RESUMEN
The urotensin 2 (uts2) gene family consists of four paralogs called uts2, uts2-related peptide (urp), urp1 and urp2. uts2 is known to exert a large array of biological effects, including osmoregulation, control of cardiovascular functions and regulation of endocrine activities. Lately, urp1 and urp2 have been shown to regulate axial straightening during embryogenesis. In contrast, much less is known about the roles of urp. The aim of the present study was to investigate the expression and the functions of urp by using the zebrafish as a model. For this purpose, we determined the expression pattern of the urp gene. We found that urp is expressed in motoneurons of the brainstem and the spinal cord, as in tetrapods. This was confirmed with a new Tg(urp:gfp) fluorescent reporter line. We also generated a urp knockout mutant by using CRISPR/Cas9-mediated genome editing and analysed its locomotor activity in larvae. urp mutant did not exhibit any apparent defect of spontaneous swimming when compared to wild-type. We also tested the idea that urp may represent an intermediary of urp1 and urp2 in their role on axial straightening. We found that the upward bending of the tail induced by the overexpression of urp2 in 24-hpf embryos was not altered in urp mutants. Our results indicate that urp does probably not act as a relay downstream of urp2. In conclusion, the present study showed that zebrafish urp gene is primarily expressed in motoneurons but is apparently dispensable for locomotor activity in the early larval stages.
Asunto(s)
Larva/metabolismo , Locomoción , Neuronas Motoras/metabolismo , Péptidos/metabolismo , Urotensinas/metabolismo , Pez Cebra/metabolismo , Animales , Animales Modificados Genéticamente , Sistemas CRISPR-Cas , Edición Génica/métodos , Hibridación in Situ , Pez Cebra/crecimiento & desarrolloRESUMEN
Urp1 and Urp2 are two neuropeptides of the urotensin II family identified in teleost fish and mainly expressed in cerebrospinal fluid (CSF)-contacting neurons. It has been recently proposed that Urp1 and Urp2 are required for correct axis formation and maintenance. Their action is thought to be mediated by the receptor Uts2r3, which is specifically expressed in dorsal somites. In support of this view, it has been demonstrated that the loss of uts2r3 results in severe scoliosis in adult zebrafish. In the present study, we report for the first time the occurrence of urp2, but not of urp1, in two tetrapod species of the Xenopus genus. In X. laevis, we show that urp2 mRNA-containing cells are CSF-contacting neurons. Furthermore, we identified utr4, the X. laevis counterparts of zebrafish uts2r3, and we demonstrate that, as in zebrafish, it is expressed in the dorsal somatic musculature. Finally, we reveal that, in X. laevis, the disruption of utr4 results in an abnormal curvature of the antero-posterior axis of the tadpoles. Taken together, our results suggest that the role of the Utr4 signalling pathway in the control of body straightness is an ancestral feature of bony vertebrates and not just a peculiarity of ray-finned fishes.
Asunto(s)
Evolución Biológica , Regulación del Desarrollo de la Expresión Génica , Filogenia , Receptores Acoplados a Proteínas G/metabolismo , Somatotipos , Urotensinas/metabolismo , Proteínas de Xenopus/metabolismo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Perfilación de la Expresión Génica , Receptores Acoplados a Proteínas G/genética , Homología de Secuencia , Proteínas de Xenopus/genética , Xenopus laevisRESUMEN
In Drosophila, the clock that controls rest-activity rhythms synchronizes with light-dark cycles through either the blue-light sensitive cryptochrome (Cry) located in most clock neurons, or rhodopsin-expressing histaminergic photoreceptors. Here we show that, in the absence of Cry, each of the two histamine receptors Ort and HisCl1 contribute to entrain the clock whereas no entrainment occurs in the absence of the two receptors. In contrast to Ort, HisCl1 does not restore entrainment when expressed in the optic lobe interneurons. Indeed, HisCl1 is expressed in wild-type photoreceptors and entrainment is strongly impaired in flies with photoreceptors mutant for HisCl1. Rescuing HisCl1 expression in the Rh6-expressing photoreceptors restores entrainment but it does not in other photoreceptors, which send histaminergic inputs to Rh6-expressing photoreceptors. Our results thus show that Rh6-expressing neurons contribute to circadian entrainment as both photoreceptors and interneurons, recalling the dual function of melanopsin-expressing ganglion cells in the mammalian retina.
Asunto(s)
Canales de Cloruro/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila/fisiología , Células Fotorreceptoras de Invertebrados/metabolismo , Rodopsina/metabolismo , Animales , Animales Modificados Genéticamente , Técnicas de Observación Conductual/instrumentación , Técnicas de Observación Conductual/métodos , Conducta Animal/fisiología , Canales de Cloruro/genética , Ojo Compuesto de los Artrópodos/citología , Ojo Compuesto de los Artrópodos/fisiología , Criptocromos/metabolismo , Proteínas de Drosophila/genética , Interneuronas/metabolismo , Masculino , Mutación , Lóbulo Óptico de Animales no Mamíferos/citología , Lóbulo Óptico de Animales no Mamíferos/metabolismo , FotoperiodoRESUMEN
Light is the major stimulus for the synchronization of circadian clocks with day-night cycles. The light-driven entrainment of the clock that controls rest-activity rhythms in Drosophila relies on different photoreceptive molecules. Cryptochrome (CRY) is expressed in most brain clock neurons, whereas six different rhodopsins (RH) are present in the light-sensing organs. The compound eye includes outer photoreceptors that express RH1 and inner photoreceptors that each express one of the four rhodopsins RH3-RH6. RH6 is also expressed in the extraretinal Hofbauer-Buchner eyelet, whereas RH2 is only found in the ocelli. In low light, the synchronization of behavioral rhythms relies on either CRY or the canonical rhodopsin phototransduction pathway, which requires the phospholipase C-ß encoded by norpA (no receptor potential A). We used norpA(P24) cry(02) double mutants that are circadianly blind in low light and restored NORPA function in each of the six types of photoreceptors, defined as expressing a particular rhodopsin. We first show that the NORPA pathway is less efficient than CRY for synchronizing rest-activity rhythms with delayed light-dark cycles but is important for proper phasing, whereas the two light-sensing pathways can mediate efficient adjustments to phase advances. Four of the six rhodopsin-expressing photoreceptors can mediate circadian entrainment, and all are more efficient for advancing than for delaying the behavioral clock. In contrast, neither RH5-expressing retinal photoreceptors nor RH2-expressing ocellar photoreceptors are sufficient to mediate synchronization through the NORPA pathway. Our results thus reveal different contributions of rhodopsin-expressing photoreceptors and suggest the existence of several circuits for rhodopsin-dependent circadian entrainment. J. Comp. Neurol. 524:2828-2844, 2016. © 2016 Wiley Periodicals, Inc.