Figure 3.
Figure 3. In place of the For2A PTEN domain, PTEN homologues fused to the FH1-FH2 domains of For2A differentially complement formin-mediated polarized growth. (A) Representative chlorophyll autofluorescence images of control RNAi (GUS-RNAi), For2 RNAi (For2AB-5′UTR), and For2 RNAi plants co-transformed with indicated constructs. Bar, 100 µm. (B) Quantification of area and circularity by chlorophyll autofluorescence shows that replacing the For2A PTEN domain with moss PTEN homologues provides full complementation with PTENA, but not with PTEND. Replacement using HsPTEN does not complement the For2 RNAi phenotype. Tagging the formin with an epitope tag (3XFLAG) does not affect its ability to complement the phenotype. Number of plants analyzed is: 101, GUS-RNAi; 101, For2AB-5′UTR; 50, +For2A; 51, +For2A-3XFLAG; 76, +PTENA-FH1FH2; 50, +PTEND-FH1FH2-3XFLAG; 50, +HsPTEN-FH1FH2-3XFLAG. Error bars represent SEM and letters above the bars indicate statistical groups with α = 0.05 from an ANOVA analysis. (C) Alignment of the phosphoinositide-binding regions of human PTEN (HsPTEN) with the For2A and For2B PTEN domains and four P. patens PTEN homologues (PpPTENA-D). The arrow indicates an arginine residue critical for catalytic activity. Note this arginine is absent in the formin PTEN domains.

In place of the For2A PTEN domain, PTEN homologues fused to the FH1-FH2 domains of For2A differentially complement formin-mediated polarized growth. (A) Representative chlorophyll autofluorescence images of control RNAi (GUS-RNAi), For2 RNAi (For2AB-5′UTR), and For2 RNAi plants co-transformed with indicated constructs. Bar, 100 µm. (B) Quantification of area and circularity by chlorophyll autofluorescence shows that replacing the For2A PTEN domain with moss PTEN homologues provides full complementation with PTENA, but not with PTEND. Replacement using HsPTEN does not complement the For2 RNAi phenotype. Tagging the formin with an epitope tag (3XFLAG) does not affect its ability to complement the phenotype. Number of plants analyzed is: 101, GUS-RNAi; 101, For2AB-5′UTR; 50, +For2A; 51, +For2A-3XFLAG; 76, +PTENA-FH1FH2; 50, +PTEND-FH1FH2-3XFLAG; 50, +HsPTEN-FH1FH2-3XFLAG. Error bars represent SEM and letters above the bars indicate statistical groups with α = 0.05 from an ANOVA analysis. (C) Alignment of the phosphoinositide-binding regions of human PTEN (HsPTEN) with the For2A and For2B PTEN domains and four P. patens PTEN homologues (PpPTENA-D). The arrow indicates an arginine residue critical for catalytic activity. Note this arginine is absent in the formin PTEN domains.

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