulation of the pax6 and eya transcripts, but not of the six transcript. Thus, luminescence was required for down regulation of the six transcript. We discuss these results in the context of symbiont-induced light-organ development. Our study indicates that the eye-specification genes are expressed in light-interacting tissues independent of their embryonic origin and are capable of responding to bacterial cues. These results offer evidence for evolutionary tinkering or the recruitment of eye development genes for use in a light-sensing photophore.

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Section: Methodsmentioning

confidence: 99%

Eye-specification genes in the bacterial light organ of the bobtail squid Euprymna scolopes, and their expression in response to symbiont cues

Peyer¹,

Pankey²,

Oakley³

et al. 2014

Mechanisms of Development

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“…Egg clutches were kept in separate containers and staged embryos were removed as needed. Staging criteria for E. scolopes can be found in Lee et al (2009a).…”

Section: Methodsmentioning

confidence: 99%

Developmental expression ofapterous/Lhx2/9in the sepiolid squidEuprymna scolopessupports an ancestral role in neural development

Farfán

Shigeno

Nödl

et al. 2009

Evolution and Development

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The transcription factors Apterous/Lhx2/9 play many pivotal roles in the development of protostomes and deuterostomes, most notably limb patterning, eye morphogenesis, and brain development. Full-length apterous/lhx2/9 homologs have been isolated from several invertebrate species, but hitherto not from a lophotrochozoan. Here, we report the isolation, characterization, and spatio-temporal expression of apterous in the sepiolid squid Euprymna scolopes. The isolated composite cDNA encodes a hypothetical protein of 448 amino acid residues with a typical LIM-homeodomain (LIM-HD) structure and the greatest overall sequence similarity to vertebrate Lhx2/9 proteins. The Euprymna scolopes apterous (Es-ap) expression patterns provided no indication of a role in the early dorso/ventral patterning or growth of the arm crown that showed expression only in two ventral cords running in parallel inside the arms and tentacles and at the base of the suckers, a region rich in nerve endings and chemosensory neurons. The Es-ap hybridization signal was also conspicuous in the eyes, olfactory organs, optic lobes, and in several lobes of the supraesophageal mass, among these the olfactory and vertical lobes, and paravertical bodies. The observed expression patterns suggest gene involvement in eye morphogenesis and neural wiring of sensory structures, including those for olfaction and vision.

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“…The whole-mount in situ hybridization method described here is modified from the protocol of Seaver et al (2001), using DIG-labeled riboprobes prepared as described in Whole-Mount In Situ Hybridization of Hawaiian Bobtail Squid (Euprymna scolopes) Embryos with DIG-Labeled Riboprobes: I. DNA Template Preparation and In Vitro Transcription of Riboprobes (Lee et al 2009a (Lee et al 2009c). <!>Formaldehyde (3.7%, prepared in filtered seawater) <!>Formaldehyde (3.7%, prepared in PTw for ISH) Glycerol (70%, prepared in PTw for ISH) Glycine (2 mg/mL, prepared in PTw for ISH) <!>NBT (Roche Applied Science 11383213001) Store in the dark at -20°C.…”

Section: Related Informationmentioning

confidence: 99%

Whole-Mount In Situ Hybridization of Hawaiian Bobtail Squid (Euprymna scolopes) Embryos with DIG-Labeled Riboprobes: II. Embryo Preparation, Hybridization, Washes, and Immunohistochemistry

Lee

McFall‐Ngai²,

Callaerts³

et al. 2009

Cold Spring Harb Protoc

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Whole-mount in situ hybridization is a technique used to localize and visualize specific gene transcripts in whole embryos by hybridizing labeled RNA probes complementary to the sequence of interest. A digoxigenin (DIG)-labeled riboprobe synthesized during in vitro transcription through the incorporation of DIG-labeled UTP is hybridized to the target sequence under stringent conditions, and excess unhybridized probe is removed during a series of washes. The location of the labeled riboprobe, and thus the mRNA sequence of interest, is then visualized by immunohistochemistry. This protocol outlines the steps involved in preparing Hawaiian bobtail squid (Euprymna scolopes) embryos, hybridizing a DIG-labeled riboprobe in whole-mount embryos, and visualizing the labeled RNA colorimetrically using an alkaline-phosphatase-conjugated anti-DIG antibody.

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The Embryonic Development of the Hawaiian Bobtail Squid (Euprymna scolopes)

Cited by 22 publications

References 6 publications

Eye-specification genes in the bacterial light organ of the bobtail squid Euprymna scolopes, and their expression in response to symbiont cues

Eye-specification genes in the bacterial light organ of the bobtail squid Euprymna scolopes, and their expression in response to symbiont cues

Developmental expression ofapterous/Lhx2/9in the sepiolid squidEuprymna scolopessupports an ancestral role in neural development

Whole-Mount In Situ Hybridization of Hawaiian Bobtail Squid (Euprymna scolopes) Embryos with DIG-Labeled Riboprobes: II. Embryo Preparation, Hybridization, Washes, and Immunohistochemistry

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