ABSTRACT
Bitter taste perception is mediated by a family of G protein-coupled receptors (T2Rs) in vertebrates. Common carp (Cyprinus carpio), which has experienced an additional round of whole genome duplication during the course of evolution, has a small number of T2R genes similar to zebrafish, a closely related cyprinid fish species, and their expression pattern at the cellular level or their cognate ligands have not been elucidated yet. Here, we showed through in situ hybridization experiments, that three common carp T2R (ccT2R) genes encoding ccT2R200-1, ccT2R202-1, and ccT2R202-2, were specifically expressed in the subsets of taste receptor cells in the lips and gill rakers. ccT2R200-1 was co-expressed with genes encoding downstream signal transduction molecules, such as PLC-ß2 and Gαia. Heterologous expression system revealed that each ccT2R showed narrowly, intermediately, or broadly tuned ligand specificity, as in the case of zebrafish T2Rs. However, ccT2Rs showed different ligand profiles from their orthologous zebrafish T2Rs previously reported. Finally, we identified three ccT2Rs, namely ccT2R200-1, ccT2R200-2, and ccT2R203-1, to be activated by natural bitter compounds, andrographolide and/or picrotoxinin, which elicited no response to zebrafish T2Rs, in a dose-dependent manner. These results suggest that some ccT2Rs may have evolved to function in the oral cavity as taste receptors for natural bitter compounds found in the habitats in a species-specific manner.
ABSTRACT
Two isolates of Tomato aspermy virus (TAV), V-TAV and C-TAV, can systemically infect Nicotiana benthamiana but only C-TAV can move systemically in N. tabacum. Any pseudorecombinants between the two strains could not move systemically in tobacco as efficiently as C-TAV. However, a pseudorecombinant consisting of RNAs 1 and 3 of V-TAV and RNA 2 of C-TAV (V1C2V3), which cannot infect tobacco systemically, generated progeny with a mutation in V1 and a recombination in C2 (V1(m)C2(r)V3), enabling the virus to move systemically. To avoid further mutation and recombination in the virus, we used Cucumber mosaic virus RNA3 (Y3) for subsequent experiments. Northern blot analyses showed that RNA4A, which encodes the 2b protein (2b), and RNA5 abundantly accumulated in V1(m)C2(r)Y3-infected tobacco. V1(m)C2(r)Y3 actually caused higher accumulation of 2b than did V1C2Y3 in Western blots, and overexpression of 2b by the PVX vector enabled V1C2Y3 to move systemically in tobacco, suggesting that 2b accumulation promotes viral systemic movement. Because RNA-silencing suppressor (RSS) activity of 2b was thought to be involved in systemic movement, we compared the RSS activity of 2b for the two TAV isolates; C-TAV 2b had stronger activity than did V-TAV 2b in tobacco in a transient protoplast assay. Our data also demonstrated that 2b and RNA5 play an important role in the evolution of members of genus Cucumovirus by generating mutant/recombinant viruses and viral systemic movement over RNA silencing.
