Healing Focus. Reason Lv 1. Reason Lv 2. Reason Lv 3. Reason Lv 4. Reason Lv 5. Black Tomefaire. Faith Lv 1. Faith Lv 2. Faith Lv 3. Faith Lv 4. Faith Lv 5. White Tomefaire. Authority Lv 1. Authority Lv 2. Authority Lv 3. Authority Lv 4. Authority Lv 5. Rally Luck. Battalion Vantage. Defensive Tactics. Offensive Tactics. Weight Armored Effect Null. Cavalry Effect Null.
Alert Stance. Flying Effect Null. A beautiful wood carving of the goddess. Appreciated by those who like art or carving, and by devout believers. A landscape painting of magnificent Lake Teutates in the clearing fog. Appreciated by those who enjoy nature or art. Beans meant to be ground and boiled into a hot beverage. Appreciated by those who enjoy bitter flavors. The type of meat is unclear, but a meat lover would appreciate it. A dish that tastes like the wilderness. Thick slices of meat covered with Noa fruit and grilled on a hotplate.
This popular dish sells out almost instantly. A perfect snack to go with your favorite drink. Spicy stew made with Teutates loach and turnips. The monastery's unique recipe features spices from Dagda. Hunks of rabbit meat are pickled in bacchus, skewered, and roasted over an open flame to create this flavorful dish. A Dagdan dish of raw fish and turnips pickled in a vinegar-based seasoning liquid.
Invention of a certain noble. A gourmet fish dish with Airmid pike, vegetables, and a sprinkle of expensive spices. Popular among nobles. Pheasant meat is pounded flat and fried. Can be served as a sort of sandwich, with cheese between two strips of meat.
Herring caught off the coast of Albinea, shredded and grilled in an earthenware pot with sliced turnips. A baked tart with stewed herring and Noa fruit mixed into the batter. Popular in Enbarr, the Imperial Capital. A simple dish. Airmid pike is pickled in vinegar and served with cabbage between two slices of bread.
Minced poultry and onions boiled with salt. The simple recipe lets high-quality ingredients speak for themselves. A gratin of bird meat topped with heaps of Gautier cheese, which is famous for its low fat content. It has a unique flavor. A dish of dried tomatoes, cabbage, chickpeas, and other vegetables, stir-fried with eggs.
Nutritious and very filling. Countless numbers are listed in this ledger. It probably belongs to someone deeply involved in commerce. A precious tool used for measuring the weight of objects. It probably belongs to someone in need of accuracy. It probably belongs to someone with a deep interest in rare things….
Iron Axe Night Sky. Vantage 1. Vantage 2. Vantage 3. Spur Res 1. Spur Res 2. Spur Res 3. Sabotage Atk 1. Sabotage Atk 2.
Sabotage Atk 3. Apotheosis Spear Rally Attack. Spur Spd 1. Drive Spd 1. Drive Spd 2. Joint Drive Spd.
Anna is a bright, cheerful young merchant, but her true nature is clouded in mystery. She's very fixated on money and is a girl of many talents, but it wasn't until Fire Emblem Awakening that she became a playable character. There's still a lot we don't know about her-she likes to keep everybody guessing! CYL3 saw the transition to different counting mechanisms involving versions of characters and ties, so change measured between CYL2 and CYL3 shouldn't be taken at face value.
Radiant Dawn. Three Houses. Fire Emblem anime. Fire Emblem: Mystery of the Emblem manga. Fire Emblem 0 Cipher. Elibe Disturbance. Kana - Shigure - Dwyer - Midori - Sophie. Byleth - Sothis. Yuri - Balthus - Constance - Hapi.
Flame Emperor - Death Knight - Metodey. Side Quest 8 Answers So, what's the best path to go with Miriel? Build 2 Answers. Ask A Question. Browse More Questions. Keep me logged in on this device. Forgot your username or password? User Info: Tisroero Tisroero 8 years ago 3 Aint 2 the one where you first meet her?
Starting this game now, but while it's downloading "Pre Thoughts". I started Awakening a few years ago but never finished. Advice for coming back. Good pairs for marriage? Where can I find Master Seals early in the game? Side Quest. After beating Victor and saving Anna, one of her new shops appeared on my map. The strange thing is, her shop disappeared after I purchased a rear item and exited the shop.
The first time you get to see how Anna fights is in Paralogue 2 of the game. She fights off bandits, trying to protect the villagers while you and your party members try to defeat Victor and his men.
She can take a hit and knock out an opponent. One thing she surely is, is overconfident. There were many times she fell in battle. Most of the times she picked the fight that she just could not win. Yes, she died during the next phase. It took me over 10 restarts to come up with a good strategy to try and save her.
After, it took me over 25 restarts to actually save her. Let Frederick and another powerful character team up with both of those characters to march towards Anna.
Sumia could have helped, but the archers are too dangerous. Three of the polymorphic positions, with one being non-synonymous, were found in the French populations. In the Portuguese populations, 10 mutations out of 30 were non-synonymous. Five polymorphisms were shared with those previously described for rFut2 and rSec1 , while two where shared only with rFut2 [ 23 ].
According to the amino acid variation, two and eleven haplotypes or variants were inferred for the French and Portuguese populations, respectively Table 2. No shared variants were detected between these two groups of populations. Variant 2 was the most common in France with a frequency of 0. For amino acid position , three different amino acids were detected: S, P and Q. The low genetic diversity observed for the French populations when comparing with the Portuguese populations is in accordance with what has been shown for other genetic markers [ 40 , 41 , 42 ].
The protein domains are represented by black boxes tm, transmembrane domain; S, stem region. Amino acid variations are shown below boxes. Positions numbering begins at the start codon and first methionine respectively.
The positions in bold refer to the polymorphic positions shared by Sec1, Fut2 and Fut1 positions , , , and and to the polymorphic positions shared by Fut2 and Fut1 positions and Asterisks represent non-synonymous substitutions. B; C Flow cytometry analysis of wild rabbit rFut1 variants. Transfection of a human null allele hFUT2 KO fused to GFP provided a negative control and transfection of the rFut2 allele variant E corresponding to the rFut2 reference sequence provided a positive control.
The upper panels show examples of results obtained with two selected rFut1 variants. In each case, the activity of rFut1 variants is compared to that of the reference rFut2 variant E.
After cloning in an expression vector, rFut1 variants 1, 2, 4, 6, 8, 9, 10, 12 and 14 were tested for their catalytic activity. Variants 5 and 7 were not tested since the amino acid variation described above was almost completely covered by the other variants. Variant 11 was not tested since cloning of the full coding sequence failed despite several attempts. CHO cells do not express type 1 precursor and therefore synthesis of H type 1 was not measured.
The presence of a GFP tail in the recombinant enzymes allowed for control and normalization of transfection and protein expression efficiencies. Synthesis of H type 2 was similar for all variants tested and much higher than for the rFut2 enzyme rFut2 variant E used as a positive control Fig 5B. By contrast, synthesis of H type 3 was either low or undetectable, whereas it was clearly visible for the rFut2 control Fig 5C.
Since, as described above, rFut1 and rFut2 mRNA are expressed at similar levels in the duodenum, these results indicate that rFut1 should be the main contributor to H type 2 synthesis and therefore to RHDV binding sites.
However, the fold variation in rFut1 and rFut2 mRNA expression appeared quite limited compared to the extreme individual variation found in rSec1 mRNA levels that varied over fold Fig 3. To this aim, six RHDV strains representative of each virus genotype were used [ 36 ]. No relationship was observed between either rFut1 or rFut2 expression.
Binding to individual duodenum extracts of six strains of RHDV representative of the virus diversity was determined as described in the materials and methods section. Plain lines separate high and low RHDV binders defined as above or below median values.
Dashed lines separate high and low expressors of rSec1 mRNA defined as animals being above or below the Fut1 and Fut2 range of expression. As described above, rFut1 appears to be the main enzyme controlling for synthesis of A, B and H type 2 in the rabbit duodenum, thus we analyzed the expression of RHDV binding sites at the surface of CHO cells co-transfected with rFut1 and an inactive rSec1 variant.
Interestingly, we observed that in the presence of rSec1, synthesis of RHDV binding sites by transfected rFut1 was significantly decreased.
The effect was amplified when a higher amount of rSec1 than of rFut1 encoding plasmid was used. To confirm that the decreased RHDV binding was due to a decreased synthesis of H type 2 ligands, the same experiment was performed using an anti-H type 2 antibody.
To further confirm these results, we then generated stable transfectants of rFut1. These cells express H type 2 epitopes and upon transfection of rSec1, the percentage of cells expressing the H epitope and attaching RHDV were clearly decreased Fig 7C. The loss of H type 2 RHDV-binding sites was only partial, likely because only a fraction of the cells was actually transfected by rSec1. These results indicate that the presence of an inactive rSec1 protein blocks synthesis of H type 2 ligands by rFut1.
In order to determine how rSec1 blocks the activity of rFut1, we performed confocal microscopy experiments to observe the intracellular localization of these enzymes. This type of glycosyltransferases is known to be expressed in the Golgi apparatus [ 43 ]. We observed that cells transfected with rFut1 containing a DSRed tag in N-terminal position showed a typical Golgi-like perinuclear labeling.
Cells transfected with a GFP-tagged rSec1 were labeled in a less well-defined cellular compartment that also may be compatible with the Golgi. Strikingly, all cells doubly expressing rFut1 and rSec1 showed a cytoplasmic labeling with both the red rFut1 and the green rSec1 tags, clearly indicating that the presence of rSec1 displaced rFut1 out of the Golgi to the cytoplasm, explaining the impairment of its function Fig 8.
Values represent means and S. Forty-eight hours after transfection, binding of either an anti-H antibody Flow cytometry plot of the anti-H antibody staining of stably rFut1-transfected CHO cells negative control in grey. Transfection of rSec1 into stably rFut1-transfected cells. Images of representative cells expressing either protein alone or co-expressing both proteins are shown. Nuclei are stained in blue. A limited and most likely ancient gene conversion between Sec1 and Fut2 in primates and several other mammalian species has been described earlier [ 4 ].
In these species gene conversion was restricted to the gene fragment coding the N-terminal domain of the catalytic region. Since in the same species Sec1 is a pseudogene presenting clear coding-sequence alterations we concluded that gene conversion with either Fut1 or Fut2 would likely no longer be observed because of deleterious effects on the Fut1 and Fut2 enzymes.
The results presented above indicating that rSec1 acts as a dominant-negative of rFut1 are fully consistent with its neofunctionalization in rabbits. Visual inspection of the alignment of the catalytic domain of the three enzymes in the different mammalian species highlighted major differences in the gene conversion extent, particularly between leporids and the remaining mammals Fig 9 and S1 Fig.
In leporids, amino acid motifs shared between the Fut2 and Sec1 encoded enzymes are visible from the beginning of the alignment and extend up to amino acid position In contrast, for the remaining mammals, motifs are only shared between amino acids and Other Fut2-Sec1 shared motifs are observed, but they extend for a few amino-acids only; in pigs, however, these motifs are larger although Sec1 is a pseudogene.
Regarding Fut1-Fut2-Sec1 shared motifs, and although they can be identified for most species, in leporids they extend from amino acid position to Interestingly, in primates, this region is located immediately downstream of the Sec1 premature stop codon in gorillas, humans and chimpanzee S1 Fig.
For each species the 3 enzymes are shown. Dark grey: positions that differ in more than one gene relatively to Xenopus tropicalis. Light grey: positions that are identical in more than one gene between the positions identified by dark grey. Black: premature stop codons. Dots mean identity with the X. The upper part of the alignment comprises amino acid positions to of Homo sapiens FUT1, the lower part comprises amino acids positions to of Homo sapiens FUT1.
Full names of each species and accession numbers of sequences are given in Supplementary Table 1. In order to determine recombination breakpoints, a GARD analysis was performed considering three distinct groups: all mammals, all mammals except leporids and only leporids.
This partition was according to the different extent of gene conversion as observed from the amino acid alignment. One recombination breakpoint was consistently detected when analyzing mammals at nucleotide of the alignment. When analyzing leporids only, the recombination breakpoint was detected at nucleotide Table 3. The phylogenetic trees S2 Fig constructed for the three genes considering the different dataset partitions and the recombination breakpoints identified by GARD Table 3 showed a wider extent of gene conversion between Fut2 and Sec1 in leporids than in the other groups of mammals analyzed.
Although gene conversion events involving Fut1 are apparent from visual inspection of Fig 9 and S1 Fig , which suggested its involvement in the gene conversion in leporids, they are not clear from the phylogenetic trees.
These carbohydrates expressed on epithelial surfaces in contact with the environment, such as the gut, the upper airways and the lower genito-urinary tract, constitute attachment factors for various pathogens.
The earlier observation that a rabbit Sec1 allele was associated with survival to a devastating RHDV outbreak led to the hypothesis that this allele was in linkage disequilibrium with a rFut2 promoter allele responsible for low expression of the corresponding Fut2 enzyme. Thus individual differences in rFut2 transcription rather than in enzyme catalytic properties could have accounted for large differences in the expression of the A, B or H glycan motifs.
Sequencing of the rFut2 promoter region revealed an association between a genetic variant and survival to RHDV in a wild rabbit population.
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