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Identification and functional analysis of pistil self-incompatibility factor HT-B of Petunia.

Puerta AR, Ushijima K, Koba T, Sassa H - J. Exp. Bot. (2009)

Bottom Line: The structural features and expression pattern of Petunia HT-B were very similar to those of Nicotiana and Solanum.Unlike other solanaceous species, expression of HT-B was also observed in self-compatible Petunia species.Quantitative analysis of the HT-B mRNA accumulation in the transgenics showed that a 100-fold reduction is not sufficient and a >1000-fold reduction is required to achieve partial breakdown of GSI.

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Science and Technology, Chiba University, 648 Matsudo, Matsudo, Chiba 271-8510, Japan.

ABSTRACT
Gametophytic self-incompatibility (GSI) in Solanaceae, Rosaceae, and Plantaginaceae is controlled by a multiallelic S-locus. The specificities of pistil and pollen are controlled by separate S-locus genes, S-RNase and SLF/SFB, respectively. Although the S-specificity is determined by the S-locus genes, factors located outside the S-locus are also required for expression of GSI. HT-B is one of the pistil non-S-factors identified in Nicotiana and Solanum, and encodes a small asparagine/aspartate-rich extracellular protein with unknown biochemical function. Here, HT-B was cloned from Petunia and characterized. The structural features and expression pattern of Petunia HT-B were very similar to those of Nicotiana and Solanum. Unlike other solanaceous species, expression of HT-B was also observed in self-compatible Petunia species. RNA interference (RNAi)-mediated suppression of Petunia HT-B resulted in partial breakdown of GSI. Quantitative analysis of the HT-B mRNA accumulation in the transgenics showed that a 100-fold reduction is not sufficient and a >1000-fold reduction is required to achieve partial breakdown of GSI.

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Amino acid sequence alignment and phylogenetic tree of HT-like proteins. (A) The amino acid sequences of HT-like proteins were aligned by CLUSTAL W (Thompson et al., 1994). Putative signal peptides are shown in lower case. A chacteristic deletion for HT-A is denoted with #. The N/D-rich domain was boxed. (B) A Neighbor–Joining tree (Saitou and Nei, 1987) of HT-like proteins. NOD24 protein of soybean (GmNOD24; M10595) was chosen as an outgroup for rooting, because it is a member of the recently proposed expanded HT family, the HT/NOD-24 family, and is distantly related to solanaceous HT-like proteins (Kondo and McClure, 2008). Numbers denote bootstrap values with 1000 replicates.
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fig1: Amino acid sequence alignment and phylogenetic tree of HT-like proteins. (A) The amino acid sequences of HT-like proteins were aligned by CLUSTAL W (Thompson et al., 1994). Putative signal peptides are shown in lower case. A chacteristic deletion for HT-A is denoted with #. The N/D-rich domain was boxed. (B) A Neighbor–Joining tree (Saitou and Nei, 1987) of HT-like proteins. NOD24 protein of soybean (GmNOD24; M10595) was chosen as an outgroup for rooting, because it is a member of the recently proposed expanded HT family, the HT/NOD-24 family, and is distantly related to solanaceous HT-like proteins (Kondo and McClure, 2008). Numbers denote bootstrap values with 1000 replicates.

Mentions: PiHT-B was isolated from pistils of P. inflata by RT-PCR using degenerate primers designed according to HT sequences of Nicotiana and Solanum. Figure 1A shows the deduced amino acid sequence of PiHT-B and other related proteins. PiHT-B encodes a 119 residue amino acid protein with a predicted cleavage site before Arg24 (SignalP, Nielsen et al., 1997). The pI of the mature protein is calculated to be 4.42. PiHT-B, unlike the previously described PiHTL (Sassa and Hirano, 2006) and like other HT-B proteins, contains a C-terminal N/D-rich domain (23 residues) with three cysteine residues at each side. HT-A proteins have a characteristic deletion of 5–7 residues near the N-terminal region of mature proteins (Kondo et al., 2002b; O'Brien et al., 2002). The corresponding region of PiHT-B was much closer to that of solanaceous HT-Bs and Petunia HTL than to HT-As (Fig. 1A). Phylogenetic analysis showed that the HT-like proteins were first classified into two groups, the HT-A/B group and the HT-L/M group (Fig. 1B). PiHT-B was categorized in the HT-A/B group, and was closely related to HT-B of Nicotiana. Identities between PiHT-B and the other solanaceous proteins ranged between 34.9% and 58.0%.


Identification and functional analysis of pistil self-incompatibility factor HT-B of Petunia.

Puerta AR, Ushijima K, Koba T, Sassa H - J. Exp. Bot. (2009)

Amino acid sequence alignment and phylogenetic tree of HT-like proteins. (A) The amino acid sequences of HT-like proteins were aligned by CLUSTAL W (Thompson et al., 1994). Putative signal peptides are shown in lower case. A chacteristic deletion for HT-A is denoted with #. The N/D-rich domain was boxed. (B) A Neighbor–Joining tree (Saitou and Nei, 1987) of HT-like proteins. NOD24 protein of soybean (GmNOD24; M10595) was chosen as an outgroup for rooting, because it is a member of the recently proposed expanded HT family, the HT/NOD-24 family, and is distantly related to solanaceous HT-like proteins (Kondo and McClure, 2008). Numbers denote bootstrap values with 1000 replicates.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2657546&req=5

fig1: Amino acid sequence alignment and phylogenetic tree of HT-like proteins. (A) The amino acid sequences of HT-like proteins were aligned by CLUSTAL W (Thompson et al., 1994). Putative signal peptides are shown in lower case. A chacteristic deletion for HT-A is denoted with #. The N/D-rich domain was boxed. (B) A Neighbor–Joining tree (Saitou and Nei, 1987) of HT-like proteins. NOD24 protein of soybean (GmNOD24; M10595) was chosen as an outgroup for rooting, because it is a member of the recently proposed expanded HT family, the HT/NOD-24 family, and is distantly related to solanaceous HT-like proteins (Kondo and McClure, 2008). Numbers denote bootstrap values with 1000 replicates.
Mentions: PiHT-B was isolated from pistils of P. inflata by RT-PCR using degenerate primers designed according to HT sequences of Nicotiana and Solanum. Figure 1A shows the deduced amino acid sequence of PiHT-B and other related proteins. PiHT-B encodes a 119 residue amino acid protein with a predicted cleavage site before Arg24 (SignalP, Nielsen et al., 1997). The pI of the mature protein is calculated to be 4.42. PiHT-B, unlike the previously described PiHTL (Sassa and Hirano, 2006) and like other HT-B proteins, contains a C-terminal N/D-rich domain (23 residues) with three cysteine residues at each side. HT-A proteins have a characteristic deletion of 5–7 residues near the N-terminal region of mature proteins (Kondo et al., 2002b; O'Brien et al., 2002). The corresponding region of PiHT-B was much closer to that of solanaceous HT-Bs and Petunia HTL than to HT-As (Fig. 1A). Phylogenetic analysis showed that the HT-like proteins were first classified into two groups, the HT-A/B group and the HT-L/M group (Fig. 1B). PiHT-B was categorized in the HT-A/B group, and was closely related to HT-B of Nicotiana. Identities between PiHT-B and the other solanaceous proteins ranged between 34.9% and 58.0%.

Bottom Line: The structural features and expression pattern of Petunia HT-B were very similar to those of Nicotiana and Solanum.Unlike other solanaceous species, expression of HT-B was also observed in self-compatible Petunia species.Quantitative analysis of the HT-B mRNA accumulation in the transgenics showed that a 100-fold reduction is not sufficient and a >1000-fold reduction is required to achieve partial breakdown of GSI.

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Science and Technology, Chiba University, 648 Matsudo, Matsudo, Chiba 271-8510, Japan.

ABSTRACT
Gametophytic self-incompatibility (GSI) in Solanaceae, Rosaceae, and Plantaginaceae is controlled by a multiallelic S-locus. The specificities of pistil and pollen are controlled by separate S-locus genes, S-RNase and SLF/SFB, respectively. Although the S-specificity is determined by the S-locus genes, factors located outside the S-locus are also required for expression of GSI. HT-B is one of the pistil non-S-factors identified in Nicotiana and Solanum, and encodes a small asparagine/aspartate-rich extracellular protein with unknown biochemical function. Here, HT-B was cloned from Petunia and characterized. The structural features and expression pattern of Petunia HT-B were very similar to those of Nicotiana and Solanum. Unlike other solanaceous species, expression of HT-B was also observed in self-compatible Petunia species. RNA interference (RNAi)-mediated suppression of Petunia HT-B resulted in partial breakdown of GSI. Quantitative analysis of the HT-B mRNA accumulation in the transgenics showed that a 100-fold reduction is not sufficient and a >1000-fold reduction is required to achieve partial breakdown of GSI.

Show MeSH
Related in: MedlinePlus