Visual pigments in a living fossil, the Australian lungfish Neoceratodus forsteri

Background. One of the greatest challenges facing the early land vertebrates was the need to effectively interpret a terrestrial environment. Interpretation was based on ocular adaptations evolved for an aquatic environment millions of years earlier. The Australian lungfish Neoceratodus forsteri is thought to be the closest living relative to the first terrestrial vertebrate, and yet nothing is known about the visual pigments present in lungfish or the early tetrapods. Results. Here we identify and characterise five visual pigments (rh1, rh2, lws, sws1 and sws2) expressed in the retina of N. forsteri. Phylogenetic analysis of the molecular evolution of lungfish and other vertebrate visual pigment genes indicates a closer relationship between lungfish and amphibian pigments than to pigments in teleost fishes. However, the relationship between lungfish, the coelacanth and tetrapods could not be absolutely determined from opsin phylogeny, supporting an unresolved trichotomy between the three groups. Conclusion. The presence of four cone pigments in Australian lungfish suggests that the earliest tetrapods would have had a colorful view of their terrestrial environment

In vitro and in vivo MMP gene expression localisation by In Situ-RT-PCR in cell culture and paraffin embedded human breast cancer cell line xenografts

BACKGROUND: Members of the matrix metalloproteinase (MMP) family of proteases are required for the degradation of the basement membrane and extracellular matrix in both normal and pathological conditions. In vitro, MT1-MMP (MMP-14, membrane type-1-MMP) expression is higher in more invasive human breast cancer (HBC) cell lines, whilst in vivo its expression has been associated with the stroma surrounding breast tumours. MMP-1 (interstitial collagenase) has been associated with MDA-MB-231 invasion in vitro, while MMP-3 (stromelysin-1) has been localised around invasive cells of breast tumours in vivo. As MMPs are not stored intracellularly, the ability to localise their expression to their cells of origin is difficult. METHODS: We utilised the unique in situ-reverse transcription-polymerase chain reaction (IS-RT-PCR) methodology to localise the in vitro and in vivo gene expression of MT1-MMP, MMP-1 and MMP-3 in human breast cancer. In vitro, MMP induction was examined in the MDA-MB-231 and MCF-7 HBC cell lines following exposure to Concanavalin A (Con A). In vivo, we examined their expression in archival paraffin embedded xenografts derived from a range of HBC cell lines of varied invasive and metastatic potential. Mouse xenografts are heterogenous, containing neoplastic human parenchyma with mouse stroma and vasculature and provide a reproducible in vivo model system correlated to the human disease state. RESULTS: In vitro, exposure to Con A increased MT1-MMP gene expression in MDA-MB-231 cells and decreased MT1-MMP gene expression in MCF-7 cells. MMP-1 and MMP-3 gene expression remained unchanged in both cell lines. In vivo, stromal cells recruited into each xenograft demonstrated differences in localised levels of MMP gene expression. Specifically, MDA-MB-231, MDA-MB-435 and Hs578T HBC cell lines are able to influence MMP gene expression in the surrounding stroma. CONCLUSION: We have demonstrated the applicability and sensitivity of IS-RT-PCR for the examination of MMP gene expression both in vitro and in vivo. Induction of MMP gene expression in both the epithelial tumour cells and surrounding stromal cells is associated with increased metastatic potential. Our data demonstrate the contribution of the stroma to epithelial MMP gene expression, and highlight the complexity of the role of MMPs in the stromal-epithelial interactions within breast carcinoma

A phylogenetic tree of the five photoreceptor opsins of and selected full-length nucleotide coding sequences of related species

<p><b>Copyright information:</b></p><p>Taken from "Visual pigments in a living fossil, the Australian lungfish "</p><p>http://www.biomedcentral.com/1471-2148/7/200</p><p>BMC Evolutionary Biology 2007;7():200-200.</p><p>Published online 25 Oct 2007</p><p>PMCID:PMC2194722.</p><p></p> The tree was constructed using the Neighbour-joining method with 1000 bootstrap replications [39]. Sarcopterygian fish (coelacanth and lungfish) are in red, agnathan fish (lamprey) are in light blue, teleost fish are in dark blue, amphibians are in green and reptiles are in purple. Genbank accession numbers are listed in Table 1. Bootstrap confidence values are at the base of each node. The rh4 opsin of (Table 1) was used as an outgroup. Scale bar indicates nucleotide substitutions per site

<it>In vitro </it>and <it>in vivo </it>MMP gene expression localisation by <it>In Situ</it>-RT-PCR in cell culture and paraffin embedded human breast cancer cell line xenografts

Abstract Background Members of the matrix metalloproteinase (MMP) family of proteases are required for the degradation of the basement membrane and extracellular matrix in both normal and pathological conditions. In vitro, MT1-MMP (MMP-14, membrane type-1-MMP) expression is higher in more invasive human breast cancer (HBC) cell lines, whilst in vivo its expression has been associated with the stroma surrounding breast tumours. MMP-1 (interstitial collagenase) has been associated with MDA-MB-231 invasion in vitro, while MMP-3 (stromelysin-1) has been localised around invasive cells of breast tumours in vivo. As MMPs are not stored intracellularly, the ability to localise their expression to their cells of origin is difficult. Methods We utilised the unique in situ-reverse transcription-polymerase chain reaction (IS-RT-PCR) methodology to localise the in vitro and in vivo gene expression of MT1-MMP, MMP-1 and MMP-3 in human breast cancer. In vitro, MMP induction was examined in the MDA-MB-231 and MCF-7 HBC cell lines following exposure to Concanavalin A (Con A). In vivo, we examined their expression in archival paraffin embedded xenografts derived from a range of HBC cell lines of varied invasive and metastatic potential. Mouse xenografts are heterogenous, containing neoplastic human parenchyma with mouse stroma and vasculature and provide a reproducible in vivo model system correlated to the human disease state. Results In vitro, exposure to Con A increased MT1-MMP gene expression in MDA-MB-231 cells and decreased MT1-MMP gene expression in MCF-7 cells. MMP-1 and MMP-3 gene expression remained unchanged in both cell lines. In vivo, stromal cells recruited into each xenograft demonstrated differences in localised levels of MMP gene expression. Specifically, MDA-MB-231, MDA-MB-435 and Hs578T HBC cell lines are able to influence MMP gene expression in the surrounding stroma. Conclusion We have demonstrated the applicability and sensitivity of IS-RT-PCR for the examination of MMP gene expression both in vitro and in vivo. Induction of MMP gene expression in both the epithelial tumour cells and surrounding stromal cells is associated with increased metastatic potential. Our data demonstrate the contribution of the stroma to epithelial MMP gene expression, and highlight the complexity of the role of MMPs in the stromal-epithelial interactions within breast carcinoma.</p

A clade credibility tree showing the relationships between lungfish and other selected vertebrate opsins

<p><b>Copyright information:</b></p><p>Taken from "Visual pigments in a living fossil, the Australian lungfish "</p><p>http://www.biomedcentral.com/1471-2148/7/200</p><p>BMC Evolutionary Biology 2007;7():200-200.</p><p>Published online 25 Oct 2007</p><p>PMCID:PMC2194722.</p><p></p> The tree was generated using Bayesian inference a Metroplis-coupled Markov chain Monte Carlo simulation. Sarcopterygian fish (coelacanth and lungfish) are in red, teleost fish are in blue, amphibians are in green and reptiles are in maroon. Genbank accession numbers are listed in Table 1. Posterior probability values are at the base of each node. The rh4 opsin of is used as an outgroup (Table 1). The probability of most relationships within the tree is 1.00 after 300,000 generations, while lower posterior probability values are found within the lws and rh1 groups