This may very well be my very last notebook post.. I know, so sad. The paper is coming together wonderfully and we pretty much have way too many awesome stories to do them all justice (TLRs, Complement cascades, WNT signaling, the list goes on). But I suppose that’s a good problem to have. Everyone’s presentations this morning were wonderful. It’s great to know that we all got such awesome projects done in such a short time. I’m going to miss FHL, and all of my wonderful classmates (I have to be sappy and include this). Looking forward to continued collaborations with you all!!
So it’s been a few days since I last posted (whoops), but I’ve been busy writing up what we have for the sea star transcriptome and in particular what complement cascade genes we have. Echinoderms were already described as having two proteins involved in the complement cascade: C3 and C2/Bf, however we found two more which were significantly deferentially expressed: Ficolin-2 and Properdin. Both of these are proteins involved in alternate activation of the complement cascade system. Looks like the infected stars are definitely responding to something though we’re still working at getting the full picture of how they’re responding and to what.
So since I keep forgetting to post double to get caught up on notebooks, I’m just going to post today and call it even! Yesterday was a mind-blowing day full of ridiculous amounts of information, all of which was extremely fascinating. It was great to get to sit in on the RCN and see so many diverse researchers come together to share their work and move forward not only in terms of research but also in terms of policy and public awareness. I was a environmental studies minor in college so I took a lot of classes which challenged me to integrate my scientific knowledge with that of students in the social sciences, but I’d been away from that kind of train of thought for a while. Needless to say yesterday was a great throw back to those days and I can’t wait to see what awesome products come out of this meeting.
So yesterday was a day of planning, lots of planning, for our upcoming talk at this weekend’s RCN. The group met back together to try and come up with a cohesive story to relay on Sunday following our extensive gene investigations. Some interesting notes from that meeting: it appears we have a lot of components of many immune-involved signaling pathways, including the TLR (sea urchins have been found to have about 150 TLRs!!!!), Complement Cascades, (C3 is well described in the Echinoderms) and Interleukins (1 and 6 have been found in Echinoderms before!). There are also lots of interesting neural and skeletal things going on (mostly with collagen) which could be important to the actual wasting part of the disease. I think we have a good game plan for the talk. After our meeting I worked on making graphs presentation ready and continuing to search through literature for an understanding of more DEGs involved in enriched GO terms. We’re at that point where transcriptomics becomes a ton of research to find out what all the lovely data you generated actually means. Today’s task: RCN!
So I got so caught up investigating genes last night I totally forgot to post! So here’s a recap of what I did yesterday! In the morning, Maya gave an awesome lecture/exercise on the applications of modeling. I definitely have a lot of respect for anyone who can both do the mathematics it takes to build models and the java scripting required for that crazy AnyLogic software. While I might not have the required skills to actually build models myself, Maya’s talk definitely got me thinking about collaborating more with people who can model in order to expand the types of questions my research can answer.
In the afternoon, team transcriptomics divided up our three groups of significantly enriched GO terms. We then started to work through each annotated gene individually to try and take all this crazy awesome data we have and make a story out of it. I’ve been working through the immune genes and there seem to be a lot of complementary interleukin and complement cascade proteins which are more expressed in infected sea stars- AWESOME! It’s cool to find multiple parts of the same pathways in our list too. Hopefully we should have the beginning of a great story to tell the RCN this weekend!
So today I thought I did a lot of work, but I have little to nothing to show for it. I continued my explorations of the DEGs today, focusing on those with greater than 2 or less than -2 fold change. I further focused in on finding DEGs with links to neurological or skeletal (cytoskeletal, collagen, cartilage, muscular, etc) functions that could explain the physical wasting of infected sea stars. After sifting through hundreds of Swiss-Prot IDs and becoming a pro at scanning the entries for important connections/terms, I have returned a narrowed down list of 82 contigs involved in neurological processes and 85 involved in the broad skeletal category. 8 contigs are involved in both groups. Obviously these lists will likely need to be narrowed down further for the discussion portion of the manuscript, but its a start. Of interesting note: there are a lot of collegenase and similar proteins, which degrade connective tissues, among those genes with high log2 fold change. Could explain why stars go from being rigid to mushy. More thoughts to come as we continue to mine the data!
So today I spent a really long time working more with my master list of DEGs from yesterday. First I worked to make some graphs for each of the significantly enriched GO terms (Fig. 1). After that I began to notice that several contigs shared the same exact annotation, down to the spID. So I made a master list of all of those. It’s something like 150 contigs! I plan on exploring that more later this week to determine whether they are the result of gene duplication or alternate splicing. What’s weird is sometimes these contigs with identical annotations will have vastly different logfold changes! If anyone know what might be going on, please let me know! Finally, after lunch I began to manually go through the annotations looking for interesting genes that weren’t previously graphed. These include genes involved in neurological processes and skeletal functions (melting and twisting explanations perhaps?) and protein ubiquitination. This will likely be a several day process, so I’ll update more when I get further into it. For now I’m only looking at genes with a logfold change of at least 2 up or down. Team Enrichment let me know if you have an issue with this. No notebook today as this was all in excel. More results soon!
Here’s what I did this morning. Team transcriptome please look and check!!
So today I focused on two things: the first was a WGCNA tutorial for the class this morning which took a while and involved a lot of fruitful parameter tweaking (though the model fit was awful). Hopefully it’ll be a useful approach moving forward to really understand our full transcriptome. Stay tuned for updates. Then this afternoon I worked on really looking over the DEGs for anything interesting. I found a ton of neurological and cytoskeleton/skeleton associated genes in the 75 or so I got through that could be super interesting. I’ll keep doing this later next week. See my notebook below and here to look at bigger versions of the figures. Happy weekend and exam cramming!
Today was a fun day of scripting and file manipulation to try and get to the end result: log change of genes of interest! I ended up looking at some genes in our most enriched GO terms and at the top 10 up and down genes in our exposed sea stars. See below for all the pretty data! And click here to look at bigger versions of the graphs.
- Hepranase degrades heprane sulfate, which can be a viral receptor…ooh! Maybe it’s down for increased viral recognition??
- Norepinephrine is down..possibly having neurological consequences?
- Quinone oxoreductase- melanin!?
Yeah that’s it.. my brain’s dead. Let me know your thoughts.