The gustin (CA6) gene polymorphism, rs2274333 (A/G), as a mechanistic link between PROP tasting and fungiform taste papilla density and maintenance

Abstract

Taste sensitivity to PROP varies greatly among individuals and is associated with polymorphisms in the bitter receptor gene TAS2R38, and with differences in fungiform papilla density on the anterior tongue surface. Recently we showed that the PROP non-taster phenotype is strongly associated with the G variant of polymorphism rs2274333 (A/G) of the gene that controls the salivary trophic factor, gustin. The aims of this study were 1) to investigate the role of gustin gene polymorphism rs2274333 (A/G), in PROP sensitivity and fungiform papilla density and morphology, and 2) to investigate the effect of this gustin gene polymorphism on cell proliferation and metabolic activity. Sixty-four subjects were genotyped for both genes by PCR techniques, their PROP sensitivity was assessed by scaling and threshold methods, and their fungiform papilla density, diameter and morphology were determined. In vitro experiments examined cell proliferation and metabolic activity, following treatment with saliva of individuals with and without the gustin gene mutation, and with isolated protein, in the two iso-forms. Gustin and TAS2R38 genotypes were associated with PROP threshold (p=0.0001 and p=0.0042), but bitterness intensity was mostly determined by TAS2R38 genotypes (p<0.000001). Fungiform papillae densities were associated with both genotypes (p<0.014) (with a stronger effect for gustin; p=0.0006), but papilla morphology was a function of gustin alone (p<0.0012). Treatment of isolated cells with saliva from individuals with the AA form of gustin or direct application of the active iso-form of gustin protein increased cell proliferation and metabolic activity (p<0.0135). These novel findings suggest that the rs2274333 polymorphism of the gustin gene affects PROP sensitivity by acting on fungiform papilla development and maintenance, and could provide the first mechanistic explanation for why PROP super-tasters are more responsive to a broad range of oral stimuli.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1. Relationship between PROP phenotype and gustin gene and TAS2R38 polymorphisms.

PROP threshold (A) and bitterness intensity ratings (3.2 mM) (B) of individuals with genotypes AA, AG and GG of gustin (CA6) polymorphism rs2274333 (A/G), and of individuals with genotypes PAV/PAV, PAV/AVI and AVI/AVI of TAS2R38. All values are mean (± SEM). n=63. Different letters indicate significant difference (p≤0.0471; Newman-Keuls test subsequent to main effects ANOVA).

Figure 2. Relationship between density of fungiform papillae and gustin gene and TAS2R38 polymorphisms.

Mean values ± SEM of density of fungiform papillae (No. /cm2) on the anterior part of the tongue of individuals with genotypes AA, AG and GG of gustin (CA6) polymorphism rs2274333 (A/G) (upper graph) and of individuals with genotypes PAV/PAV, PAV/AVI and AVI/AVI of TAS2R38 (lower graph). n=63. Different letters indicate significant difference (p≤0.0379; Newman-Keuls test subsequent to main effects ANOVA). Examples of the 6-mm-diameter stained area of the tongue tip where measures were taken are shown to the right of the graphs.

Figure 3. Relationship between fungiform papillae diameter and gustin gene and TAS2R38 polymorphisms.

Mean values ± SEM of the diameter of fungiform papillae of individuals with genotypes AA, AG and GG of gustin (CA6) polymorphism rs2274333 (A/G) (upper graph) and of individuals with genotypes PAV/PAV, PAV/AVI and AVI/AVI of TAS2R38 (lower graph). n=63. Different letters indicate significant difference (p≤ 0.00053; Newman-Keuls test subsequent to main effects ANOVA).

Figure 4. Relationship between fungiform papillae distortion and gustin gene polymorphism.

Standard deviation (SD) of diameter of fungiform papillae (A) and percentage of distorted fungiform papillae (B) in individuals with genotypes AA, AG and GG of gustin (CA6) polymorphism rs2274333 (A/G). All values are mean (± SEM). n=63. Different letters indicate significant difference (p≤0.00019; Newman-Keuls test subsequent to one-way ANOVA).

Figure 5. Effect of gustin gene polymorphism rs2274333 (A/G) in vitro experiments.

A, Number of cells, expressed as percentage of control, after treatments with saliva of subjects with genotype AA (n=12) or with saliva of subjects with genotype GG (n=12); different letters indicate significant difference (p=0.0135; Student’s t test). B, Fluorescence emission at a wavelength of 590 nm obtained from cells treated for 72 h with saliva of subjects with genotype AA, genotype GG and control; n=12; different letters indicate significant differences (p≤0.00023; Newman-Keuls test subsequent to one-way ANOVA). C, Fluorescence emission at a wavelength of 590 nm obtained from cells treated for 72 h with the two iso-forms of isolated gustin (gustin Ser90 or gustin Gly90) + Zn, control + Zn, or control; n=33; different letters indicate significant differences (p≤0.00067; Newman-Keuls test subsequent to one-way ANOVA).