Os01g0742500 - Revision history

192.168.72.52 at 04:55, 14 May 2015

2015-05-14T04:55:56Z

Xysj2012: /* Structured Information */

2015-05-13T07:28:32Z

‎Structured Information

Zhanglide: /* Mutation */

2014-05-29T04:48:33Z

‎Mutation

Zhanglide: /* Mutation */

2014-05-29T04:45:19Z

‎Mutation

Zhanglide: /* Mutation */

2014-05-29T04:44:55Z

‎Mutation

Zhanglide: /* Mutation */

2014-05-29T04:44:20Z

‎Mutation

Zhanglide: /* Mutation */

2014-05-27T14:18:16Z

‎Mutation

Zhanglide: /* Mutation */

2014-05-27T14:17:52Z

‎Mutation

Zhanglide: /* Mutation */

2014-05-27T13:55:14Z

‎Mutation

Zhanglide: /* Function */

2014-05-27T13:46:37Z

‎Function

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 ''[[Figure 1. Transformation of catalytically inactive mutants for OsHXK5 and OsHXK6 into a yeast hexokinase mutant. A, Schematic representation of OsHXK5 and OsHXK6 and their catalytically inactive mutation sites. Mitochondrial targeting signals and NLSs are indicated as white (M) and black (N) rectangles. 1, 2, and A indicate the conserved phosphate 1, 2, and adenosine interaction regions within the ATP-binding site, respectively. The region S indicates the conserved sugar-binding domain. B, Complementation of the hexokinase-deficient yeast triple mutant YSH7.4-3C (hxk1, hxk2, glk1) with OsHXK5, OsHXK6, and their catalytically inactive mutant alleles. The transformed colonies were streaked on the SD-Ura medium (synthetic defined minimal medium lacking uracil) containing 2% D-Glc as a sole carbon source and grown for 3 d at 30�C (top). The YSH7.4-3C mutant strain transformed with the pDR196 vector was used as a control. As control experiment, YSH7.4-3C mutant strains transformed with pDR196, OsHXK5, OsHXK6, and their catalytically inactive mutant alleles were streaked on the SD-Ura medium containing 2% D-Gal (middle). Expression levels of HXK5, HXK6, and their mutant alleles in these strains were measured by RT-PCR analysis (bottom).]]''
-==Labs working on this gene==
+==Structured Information==
-# Plant Metabolism Research Center and Graduate School of Biotechnology
+     [[Category:Genes]][[Category:Oryza Sativa Japonica Group]][[Category:Japonica Chromosome 1]]
 ==References==

@@ Line 46: / Line 46: @@
 ==Structured Information==
 [[Category:Genes]]
 [[Category:Japonica mRNA]]

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 The expression of OsHXK5, OsHXK6, or their mutant alleles complemented the glucose insensitive2-1 mutant, thereby resulting in wild-type characteristics in seedling development, Glc-dependent gene expression, and plant growth.
 It has been reported in Arabidopsis that the sugar sensing and signaling functions of AtHXK1 do not depend on its Glc phosphorylation activity. To uncouple the sugar sensing and signaling activities from Glc phosphorylation, we employed a targeted mutagenesis experiment to generate catalytically inactive mutants of the candidate rice Glc sensors OsHXK5 and OsHXK6. In the mutant alleles, ATP binding was eliminated by mutating the conserved Gly (G) in the phosphate 1 domain of the ATP-binding site to Asp (D) and phosphoryl transfer was prevented by mutating the conserved Ser (S) in the sugar-binding domain to Ala. These mutant alleles were referred to as OsHXK5-G113D, OsHXK5-S186A, OsHXK6-G112D, and OsHXK6-S185A, according to their mutation sites. To determine whether enzyme catalytic activity was abolished in the mutant alleles, the individual cDNA clones were tested to complement the yeast triple mutant YSH7.4-3C (hxk1, hxk2, glk1), which lacks endogenous hexokinase activity. While yeast cells transformed with wild-type cDNAs of OsHXK5 and OsHXK6 were able to grow on selection medium containing Glc as the sole carbon source , yeast cells transformed with the OsHXK mutant alleles or the empty pDR196 vector did not grow on the selection medium. In the control experiment, all transformed yeast cells grew on the Gal-containing medium. In addition, expressions of HXK5, HXK6, and their catalytically inactive mutant alleles were confirmed by reverse transcription (RT)-PCR analysis. These findings demonstrate that the mutant OsHXKs lacked catalytic activity.
 [[File:njmzld.jpg]]
 ==Labs working on this gene==

@@ Line 25: / Line 25: @@
 The expression of OsHXK5, OsHXK6, or their mutant alleles complemented the glucose insensitive2-1 mutant, thereby resulting in wild-type characteristics in seedling development, Glc-dependent gene expression, and plant growth.
 It has been reported in Arabidopsis that the sugar sensing and signaling functions of AtHXK1 do not depend on its Glc phosphorylation activity. To uncouple the sugar sensing and signaling activities from Glc phosphorylation, we employed a targeted mutagenesis experiment to generate catalytically inactive mutants of the candidate rice Glc sensors OsHXK5 and OsHXK6. In the mutant alleles, ATP binding was eliminated by mutating the conserved Gly (G) in the phosphate 1 domain of the ATP-binding site to Asp (D) and phosphoryl transfer was prevented by mutating the conserved Ser (S) in the sugar-binding domain to Ala. These mutant alleles were referred to as OsHXK5-G113D, OsHXK5-S186A, OsHXK6-G112D, and OsHXK6-S185A, according to their mutation sites. To determine whether enzyme catalytic activity was abolished in the mutant alleles, the individual cDNA clones were tested to complement the yeast triple mutant YSH7.4-3C (hxk1, hxk2, glk1), which lacks endogenous hexokinase activity. While yeast cells transformed with wild-type cDNAs of OsHXK5 and OsHXK6 were able to grow on selection medium containing Glc as the sole carbon source , yeast cells transformed with the OsHXK mutant alleles or the empty pDR196 vector did not grow on the selection medium. In the control experiment, all transformed yeast cells grew on the Gal-containing medium. In addition, expressions of HXK5, HXK6, and their catalytically inactive mutant alleles were confirmed by reverse transcription (RT)-PCR analysis. These findings demonstrate that the mutant OsHXKs lacked catalytic activity.
-[[File:zldnjm.jpg]]
+[[File:njmzld.jpg]]
 ==Labs working on this gene==

@@ Line 25: / Line 25: @@
 The expression of OsHXK5, OsHXK6, or their mutant alleles complemented the glucose insensitive2-1 mutant, thereby resulting in wild-type characteristics in seedling development, Glc-dependent gene expression, and plant growth.
 It has been reported in Arabidopsis that the sugar sensing and signaling functions of AtHXK1 do not depend on its Glc phosphorylation activity. To uncouple the sugar sensing and signaling activities from Glc phosphorylation, we employed a targeted mutagenesis experiment to generate catalytically inactive mutants of the candidate rice Glc sensors OsHXK5 and OsHXK6. In the mutant alleles, ATP binding was eliminated by mutating the conserved Gly (G) in the phosphate 1 domain of the ATP-binding site to Asp (D) and phosphoryl transfer was prevented by mutating the conserved Ser (S) in the sugar-binding domain to Ala. These mutant alleles were referred to as OsHXK5-G113D, OsHXK5-S186A, OsHXK6-G112D, and OsHXK6-S185A, according to their mutation sites. To determine whether enzyme catalytic activity was abolished in the mutant alleles, the individual cDNA clones were tested to complement the yeast triple mutant YSH7.4-3C (hxk1, hxk2, glk1), which lacks endogenous hexokinase activity. While yeast cells transformed with wild-type cDNAs of OsHXK5 and OsHXK6 were able to grow on selection medium containing Glc as the sole carbon source , yeast cells transformed with the OsHXK mutant alleles or the empty pDR196 vector did not grow on the selection medium. In the control experiment, all transformed yeast cells grew on the Gal-containing medium. In addition, expressions of HXK5, HXK6, and their catalytically inactive mutant alleles were confirmed by reverse transcription (RT)-PCR analysis. These findings demonstrate that the mutant OsHXKs lacked catalytic activity.
-[[File:zldnjm.jpg]][[File:njmzld.jpg]]
+[[File:zldnjm.jpg]]
 ==Labs working on this gene==

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 These OsHXKs including OsHXK6 retain a dual-targeting ability to mitochondria and nuclei. Some results also provide evidence that rice OsHXK5 and OsHXK6 can function as Glc sensors.
 The OsHXK5DmTP-GFP and OsHXK6DmTP-GFP fusion proteins, which lack N-terminal mitochondrial targeting peptides, were present mainly in the nucleus with a small amount of the proteins seen in the cytosol. In addition, the OsHXK5NLS-GFP and OsHXK6NLS-GFP fusion proteins harboring nuclear localization signals were targeted predominantly in the nucleus, suggesting that these OsHXKs retain a dual-targeting ability to mitochondria and nuclei.
 ===Expression===