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System for stable ?-estradiol-inducible gene expression in the moss Physcomitrella patens.

ABSTRACT: Inducible transgene expression provides a useful tool to analyze gene function. The moss Physcomitrellapatens is a model basal land plant with well-developed research tools, including a high efficiency of gene targeting and substantial genomics resources. However, current systems for controlled transgene expression remain limited. Here we report the development of an estrogen receptor mediated inducible gene expression system, based on the system used in flowering plants. After identifying the appropriate promoters to drive the chimeric transducer, we succeeded in inducing transcription over 1,000-fold after 24 h incubation with ?-estradiol. The P. patens system was also effective for high-level long-term induction of gene expression; transcript levels of the activated gene were maintained for at least seven days on medium containing ?-estradiol. We also established two potentially neutral targeting sites and a set of vectors for reproducible expression of two transgenes. This ?-estradiol-dependent system will be useful to test genes individually or in combination, allowing stable, inducible transgenic expression in P. patens.

SUBMITTER: Kubo M

PROVIDER: S-EPMC3785464 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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System for stable β-estradiol-inducible gene expression in the moss Physcomitrella patens.

Kubo Minoru M Imai Akihiro A Nishiyama Tomoaki T Ishikawa Masaki M Sato Yoshikatsu Y Kurata Tetsuya T Hiwatashi Yuji Y Reski Ralf R Hasebe Mitsuyasu M

PloS one 20130927 9

Inducible transgene expression provides a useful tool to analyze gene function. The moss Physcomitrellapatens is a model basal land plant with well-developed research tools, including a high efficiency of gene targeting and substantial genomics resources. However, current systems for controlled transgene expression remain limited. Here we report the development of an estrogen receptor mediated inducible gene expression system, based on the system used in flowering plants. After identifying the a ...[more]

PMID: 24086772

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Project description:BACKGROUND: Hexokinase catalyzes the phosphorylation of glucose and fructose, but it is also involved in sugar sensing in both fungi and plants. We have previously described two types of hexokinases in the moss Physcomitrella. Type A, exemplified by PpHxk1, the major hexokinase in Physcomitrella, is a soluble protein that localizes to the chloroplast stroma. Type B, exemplified by PpHxk2, has an N-terminal membrane anchor. Both types are found also in vascular plants, and localize to the chloroplast stroma and mitochondrial membranes, respectively. RESULTS: We have now characterized all 11 hexokinase encoding genes in Physcomitrella. Based on their N-terminal sequences and intracellular localizations, three of the encoded proteins are type A hexokinases and four are type B hexokinases. One of the type B hexokinases has a splice variant without a membrane anchor, that localizes to the cytosol and the nucleus. However, we also found two new types of hexokinases with no obvious orthologs in vascular plants. Type C, encoded by a single gene, has neither transit peptide nor membrane anchor, and is found in the cytosol and in the nucleus. Type D hexokinases, encoded by three genes, have membrane anchors and localize to mitochondrial membranes, but their sequences differ from those of the type B hexokinases. Interestingly, all moss hexokinases are more similar to each other in overall sequence than to hexokinases from other plants, even though characteristic sequence motifs such as the membrane anchor of the type B hexokinases are highly conserved between moss and vascular plants, indicating a common origin for hexokinases of the same type. CONCLUSIONS: We conclude that the hexokinase gene family is more diverse in Physcomitrella, encoding two additional types of hexokinases that are absent in vascular plants. In particular, the presence of a cytosolic and nuclear hexokinase (type C) sets Physcomitrella apart from vascular plants, and instead resembles yeast, where all hexokinases localize to the cytosol. The fact that all moss hexokinases are more similar to each other than to hexokinases from vascular plants, even though both type A and type B hexokinases are present in all plants, further suggests that the hexokinase gene family in Physcomitrella has undergone concerted evolution.

Dataset Information

System for stable ?-estradiol-inducible gene expression in the moss Physcomitrella patens.

Publications

System for stable β-estradiol-inducible gene expression in the moss Physcomitrella patens.

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