Metamorphosis, a critical stage in the development of toads and frogs, involves rapid levels of morphological change. In the current study, we have used microarray analysis to identify shifts in gene expression between tadpole and toadlet stages of the cane toad, Bufo (Chaunus) marinus. Here, we report on nine genes that show the greatest induction during metamorphosis; the gut-associated gastrokine and trefoil factor, blood components haemoglobins alpha/beta, apolipoprotein and serum albumin, a nasal gene olfactomedin, a lens gene gamma-crystallin, and a novel gene with low homology to frog harderin. We present both temporal and spatial expression patterns of these genes identified in developing and adult cane toads. This study extends our knowledge of the molecular basis of toad metamorphosis, and not only offers insights to the genes induced during the general remodelling that occurs but also reveals possible targets for control and manipulation of amphibian pest species, for example, the cane toad in Australia.
"Another goal for a microarray study was to identify genes upregulated during metamorphosis that could be used in a strategy to control the invasive cane toad, B. marinus (Halliday et al., 2008). The authors produced a 5170-spot cDNA array of PCR products from randomly selected clones from a toadlet cDNA library. "
[Show abstract][Hide abstract] ABSTRACT: A major goal for understanding the role of thyroid hormone (TH) in development has been to identify genes regulated by TH in different tissues during frog metamorphosis. The exquisite dependence of metamorphosis on TH also provides a model to study TH endocrine disruption. To identify such TH-regulated genes and select biomarkers for TH endocrine disruption, global gene expression analyses in tadpoles using microarrays have been done in 21 studies, involving five frog species, seven organs, and four endocrine disrupting chemicals. As expected, each organ has a unique set of genes associated with its tissue-specific metamorphic outcome, and functions ascribed to many of these genes correspond to histological changes induced by TH. Also, the large number of transcription factors identified in microarrays is consistent with the molecular mechanisms of TH action. On the other hand, microarray analysis has also revealed interesting findings not predicted from previous morphological or molecular studies. Furthermore, endocrine disruption studies identified candidate biomarkers for TH disruption, and the mechanisms of action of several endocrine disrupting chemicals have been examined. The microarray studies described here have produced a wealth of data on gene expression that requires further functional studies to elucidate the roles of these genes in development and endocrine disruption.
Current Topics in Developmental Biology 01/2013; 103:329-64. DOI:10.1016/B978-0-12-385979-2.00012-5 · 4.68 Impact Factor
"Cane toad genes that are expressed in metamorphs but not in tadpoles therefore represent ideal targets to block and thus manipulate aspects of development. One documented example of the transition from a larval to an adult form in amphibians is haemoglobin  and we have used microarray analysis to establish that adult haemoglobin is significantly upregulated during cane toad metamorphosis . Injecting tadpoles with adult globin interfered with expression of this protein in Rana catesbeiana, and induced changes in gene expression profiles of metamorphs . "
[Show abstract][Hide abstract] ABSTRACT: The cane toad, Bufo (Chaunus) marinus, is one of the most notorious vertebrate pests introduced into Australia over the last 200 years and, so far, efforts to identify a naturally occurring B. marinus-specific pathogen for use as a biological control agent have been unsuccessful. We explored an alternative approach that entailed genetically modifying a pathogen with broad host specificity so that it no longer caused disease, but carried a gene to disrupt the cane toad life cycle in a species specific manner.
The adult beta globin gene was selected as the model gene for proof of concept of autoimmunity as a biocontrol method for cane toads. A previous report showed injection of bullfrog tadpoles with adult beta globin resulted in an alteration in the form of beta globin expressed in metamorphs as well as reduced survival. In B. marinus we established for the first time that the switch from tadpole to adult globin exists. The effect of injecting B. marinus tadpoles with purified recombinant adult globin protein was then assessed using behavioural (swim speed in tadpoles and jump length in metamorphs), developmental (time to metamorphosis, weight and length at various developmental stages, protein profile of adult globin) and genetic (adult globin mRNA levels) measures. However, we were unable to detect any differences between treated and control animals. Further, globin delivery using Bohle iridovirus, an Australian ranavirus isolate belonging to the Iridovirus family, did not reduce the survival of metamorphs or alter the form of beta globin expressed in metamorphs.
While we were able to show for the first time that the switch from tadpole to adult globin does occur in B. marinus, we were not able to induce autoimmunity and disrupt metamorphosis. The short development time of B. marinus tadpoles may preclude this approach.
PLoS ONE 01/2011; 6(1):e14576. DOI:10.1371/journal.pone.0014576 · 3.23 Impact Factor
"However, to confirm such co-distribution within smooth muscle, we used gastrokine-1 polyclonal antibody in other tissues and found, in agreement with recent related data on genes induced during early development stages , that gastrokine was completely absent in skeletal muscle. However, also in agreement with this work, we found that gastrokine-1 was weakly present in bovine heart and, more particularly, targeted to Hiss fibers (personal data, not shown). "
[Show abstract][Hide abstract] ABSTRACT: The potential role and function of gastrokine-1 (GNK1) in smooth muscle cells is investigated in this work by first establishing a preparative protocol to obtain this native protein from freshly dissected chicken gizzard. Some unexpected biochemical properties of gastrokine-1 were deduced by producing specific polyclonal antibody against the purified protein. We focused on the F-actin interaction with gastrokine-1 and the potential role and function in smooth muscle contractile properties.
GNK1 is thought to provide mucosal protection in the superficial gastric epithelium. However, the actual role of gastrokine-1 with regards to its known decreased expression in gastric cancer is still unknown. Recently, trefoil factors (TFF) were reported to have important roles in gastric epithelial regeneration and cell turnover, and could be involved in GNK1 interactions. The aim of this study was to evaluate the role and function of GNK1 in smooth muscle cells.
From fresh chicken gizzard smooth muscle, an original purification procedure was used to purify a heat soluble 20 kDa protein that was sequenced and found to correspond to the gastrokine-1 protein sequence containing one BRICHOS domain and at least two or possibly three transmembrane regions. The purified protein was used to produce polyclonal antibody and highlighted the smooth muscle cell distribution and F-actin association of GNK1 through a few different methods.
Altogether our data illustrate a broader distribution of gastrokine-1 in smooth muscle than only in the gastrointestinal epithelium, and the specific interaction with F-actin highlights and suggests a new role and function of GNK1 within smooth muscle cells. A potential role via TFF interaction in cell-cell adhesion and assembly of actin stress fibres is discussed.
PLoS ONE 02/2008; 3(12):e3854. DOI:10.1371/journal.pone.0003854 · 3.23 Impact Factor
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