FLS2

"FLS" genes have been discovered to be involved in flagellin reception of bacteria. FLS1 was the original gene discovered shown to correspond with a specific ecotype within "Arabidopsis thaliana". Even so, further studies have shown a second FLS gene known as FLS2 that is also associated with flagellin reception. "FLS2" and FLS1 are different genes with different responsibilities, but are related genetically. "FLS2" has a specific focus in plant defense and is involved in promoting the MAP kinase cascade. Mutations in the "FLS2" gene can cause bacterial infection by lack of response to flg22. Therefore, "FLS2"’s primary focus is association with flg22 while its secondary focus is the involvement of promoting the MAP kinase cascade in plant defense.
"FLS2" and its similarities to receptor-like kinases HOLAKHEASEN.
"FLS2" is considered a receptor like kinase (RLK). RLKs are very similar to tyrosine kinases but different in the fact that tyrosine kinases are present in mammals and RLKs are plant specific. RLKs are transmembrane proteins that consists of inner, outer, and central membrane regions. Being proteins, they also have an amino-terminus domain and a carboxyl-terminal kinase domain. RLKs play an important role in plant reception which will be shown through "FLS2". BRASSINOSTEROID INSENSITIVE 1 (BRI1) is another important RLK with a lot of similarities to FLS2. Despite share many of the same signalling components, FLS2 and BRI1 have very different effects on the cell.
"FLS2" Structure.
"FLS2" consists of 3 domains: an extracellular, a transmembrane, and an intracellular. The extracellular domain is known as the Leucine-rich repeat (LRR) domain. It is in this region, which is the amino-terminus, where it is said to have direct interaction with flagellin initiating the response of "FLS2" to flg22. The transmembrane domain is where proteins transition from extracellular to intracellular. This region is usually very thermodynamically stable and occurs only in the phospholipid membrane between cells. The intracellular domain is the serine/threonine kinase domain. In this domain, phosphorylation catalyzes a protein kinase cascade leading to a response. In "FLS2", this response elicits changes in growth and plant defenses.
Ectopic Expression of "FLS2".
One experiment performed to show involvement of "FLS2" in flagellin perception was an ectopic expression of "FLS2" using the CaMV 35S gene promoter in one wild type "Arabidopsis thaliana" plant and two supposed mutant "Arabidopsis thaliana" plants. The wild type "Arabidopsis" plant was indicated by Col-0 and the two mutants were indicated by 35S::"FLS2"-"col2" and 35S::"FLS2"-"col4". In this experiment, callose deposition and production of active oxygen species were the variables considered to show flagellin perception after treatment of flg22 as indicated by previous research (Gomez-Gomez, et al. 1999). Results for this study showed a few things. The first thing it showed was a response in callose deposition 12 hours after treatment in only one plant type which was the mutant 35S::"FLS2"-"col2". In the wild type and mutant 35S::"FLS2"-"col4", no callose deposition was detected in plants that exhibited reduced levels of "FLS2". In direct correlation to the callose deposition response, the second thing observed was a luminol-based assay to show oxidative burst in leaf tissues of the three plants types. The luminol-based assay showed a rapid and strong oxidative response in mutant 35S::"FLS2"-"col2" with treatment of 10nM flg22, whereas a 10nM treatment of flg22 in the mutant 35S::"FLS2"-"col4" was unable to induce any oxidative response that would occur naturally in the wild type "Arabidopsis" plant. Even so, higher treatment levels of flg22 would elicit an oxidative response but not significant compared to the wild type. The last thing observed in this experiment was treatment of flg22 in regards to growth inhibition to T2 seedlings. Oddly enough, growth inhibition in 35S::"FLS2"-"col2" was greater than the wild type under flg22 treatment, whereas 35S::"FLS2"-"col4" showed a reduced growth inhibition compared to wild type plants under flg22 treatment which is inverse of the oxidative response in these two mutants relative to the wild type.
Involvement of "FLS2" in Defense responses.
In the same way that mammals and insects have an innate immunity response to pathogens, plants have a defense system that is similar yet modified. The biggest difference between plants, mammals, and insects is that mammals and insects are not stationary like plants, therefore a plant’s defense system is more complex and more reliant of signaling pathways to notify the plant of what could be potentially attacking. With that in mind, it is important that all gram positive and gram negative bacteria share flagellin. With plants being stationary, a specific molecular pattern of flagellin can be recognized by plants to determine phytopathogenic bacteria in plant defense. As discovered by the ectopic expression of "FLS2" study, "FLS2" is directed related to flagellin perception, so by discovery and by natural way of the plant, "FLS2" is the gene responsible for perception of flagellin which signals a plant defense response. Things are not all settled here because, as mentioned earlier, gram positive and gram negative bacteria share flagellin which means that plants cannot distinguish between avirulent and virulent bacteria. This could provide extremely useful when an abundance of pathogenic bacteria are present, however this can also prove inefficient when plant defense response is occurring due to flagellin perception of avirulent bacteria, or so one would think. In actuality, avirulent bacteria defense responses are higher than virulent bacteria defense response due to the presence of multiple specific elicitors. Hutcheson (1997) found that the tertiary and quaternary structures of protein elicitors are dependent on elicitor activity. Even so, resistant gene products are commonly found in Leucine rich repeat domains which is the extracellular domain of "FLS2", and "hrp" genes are directly related to disease incitement and hypersensitive response to resistant plants. In short, the more elicitors that are present within flagellin perception, the more plant defense response will occur regardless of virulence. It is just more common for avirulent pathogen to produce more elicitors.
Expression of "FLS2" from "Arabidopsis" in Tomato Cells.
The design of this experiment is similar to previous literature from and but unlike those two publications, in this publication, the promoter used was the 35S cauliflower mosaic virus promoter used to express "FLS2" coding sequences fused to a triple c-myc tag. What happens to a "FLS2" gene in "Arabidopsis" is known but this study looks at how the "FLS2" gene in "Arabidopsis" may affect a plant that contains a "FLS2" gene that does not respond similarly to "Arabidopsis". Figure 1 in shows how tomato cells with no introduction to an "Arabidopsis FLS2" gene show no response to the perception of either flg15 or flg22. However, when viewing the "Arabidopsis" "FLS2" gene and the tomato cells with the "Arabidopsis FLS2" gene added, it is clear that the responses are similar due to the presence of the "Arabidopsis" "FLS2" gene.
Alkalinization Response Induced by the Flagellin Derivative flg22-ΔA16/17.
Alkalinization has been used to show ion fluxes across membranes in previous literature. In this study, alkalinization induced by the flagellin derivative flg22-ΔA16/17 in tomato plants expressing the "Arabidopsis FLS2" gene. Extracellular alkalinization was used to elicit a response in these type plants. Alkalinization had no effect on non-treated tomato plants, but increased binding activity to "FLS2" was shown with no increase in sensitivity toward flg22 in the tomato plants expressing the "Arabidopsis" "FLS2" gene. This lack of increase in sensitivity could be attributed to a number of factors that are unknown, yet hypothesized. Even so, the binding of tomato plants expressing "Arabidopsis" "FLS2" gene showed similar sensitivity of flg15 to that of normal tomato plants. This indicates that both the "Arabidopsis FLS2" gene and the tomato plant "FLS2" gene are both working in tomato plants expressing "Arabidopsis" "FLS2" gene. The "Arabidopsis FLS2" gene responds to flg22 and the natural tomato "FLS2" gene responds to flg15. Furthermore, the tomato "FLS2" gene was analyzed for any derivatives that might be responsible for gene expression. Although less active than flg22, its derivative flg22-ΔA16/17 was found to elicit a medium alkalinization response in "Arabidopsis FLS2" gene in both "Arabipdopsis" and tomato plants expressing "Arabidopsis" "FLS2" gene. Essentially, the majority of flagellin perception occurring in tomato plants expressing "Arabidopsis" "FLS2" gene is occurring because of the "Arabidopsis" "FLS2" gene while a minor portion of flagellin perception can be attributed to the tomato "FLS2" gene.