Abstract
Main conclusion
Silencing of CI-sHsps by RNAi negatively affected the seed germination process and heat stress response of rice seedlings. Seed size of RNAiCI-sHsp was reduced as compared to wild-type plants.
Abstract
Small heat shock proteins (sHsps) are the ATP-independent chaperones ubiquitously expressed in response to diverse environmental and developmental cues. Cytosolic sHsps constitute the major repertoire of sHsp family. Rice cytosolic class I (CI)-sHsps consists of seven members (Hsp16.9A, Hsp16.9B, Hsp16.9C, Hsp17.4, Hsp17.7, Hsp17.9A and Hsp18). Purified OsHsp17.4 and OsHsp17.9A proteins exhibited chaperone activity by preventing formation of large aggregates with model substrate citrate synthase. OsHsp16.9A and OsHsp17.4 showed nucleo-cytoplasmic localization, while the localization of OsHsp17.9A was preferentially in the nucleus. Transgenic tobacco plants expressing OsHsp17.4 and OsHsp17.9A proteins and Arabidopsis plants ectopically expressing OsHsp17.4 protein showed improved thermotolerance to the respective trans-hosts during the post-stress recovery process. Single hairpin construct was designed to generate all CI-sHsp silenced (RNAiCI-sHsp) rice lines. The major vegetative and reproductive attributes of the RNAiCI-sHsp plants were comparable to the wild-type rice plants. Basal and acquired thermotolerance response of RNAiCI-sHsp seedlings of rice was mildly affected. The seed length of RNAiCI-sHsp rice plants was significantly reduced. The seed germination process was delayed and seed thermotolerance of RNAiCI-sHsp was negatively affected than the non-transgenic seeds. We, thus, implicate that sHsp genes are critical in seedling thermotolerance and seed physiology.
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Abbreviations
- CI/CII-sHsps:
-
Class I/class II small heat shock proteins
- ECL:
-
Enhanced chemiluminiscence
- HS:
-
Heat stress
References
Agarwal M (2002) Molecular characterization of high and low molecular weight heat shock proteins from rice (Oryza sativa L.). PhD thesis, Univ Delhi
Almoguera C, Prieto-Dapena P, Jordano J (1998) Dual regulation of a heat shock promoter during embryogenesis: stage-dependent role of heat shock elements. Plant J 13:437–446. https://doi.org/10.1046/j.1365-313X.1998.00044.x
Bart R, Chern M, Park C-J et al (2006) A novel system for gene silencing using siRNAs in rice leaf and stem-derived protoplasts. Plant Methods 2:13. https://doi.org/10.1186/1746-4811-2-13
Basha E, Jones C, Wysocki V, Vierling E (2010) Mechanistic differences between two conserved classes of small heat shock proteins found in the plant cytosol. J Biol Chem 285:11489–11497. https://doi.org/10.1074/jbc.M109.074088
Basha E, O’Neill H, Vierling E (2012) Small heat shock proteins and α-crystallins: dynamic proteins with flexible functions. Trends Biochem Sci 37:106–117. https://doi.org/10.1016/j.tibs.2011.11.005
Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254. https://doi.org/10.1016/0003-2697(76)90527-3
Cashikar AG, Duennwald M, Lindquist SL (2005) A chaperone pathway in protein disaggregation: HSP26 alters the nature of protein aggregates to facilitate reactivation by HSP104. J Biol Chem 280:23869–23875. https://doi.org/10.1074/jbc.M502854200
Cheabu S, Ngam PM, Arikit S et al (2018) Effects of heat stress at vegetative and reproductive stages on spikelet fertility. Rice Sci 25:218–226. https://doi.org/10.1016/j.rsci.2018.06.005
Chen X, Lin S, Liu Q et al (2014) Expression and interaction of small heat shock proteins (sHsps) in rice in response to heat stress. Biochim Biophys Acta 1844:818–828. https://doi.org/10.1016/j.bbapap.2014.02.010
Chomczynski P, Sacchi N (2006) The single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction: twenty-something years on. Nat Protoc 1:581–585. https://doi.org/10.1038/nprot.2006.83
Clough SJ, Bent AF (1998) Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J 16:735–743. https://doi.org/10.1046/j.1365-313X.1998.00343.x
Cui Y, Xu G, Wang M et al (2013) Expression of OsMSR3 in Arabidopsis enhances tolerance to cadmium stress. Plant Cell Tissue Organ Cult 113:331–340. https://doi.org/10.1007/s11240-012-0275-x
Dafny-Yelin M, Tzfira T, Vainstein A, Adam Z (2008) Non-redundant functions of sHSP-CIs in acquired thermotolerance and their role in early seed development in Arabidopsis. Plant Mol Biol 67:363–373. https://doi.org/10.1007/s11103-008-9326-4
Frank G, Pressman E, Ophir R et al (2009) Transcriptional profiling of maturing tomato (Solanum lycopersicum L.) microspores reveals the involvement of heat shock proteins, ROS scavengers, hormones, and sugars in the heat stress response. J Exp Bot 60:3891–3908. https://doi.org/10.1093/jxb/erp234
Gietz RD, Schiestl RH (2008) High-efficiency yeast transformation using the LiAc/SS carrier DNA/PEG method. Nat Protoc 2:31–35. https://doi.org/10.1038/nprot.2007.13
González-Schain N, Dreni L, Lawas LMF et al (2016) Genome-wide transcriptome analysis during anthesis reveals new insights into the molecular basis of heat stress responses in tolerant and sensitive rice varieties. Plant Cell Physiol 57:57–68. https://doi.org/10.1093/pcp/pcv174
Guan JC, Jinn TL, Yeh CH et al (2004) Characterization of the genomic structures and selective expression profiles of nine class I small heat shock protein genes clustered on two chromosomes in rice (Oryza sativa L.). Plant Mol Biol 56:795–809. https://doi.org/10.1007/s11103-004-5182-z
Guo H-X, Zeng W-Z, Wang C-Y et al (2016) Expression analysis of two reverse duplicated small heat shock protein genes in rice (Oryza sativa L.). J Integr Agric 15:1685–1692. https://doi.org/10.1016/S2095-3119(15)61224-2
Ham D-J, Moon J-C, Hwang S-G, Jang CS (2013) Molecular characterization of two small heat shock protein genes in rice: their expression patterns, localizations, networks, and heterogeneous overexpressions. Mol Biol Rep 40:6709–6720. https://doi.org/10.1007/s11033-013-2786-x
Haslbeck M, Vierling E (2015) A first line of stress defense: small heat shock proteins and their function in protein homeostasis. J Mol Biol 427:1537–1548. https://doi.org/10.1016/j.jmb.2015.02.002
Huang Y-C, Niu C-Y, Yang C-R, Jinn T-L (2016) The heat-stress factor HSFA6b connects ABA signaling and ABA-mediated heat responses. Plant Physiol 172:1182–1199. https://doi.org/10.1104/pp.16.00860
Jagadish SVK, Muthurajan R, Oane R et al (2010) Physiological and proteomic approaches to address heat tolerance during anthesis in rice (Oryza sativa L.). J Exp Bot 61:143–156. https://doi.org/10.1093/jxb/erp289
Ju Y, Tian H, Zhang R et al (2017) Overexpression of OsHSP18.0-CI enhances resistance to bacterial leaf streak in rice. Rice 10:1–11. https://doi.org/10.1186/s12284-017-0153-6
Jung YJ, Nou IS, Kang KK (2014) Overexpression of Oshsp16.9 gene encoding small heat shock protein enhances tolerance to abiotic stresses in rice. Plant Breed Biotech 2:370–379
Kim M, Kim H, Lee W et al (2015) Quantitative shotgun proteomics analysis of rice anther proteins after exposure to high temperature. Int J Genom 2015:1–9
Koo HJ, Park SM, Kim KP et al (2015) Small heat shock proteins can release light dependence of tobacco seed during germination. Plant Physiol 167:1030–1038. https://doi.org/10.1104/pp.114.252841
Kotak S, Vierling E, Baumlein H, von Koskull-Doring P (2007) A novel transcriptional cascade regulating expression of heat stress proteins during seed development of Arabidopsis. Plant Cell 19:182–195. https://doi.org/10.1105/tpc.106.048165
Lee U, Wie C, Escobar M et al (2005) Genetic analysis reveals domain interactions of Arabidopsis Hsp100/ClpB and cooperation with the small heat shock protein chaperone system. Plant Cell 17:559–571. https://doi.org/10.1105/tpc.104.027540.1
Lee DG, Ahsan N, Lee SH et al (2007) A proteomic approach in analyzing heat-responsive proteins in rice leaves. Proteomics 7:3369–3383. https://doi.org/10.1002/pmic.200700266
Li G, Jain R, Chern M et al (2017) The sequences of 1,504 mutants in the model rice variety Kitaake facilitate rapid functional genomic studies. Plant Cell 29:1218–1231. https://doi.org/10.1105/tpc.17.00154
Lin SK, Chang MC, Tsai YG, Lur HS (2005) Proteomic analysis of the expression of proteins related to rice quality during caryopsis development and the effect of high temperature on expression. Proteomics 5:2140–2156. https://doi.org/10.1002/pmic.200401105
Löw D, Brändle K, Nover L, Forreiter C (2000) Cytosolic heat-stress proteins Hsp17.7 class I and Hsp17.3 class II of tomato act as molecular chaperones in vivo. Planta 211:575–582. https://doi.org/10.1007/s004250000315
Ma W, Guan X, Li J et al (2019) Mitochondrial small heat shock protein mediates seed germination via thermal sensing. Proc Natl Acad Sci USA 116:4716–4721. https://doi.org/10.1073/pnas.1815790116
Malik MK, Slovin JP, Hwang CH, Zimmerman JL (1999) Modified expression of a carrot small heat shock protein gene, Hsp17.7, results in increased or decreased thermotolerance. Plant J 20:89–99. https://doi.org/10.1046/j.1365-313X.1999.00581.x
McLoughlin F, Basha E, Fowler ME et al (2016) Class I and II small heat-shock proteins protect protein translation factors during heat stress. Plant Physiol 172:1221–1236. https://doi.org/10.1104/pp.16.00536
Murakami T, Matsuba S, Funatsuki H et al (2004) Over expression of small heat shock, sHSP17.7 confers both heat tolerance and UV-B resistance to rice plants. Mol Breed 13:165–175
Murashige T, Skoog F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 15:473–497. https://doi.org/10.1111/j.1399-3054.1962.tb08052.x
Neta-Sharir I, Isaacson T, Lurie S, Weiss D (2005) Dual role for tomato heat shock protein 21: protecting photosystem II from oxidative stress and promoting color changes during fruit maturation. Plant Cell 17:1829–1838. https://doi.org/10.1105/tpc.105.031914
O’Farrell PH (1975) High resolution two-dimensional electrophoresis of proteins. J Biol Chem 250:4007–4021
Prieto-Dapena P, Castano R, Almoguera C, Jordano J (2006) Improved resistance to controlled deterioration in transgenic seeds. Plant Physiol 142:1102–1112. https://doi.org/10.1104/pp.106.087817
Sarkar NK, Kim YK, Grover A (2009) Rice sHsp genes: genomic organization and expression profiling under stress and development. BMC Genom 10:393. https://doi.org/10.1186/1471-2164-10-393
Sarkar NK, Kim YK, Grover A (2014) Coexpression network analysis associated with call of rice seedlings for encountering heat stress. Plant Mol Biol 84:125–143. https://doi.org/10.1007/s11103-013-0123-3
Sato Y, Yokoya S (2008) Enhanced tolerance to drought stress in transgenic rice plants overexpressing a small heat-shock protein, sHSP17.7. Plant Cell Rep 27:329–334. https://doi.org/10.1007/s00299-007-0470-0
Sun W, Bernard C, van de Cotte B et al (2001) At-HSP17.6A, encoding a small heat-shock protein in Arabidopsis, can enhance osmotolerance upon overexpression. Plant J 27:407–415. https://doi.org/10.1046/j.1365-313X.2001.01107.x
Suzuki K, Itai R, Suzuki K et al (1998) Formate dehydrogenase, an enzyme of anaerobic metabolism, is induced by iron deficiency in barley roots. Plant Physiol 116:725–732. https://doi.org/10.1104/pp.116.2.725
Tripp J, Mishra SK, Scharf KD (2009) Functional dissection of the cytosolic chaperone network in tomato mesophyll protoplasts. Plant Cell Environ 32:123–133. https://doi.org/10.1111/j.1365-3040.2008.01902.x
Volkov RA, Panchuk II, Schöffl F (2005) Small heat shock proteins are differentially regulated during pollen development and following heat stress in tobacco. Plant Mol Biol 57:487–502. https://doi.org/10.1007/s11103-005-0339-y
Wang A, Yu X, Mao Y et al (2015) Overexpression of a small heat-shock-protein gene enhances tolerance to abiotic stresses in rice. Plant Breed 134:384–393. https://doi.org/10.1111/pbr.12289
Waters ER (2013) The evolution, function, structure, and expression of the plant sHSPs. J Exp Bot 64:391–403. https://doi.org/10.1093/jxb/ers355
Waters ER, Aevermann BD, Sanders-Reed Z (2008) Comparative analysis of the small heat shock proteins in three angiosperm genomes identifies new subfamilies and reveals diverse evolutionary patterns. Cell Stress Chaperones 13:127–142. https://doi.org/10.1007/s12192-008-0023-7
Wehmeyer N, Vierling E (2000) The expression of small heat shock proteins in seeds responds to discrete developmental signals and suggests a general protective role in desiccation tolerance. Plant Physiol 122:1099–1108. https://doi.org/10.1104/pp.122.4.1099
Wehmeyer N, Hernandez LD, Finkelstein RR, Vierling E (1996) Synthesis of small heat-shock proteins is part of the developmental program of late seed maturation. Plant Physiol 112:747–757. https://doi.org/10.1104/pp.112.2.747
Wesley SV, Helliwell CA, Smith NA et al (2001) Construct design for efficient, effective and high-throughput gene silencing in plants. Plant J 27:581–590
Yeh CH, Yeh KW, Wu SH et al (1995) A recombinant rice 16.9-kDa heat shock protein can provide thermoprotection in vitro. Plant Cell Physiol 36:1341–1348
Yeh CH, Chang PF, Yeh KW et al (1997) Expression of a gene encoding a 16.9-kDa heat-shock protein, Oshsp16.9, in Escherichia coli enhances thermotolerance. Proc Natl Acad Sci USA 94:10967–10972. https://doi.org/10.1073/pnas.94.20.10967
Young LS, Yeh CH, Chen YM, Lin CY (1999) Molecular characterization of Oryza sativa 16.9 kDa heat shock protein. Biochem J 344(Pt 1):31–38. https://doi.org/10.1042/0264-6021:3440031
Zhao C, Liu B, Piao S et al (2017) Temperature increase reduces global yields of major crops in four independent estimates. Proc Natl Acad Sci USA 114:9326–9331. https://doi.org/10.1073/pnas.1701762114
Zhou Y, Chen H, Chu P et al (2012) NnHSP17.5, a cytosolic class II small heat shock protein gene from Nelumbo nucifera, contributes to seed germination vigor and seedling thermotolerance in transgenic Arabidopsis. Plant Cell Rep 31:379–389. https://doi.org/10.1007/s00299-011-1173-0
Zivy M, Thiellement H, de Vienne D, Hofmann JP (1984) Study on nuclear and cytoplasmic genome expression in wheat by two-dimensional gel electrophoresis. Theor Appl Genet 68:335–345
Zou J, Liu C, Liu A et al (2012) Overexpression of OsHsp17.0 and OsHsp23.7 enhances drought and salt tolerance in rice. J Plant Physiol 169:628–635. https://doi.org/10.1016/j.jplph.2011.12.014
Acknowledgements
We thank Tsung-Luo Jinn, National Taiwan University, Taiwan for providing 35S: EYFP plasmid. AG thanks Department of Biotechnology (BT/AGIII/CARI/01/2012 dated 11.07.2013) and J.C. Bose fellowship award, Department of Science and Technology, Government of India for the financial support.
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Sarkar, N.K., Kotak, S., Agarwal, M. et al. Silencing of class I small heat shock proteins affects seed-related attributes and thermotolerance in rice seedlings. Planta 251, 26 (2020). https://doi.org/10.1007/s00425-019-03318-9
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DOI: https://doi.org/10.1007/s00425-019-03318-9