Expression and Developmental Regulation of the Hydra-RFamide and Hydra-LWamide Preprohormone Genes inHydra:Evidence for Transient Phases of Head Formation

Abstract

Hydra magnipapillata has three distinct genes coding for preprohormones A, B, and C, each yielding a characteristic set of Hydra-RFamide (Arg-Phe-NH2) neuropeptides, and a fourth gene coding for a preprohormone that yields various Hydra-LWamide (Leu-Trp-NH2) neuropeptides. Using a whole-mount double-labeling in situ hybridization technique, we found that each of the four genes is specifically expressed in a different subset of neurons in the ectoderm of adult Hydra. The preprohormone A gene is expressed in neurons of the tentacles, hypostome (a region between tentacles and mouth opening), upper gastric region, and peduncle (an area just above the foot). The preprohormone B gene is exclusively expressed in neurons of the hypostome, whereas the preprohormone C gene is exclusively expressed in neurons of the tentacles. The Hydra-LWamide preprohormone gene is expressed in neurons located in all parts of Hydra with maxima in tentacles, hypostome, and basal disk (foot). Studies on animals regenerating a head showed that the prepro-Hydra-LWamide gene is expressed first, followed by the preprohormone A and subsequently the preprohormone C and the preprohormone B genes. This sequence of events could be explained by a model based on positional values in a morphogen gradient. Our head-regeneration experiments also give support for transient phases of head formation: first tentacle-specific preprohormone C neurons (frequently associated with a small tentacle bud) appear at the center of the regenerating tip, which they are then replaced by hypostome-specific preprohormone B neurons. Thus, the regenerating tip first attains a tentacle-like appearance and only later this tip develops into a hypostome. In a developing bud of Hydra, tentacle-specific preprohormone C neurons and hypostome-specific preprohormone B neurons appear about simultaneously in their correct positions, but during a later phase of head development, additional tentacle-specific preprohormone C neurons appear as a ring at the center of the hypostome and then disappear again. Nerve-free Hydra consisting of only epithelial cells do not express the preprohormone A, B, or C or the LWamide preprohormone genes. These animals, however, have a normal phenotype, showing that the preprohormone A, B, and C and the LWamide genes are not essential for the basic pattern formation of Hydra.

Head regeneration in wild-type hydra requires de novo neurogenesis

“…As well as the nematocyte pathway, hyCOUP-TF, CnASH and CREB are also expressed in the neuronal lineage Hayakawa et al, 2004;Chera et al, 2007), whereas other genes display a neuronal-specific expression, such as the PRD-class prdl-a (Gauchat et al, 1998) and gsc (Broun et al, 1999), the ANTPclass msh and the RFamide neuropeptide genes (Mitgutsch et al, 1999). Their RT-PCR expression analysis in cnox-2(-) and control hydra showed a significant downregulation of prdl-a, gsc and RFamide-B, whereas hyCOUP-TF was found to be upregulated (Fig.…”

Neuronal evolution: analysis of regulatory genes in a first-evolved nervous system, the hydra nervous system

“…Riboprobes were labelled with either digoxigenin (DIG) or fluorescein following the supplier recommendations (Roche) and in situ hybridizations were performed on whole mount animals, following the procedure described in Grens et al (1995). Double in situ were performed according to Mitgutsch et al (1999). During the NBT/BCIP and/or Fast Red (Sigma) staining, appearance of the gene expression patterns was followed on Stemi SV11 stereomicroscope (Zeiss).…”

Peduncle of Hydra and the heart of higher organisms share a common ancestral origin

“…The effect of Hydra RFamide I-III (ϽQWLGGRFa, ϽQWFNGRFa, KPHLRGRFa) (Mitgutsch et al, 1999) on the contractile activity was examined by treating animals with the peptides at 10 -6 , 10…”

“…Neurons present in both the tentacles and the peduncle express a gene, preprohormone A, that encodes the peptides, Hydra RFamide I-IV (ϽQWLGGRFa, ϽQWFNGRFa, KPHLRGRFa, and HLRGRFa) (Mitgutsch et al, 1999). Preprohormone B encoding RFamide I and II and preprohormone C encoding RFamide I is expressed in hypostomal and tentacle neurons, respectively.…”