Green Fluoresence Protein
I’ve seen this green color in the past. It seems relatively obvious now that the green color that we are looking at in the reefs is wild Green Fluorescing Protein (GFP)1. It is one of the most important biologic markers used today. It was initially discovered in a jellyfish, Aequoria Victoria, and reef polyps. Its development as a synthesized reagent was awarded the Nobel prize. It is one of a class of reagents that radiate fluorescent green, red, yellow and blue when exposed to special wavelengths of light. They are used as bioluminescent markers in biochemistry and cell biology in a wide array of research and industry. I used this marker some years ago when using molecular imaging to search for circulating tumor cells (CTC) at the Barbara Ann Karmanos Cancer Institute, Tumor Biology and Microenvironment Program, the School of Medicine and the Bioengineering departments of Wayne State University and the associated NIH National Cancer Institute in Detroit.
Here are two examples of green and blue fluorescent proteins seen in two different coral species as they respond differently to the same blue light. Note that the GFP reef corals are stricken with SCTPD while the red fluorescent protein (RFP) reef coral seems to be unaffected.
The colorful green bioluminescence occurs when the GFP is exposed to a particular energy of blue light. The light excites electrons in the molecule which, when they return to their rest state, emit green light. In this case blue light exposure radiates green fluorescence. Cells continually manufacture the florescent protein for replacement as it matures.
Wouldn’t it be ironic to think that the polyps that gave us the GFP may help us to understand the cause of their disease?
I’ll make this very clear. This is just my conjecture. I am not an expert in any of the following issues. This is the realm of highly sophisticated experts in multiple specialized fields.
Thoughts on tracing down an idea for cause and effect
There may be a relation between protein structure and spectroscopic function. Loss of fluorescence may indicate that the GFP molecule is misfolded, split or destroyed. There are some things that can cause these molecules to stop performing including: insufficient molecular oxygen, increase in temperature above 70 degrees Celsius (158 F); exposure to an acid with a pH less than 5.4, age degradation, or protein modification. Some of these are not likely because the water temperature, acidity, and the rate of aging of the reef in the ocean are relatively constant. Temperature, acidity and aging is not what is at work in the polyps or in the reef. What is not constant is a biologic process which may alter the GFP. An virus could invade a cell and change a proteins including GFP. Lack of fluorescence may be a direct symptom of a disease. GFP labels living cells therefore it can be used to monitor infectious processes in plants and animals.3 GFP behavior part of a larger process causing death of the polyp complex
Is there a direct correlation between the lack of the GFP and the cause of SCTLD?
Establishing a causal relationship. Is the disease infectious:
In an earlier posts Fluorescent Photos to Measure Reef Viability (2) and Hunting the Cause of Stony Coral Tissue Loss. Looking at the Reefs with Scientific Vigor (3) I suggested a test of infection showing a controlled method of applying the diminished green polyps or BFP (blue fluorescing protein) polyps to GFP polyps. If the trial infection works then the cause is either a bacteria or a virus. This may depend upon identification and use of a vector organism which may be critical for the test to work.
If there were a virus in a polyp or associated zooxanthellae, a comparison of DNA before, during and after exposure could be made. Sample both at various stages of the disease using the GFP as an indicator of the staging. Additionally, special techniques of microscopic dissection, staining and a variety of light and electron microscopy should show some evidence. The polyps and the zooxanthellae are higher level organisms and their DNA should to be decoded. If the disease is virus specific, PCR testing may be done to discover it. DNA sequencing and Crisper technology might be used to find a place in the DNA where a lethal virus could attach.
Discussion of findings:
Crisper technology could be used to modify the DNA to engineer a resistant strain of stony coral. The strain could be cloned and reproduced in vast colonies. Multiple coral polyp types could be added to make a more representative colony. The harvested viral resistant polyp strains could be explanted back onto the dead wild coral reefs that were cleaned of dead coral tissue, algae and debris. These are all untried technologies.
Spread of the disease:
Polyps are filter feeders. They harvest their sustenance from the organisms in the float of water of the ocean directly into their gut. This sounds like a combination of opportunity: portal of entry, susceptible host, and an obviously virulent pathogen. Viruses are frequently carried by arthropods (insects) that can spread the virus in the viral particle of their bodies. They inject the particle into host cells using their sharp, piercing or cutting anatomy as do mosquitos, ticks, flees and others. Examples of this include Zika, West Nile fever, Deer Tick fever. Viruses can also spread by contact through the air in water droplets. (Such as the Corona virus). An arthropodal nymph acting as a vector could easily be carried by currents, ships and wind for hundreds or even thousands of miles where it could start another colonial infection.
There may be a number of contributing factors such as rising water temperatures, pollution, habitat destruction, water depth, invasive species, symbiotic species relationships which may exacerbate the process.
Search for complicating factors:
- How long does GFP continue to fluoresce without replacement
- Discover the virus pathogen and its structure
- Unravel the complexities of the method of action of the pathogen
- Geographically locate areas of afflicted coral
- Calculate the rate of spread, rate of infection, time from onset to death
- Identify the origin of the disease and the vector
- Identify potential causes of increased susceptibility of the reefs
- Search geological and anthropological history for episodes of similar reef diseases
Discussion on actions:
- Consider possible unintended consequences of well intended actions
- Search for other possible causes such as prions, rickettsiae and other outliers.
- Form a team of experts in the necessary fields
- Estimate the cost of the project in stages
- Form a group of interested parties of influence
- Promote the idea seeking funding and permission to act on findings.
- Begin the search for the cause and form an action plan
- Begin at a fixed base location with relative separation from recreational or commercial activity
- Start small with two isolated sites each with differing characteristics in reef health
- Engage in work to slow or stop SDTLD
Not much is known about reef ecosystems, polyps and their physiology, diseases, and pathogens. Additionally, they are part of a larger oceanic system of which we have limited knowledge. The current blog is biased by knowledge of land based diseases. There may be causes of which we are completely unaware. It is a voyage of discovery aboard Everglades Ark.
I am very interested in your thoughts on the subject of reefs, polyps, fluorescence, viral biology, or any of the topics which are presented. Additionally, if you are interested in helping or would like to report on your findings please contact me.
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- The GFP page
- Fluorescent Proteins
- Split green fluorescent protein as a tool to study infection with a plant pathogen, Cauliflower mosaic virusFluorescent protein microscopy
- Algorithms and Datasets for Colour Science
- Spectrum or excitation and emission of protein fluorescence microscopy
#RFP #GFP #fluorescing protein #vector #SDTLD #stony coral tissue loss disease #virus #crisper #spectroscopy #polyp #crisper #DNA