Structural and Biochemical Analysis of the Hordeum vulgare L. HvGR-RBP1 Protein, a Glycine-Rich RNA-Binding Protein Involved in the Regulation of Barley Plant Development and Stress Resp
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- 作者：Brian P. Tripet ; Katelyn E. Mason ; Brian J. Eilers ; Jennifer Burns ; Paul Powell ; Andreas M. Fischer ; Val茅rie Copi茅
- 刊名：Biochemistry
- 出版年：2014
- 出版时间：December 23, 2014
- 年：2014
- 卷：53
- 期：50
- 页码：7945-7960
- 全文大小：779K
- ISSN：1520-4995
文摘
The timing of whole-plant senescence influences important agricultural traits such as yield and grain protein content. Post-transcriptional regulation by plant RNA-binding proteins is essential for proper control of gene expression, development, and stress responses. Here, we report the three-dimensional solution NMR structure and nucleic acid-binding properties of the barley glycine-rich RNA-binding protein HvGR-RBP1, whose transcript has been identified as being >45-fold up-regulated in early鈥攁s compared to late鈥攕enescing near-isogenic barley germplasm. NMR analysis reveals that HvGR-RBP1 is a multidomain protein comprising a well-folded N-terminal RNA Recognition Motif (RRM) and a structurally disordered C-terminal glycine-rich domain. Chemical shift differences observed in 2D 1H鈥?sup>15N correlation (HSQC) NMR spectra of full-length HvGR-RBP1 and N-HvGR-RBP1 (RRM domain only) suggest that the two domains can interact both in-trans and intramolecularly, similar to what is observed in the tobacco NtGR-RBP1 protein. Further, we show that the RRM domain of HvGR-RBP1 binds single-stranded DNA nucleotide fragments containing the consensus nucleotide sequence 5鈥?TTCTGX-3鈥?with low micromolar affinity in vitro. We also demonstrate that the C-terminal glycine-rich (HvGR) domain of Hv-GR-RBP1 can interact nonspecifically with ssRNA in vitro. Structural similarities with other plant glycine-rich RNA-binding proteins suggest that HvGR-RBP1 may be multifunctional. Based on gene expression analysis following cold stress in barley and E. coli growth studies following cold shock treatment, we conclude that HvGR-RBP1 functions in a manner similar to cold-shock proteins and harbors RNA chaperone activity. HvGR-RBP1 is therefore not only involved in the regulation of barley development including senescence, but also functions in plant responses to environmental stress.