The Wnt pathway! http://web.stanford.edu/group/nusselab/cgi-bin/wnt/reporters. Play the video to see a baby Zebrafish expressing Tcf/Lef-miniP:dGFP.
My post today will be covering the weekend as well as today, so it will be appropriately long. The last few days have been quite busy as we work through all the sea star transcriptome data and got to participate in the RCN meeting on Marine Infectious diseases. It was an honor meet so many scientists focused on disease ecology and to hear many great talks over the weekend.
I have struggling with posting every day since I like to have a complete story to tell when I write, however, looking back at my posts and those of my classmates I can see what a great tool it will be moving forward. This is something I will be working on in the future to incorporate into my daily note taking. It is different to post with an audience in mind, but good I think. My work is always better when I need to explain it to someone else and that is how this feels, just more permanent.
The last few days I have focused on the significantly enriched sea star genes falling within the “adhesion” biological function category. I took a subset of the genes, and Reyn and Ruth looked at the other. For my set, I went through each one and looked up the function of the protein it encoded and other proteins associated with it. It was difficult to find papers focusing on echinoderms for all of the genes, but for the ones that could find them for I did. I noted if the gene was higher of lower expressed in the control vs. experimental animals, what the value of the log change was, and what the p-value was.
Clear grouping of gene functions started to turn up right away. There were several genes involved in the extracellular matrix and another set in mutable collagenous tissue. Here is the list of genes I looked at with notes on the reference relating to them and a summery of their function taken from UniProt: Adhesion-ME References listed
After going through the list I grouped the genes based on function (highlighted in the doc) and started looking up papers based on the potential interacting pathways. While the Wnt pathway was not well represented in my list I had many genes that were regulated by it so started searching for the related Wnt genes in the lists of immune and cytokine genes. In the diseased animals there is a general lowering of expression relative to the controls in Wnt and Frizzled, and an increased expression of genes associated with the Adherin Junctions (cell-cell adhesion). The Wnt/beta-Catenin pathway regulates the extracellular matrix through activation of beta-Catenin, but is also involved in mutable collagenous tissue through beta-Catenin mediated integrin signaling. The disruption of the Wnt/beta-Catenin pathway is associated with disease in vertebrates. A disruption of this pathway is seen in many types of cancer. It is well characterized in echinoderms due to the sea urchin genome. I then compared the list of genes I had hand curated to the genes discussed in Croce_2006_EvolutionConservedWntPathways_SeaUrchin. Since this paper describes only the genes found and characterized in the sea star genome there is more confidence that these are coding for a protein in the Wnt/beta-Catenin pathway. My next step was to search all the significant enriched genes on the list against those mentioned in Croce et al 2006.
So…what is going on in our challenge experiment Pycnopodias? The table below contains the list that I have evaluated and placed as a likely part of the wnt pathway or linked to through the literature. I have started to map them onto a pathway (human) of the wnt/beta-catenin pathway. I will wait until that is more complete before uploading it. There are several more wnt genes found in the immune category that I have yet to look at in relation to the ones I have here to see where they fit. A few of the ones here (the 2 spondins in particulare) have literature on them but I have not determined if it is relevant to echinoderms. This list does not reflect the interactions but the one I will post tomorrow will.
|log2FoldChange||pvalue||ID||Protein names||Broad Group||Found in Croce et al 2006?|
|6.69463||8.14E-21||Q8IUX8||Epidermal growth factor-like protein 6||Adhesion||no|
|4.483540943||1.79E-06||Q9QYP1||Low-density lipoprotein receptor-related protein 4||Immune||no|
|2.939162||4.26E-06||Q9Y2D8||Afadin- and alpha-actinin-binding protein (ADIP)||Adhesion||no|
|2.638696126||2.61E-10||Q99N43||Kremen protein 1||Immune||yes|
|2.178693||7.64E-04||Q9C0A0||Contactin-associated protein-like 4||Adhesion||no|
|2.147782||4.10E-03||Q5RD64||Contactin-associated protein-like 2||Adhesion||no|
|1.539407||0.001994||O14522||Receptor-type tyrosine-protein phosphatase T||Adhesion||no|
|-1.41984295||0.006131503||Q8NCW0||Kremen protein 2||Immune||Yes|
|-1.930118114||0.003009238||Q6FHJ7||Secreted frizzled-related protein 4||Immune||yes|
|-3.66822||5.43E-07||Q9VBW3||Tyrosine kinase receptor Cad96Ca||Adhesion||no|
Clevers_2006_WntBetaCateninSigDisease covers wnt/beta-catenin signaling involvement in human disease. Deregulation of the system leads to cancers in many cases. Another paper on inhibitors and activators is particularly interesting since it several of the genes I have identified are inhibitors (Cruciat_2013_WntInhibitors&Activators).
After completing the analysis of the wnt gene I will look at the adhesion set of genes. The two are linked so there will be overlap in the interacting proteins. The first step in looking at this (as with the last set) will be to find as many of my significantly enriched genes as I can that are associated with echinoderms. The echinoderm adhesome is described here: Whittaker_2006_TheEchinodermAdhesome.
The last step is to merge to the two sets of interacting proteins that I am most confident in (from the echinoderm literature) so the connections between them become clearer. After that I will add the genes annotated to other organisms to fill in where they fit.