Gene characterization and molecular pathway analysis of reverse thermosensitive genic male sterility in eggplant (Solanum melongena L.)

Abstract

The naturally occurring mutant eggplant line 05ms was identified with reverse thermosensitive genic male sterility (rTGMS), but its temperature-responsive fertility mechanisms remain largely unknown. Here, we studied the flower morphology, anther cellular structure, and genome-wide gene expression of this rTGMS line. Candidate genes for thermosensitive male sterility during the microspore development of 05ms and the temperature-insensitive line S63 under low-temperature (LT) and high-temperature (HT) conditions were identified. Under LT, tapetum cells were vacuolated and had delayed disintegration in 05ms. RNA-seq analysis indicated that DEGs were enriched in the KEGG pathways ‘plant hormone signal transduction’, ‘starch and sucrose metabolism’, and ‘phenylpropanoid biosynthesis’. We identified two genes, 4CLL1 (Sme2.5_00368.1_g00010.1) and CKI1 (Sme2.5_10056.1_g00002.1), which could potentially regulate eggplant anther development and may be candidate genes for rTGMS. Finally, we propose a working model of anther abortion for rTGMS in eggplant. CKI1 responds to LT stress and causes expression changes in genes related to anther development, such as 4CLL1, and the cellular structure of the tapetum becomes abnormal, causing male sterility. The findings of this study explain the underlying molecular mechanisms of male sterility in eggplant rTGMS lines.

References

[1] Chen, L. & Liu, Y. G. Male sterility and fertility restoration in crops. Annu. Rev. Plant Biol. 65, 579-606 (2014).

[2] Shemeshmayer, E. et al. Garlic (Allium sativum L.) fertility: transcriptome and proteome analyses provide insight into flower and pollen development. Front. Plant Sci. 6, 271 (2015).

[3] Shu, J. et al. Detection of the diversity of cytoplasmic male sterility sources in broccoli (Brassica Oleraceavar.Italica) using mitochondrial markers. Front. Plant Sci. 7, 927 (2016).

[4] Lee, Y. P. et al. Discovery of a novel cytoplasmic male-sterility and its restorer lines in radish (Raphanus sativus L.). Theor. Appl. Genet. 117, 905-913 (2008).

[5] Qiu, Y., Liao, L., Jin, X., Mao, D. & Liu, R. Analysis of the meiotic transcriptome reveals the genes related to the regulation of pollen abortion in cytoplasmic male-sterile pepper (Capsicum annuum L.). Gene 641, 8 (2018).

[6] Yoshimi, M. et al. Variations in the structure and transcription of the mitochondrial atp and cox genes in wild Solanum species that induce male sterility in eggplant (S. melongena). Theor. Appl. Genet. 126, 1851-1859 (2013).

[7] Fan, Y. & Zhang, Q. Genetic and molecular characterization of photoperiod and thermo-sensitive male sterility in rice. Plant Reprod. 31, 1-12 (2017).

[8] Xue, Z. et al. Deficiency of a triterpene pathway results in humidity-sensitive genic male sterility in rice. Nat. Commun. 9, 604-613 (2018).

[9] Wang, W. et al. Comparative transcriptomes profiling of photoperiod-sensitive male sterile rice nongken 58S during the male sterility transition between short-day and long-day. BMC Genomics. 12, 462- 462 (2011).

[10] Huang, J. Z., Zhi-Guo, E., Zhang, H. L. & Shu, Q. Y. Workable male sterility systems for hybrid rice: genetics, biochemistry, molecular biology, and utilization. Rice 7, 1-14 (2014).

[11] Jia, J. H. et al. Molecular mapping of the reverse thermo-sensitive genic male-sterile gene (rtms1) in rice. Theor. Appl. Genet. 103, 607-612 (2001).

[12] Yu, C. et al. Premature breakdown of tapetum associated with reverse thermo-sensitive genic male-sterile line Huiyou50S in rapeseed (Brassica napus). Acta Physiologiae Plant. 38, 54 (2016).

[13] Omidvar, V., Mohorianu, I., Dalmay, T. & Fellner, M. Identification of miRNAs with potential roles in regulation of anther development and male-sterility in 7B-1 male-sterile tomato mutant. BMC Genomics. 16, 878 (2015).

[14] Sawhney, V. K. Temperature control of male sterility in a tomato mutant. J. Hered. 74, 51-54 (1983).

[15] Kim, Y. M. et al. Characterization and inheritance of a novel thermo-sensitive restoration of cytoplasmic male sterility in Capsicum annuum. Sci. Hortic. Amst. 164(Complete), 512-520 (2013).

[16] Tang, Z., Zhang, L., Yang, D. I., Zhao, C. & Zheng, Y. Cold stress contributes to aberrant cytokinesis during male meiosis I in a wheat thermosensitive genic male sterile line. Plant Cell Environ. 34, 389-405 (2011).

[17] Li, B., Gao, X. R., Wu, Y. R., Wang, H. C. & Pan, X. Q. On discover, identification and genetic analysis of a new male sterile line in eggplant. Chin. J. Southwest China Norm. Univ. (Nat. Sci. Ed.). 38, 59-62 (2013).

[18] Zhou, H. et al. Development of commercial thermo-sensitive genic male sterile rice accelerates hybrid rice breeding using the CRISPR/Cas9-mediated TMS5 editing system. Sci. Rep. 6, 37395 (2016).

[19] Zhou, H. et al. RNase Z(S1) processes UbL40 mRNAs and controls thermosensitive genic male sterility in rice. Nat. Commun. 5, 4884 (2014).

[20] Millar, A. A. & Gubler, F. The Arabidopsis GAMYB-like genes, MYB33 and MYB65, are microRNA-regulated genes that redundantly facilitate anther development. Plant Cell. 17, 705-721 (2005).

[21] Mizuno, S. et al. Receptor-like protein kinase 2 (RPK2) is a novel factor controlling anther development in Arabidopsis thaliana. Plant J. 50, 751-766 (2010).

[22] Yu, J. et al. Two rice receptor-like kinases maintain male fertility under changing temperatures. Proc. Natl Acad. Sci. USA 114, 12327-12332 (2017).

[23] Zhang, H. et al. Mutation in CSA creates a new photoperiod-sensitive genic male sterile line applicable for hybrid rice seed production. PNAS 110, 76-81 (2013).

[24] min, L. et al. Sugar and auxin signaling pathways respond to high-temperature stress during anther development as revealed by transcript profiling analysis in cotton. Plant Physiol. 164, 1293 (2014).

[25] Hirakawa, H. et al. Draft genome sequence of eggplant (Solanum melongena L.): the representative solanum species indigenous to the old world. DNA Res. Int. J. rapid Publ. Rep. genes genomes 21, 649-660 (2014).

[26] Yang, Y. et al. The key genes and pathways related to male sterility of eggplant revealed by comparative transcriptome analysis. BMC Plant Biol. 18, 209-221 (2018).

[27] Han, Y. et al. Fine mapping of a male sterility gene ms-3 in a novel cucumber (Cucumis sativus L.) mutant. Theor. Appl. Genet. 131, 1-12 (2018).

[28] Zhang, Z. B. et al. Transcription factor AtMYB103 is required for anther development by regulating tapetum development, callose dissolution and exine formation in Arabidopsis. Plant J. 52, 528-538 (2010).

[29] Xu, J. et al. Aborted microspores acts as a master regulator of pollen wall formation in Arabidopsis. Plant Cell. 26, 1544 (2014).

[30] Singh, R. P., Prasad, P. V. V. & Reddy, K. R. Chapter two-impacts of changing climate and climate variability on seed production and seed industry. Adv. Agron. 118, 49-110 (2013).

[31] Oliver, S. N. Cold-induced repression of the rice anther: specific cell wall invertase gene OSINV4 is correlated with sucrose accumulation and pollen sterility. Plant, Cell Environ. 28, 1534-1551 (2005).

[32] Fu, Z. et al. The rice basic helix-loop-helix transcription factor TDR Interacting protein2 is a central switch in early anther development. Plant Cell. 26, 1512 (2014).

[33] Schneider, K. et al. The substrate specificity-determining amino acid code of 4-coumarate: CoA ligase. Proc. Natl Acad. Sci. USA 100, 8601-8606 (2003).

[34] Li, Y. et al. OsACOS12, an orthologue of Arabidopsis acyl-CoA synthetase5, plays an important role in pollen exine formation and anther development in rice. BMC Plant Biol. 16, 256 (2016).

[35] Yang, X. et al. Rice fatty acyl-CoA synthetase OsACOS12 is required for tapetum programmed cell death and male fertility. Planta 246, 1-18 (2017).

[36] Souza, C. D. A., Barbazuk, B., Ralph, S. G., Bohlmann, J. & Douglas, C. J. Genome-wide analysis of a land plant-specific acyl:coenzymeA synthetase (ACS) gene family in Arabidopsis, poplar, rice and Physcomitrella. N. Phytologist. 179, 987-1003 (2008).

[37] Lin, Y. Functional analysis of anther-specific genes essential for pollen exine development and male fertility in tobacco. Ph. D thesis from The University of Hong Kong (2012).

[38] Tuazon, P. T. & Traugh, J. A. Casein kinase I and II-multipotential serine protein kinases: structure, function, and regulation. Adv. Second Messenger Phosphoprot. Res. 23, 123 (1991).

[39] Moriya, H. & Johnston, M. Glucose sensing and signaling in Saccharomyces cerevisiae through the Rgt2 glucose sensor and casein kinase I. PNAS 101, 1572-1577 (2004).

[40] Li, Y. et al. Promoters of Arabidopsis Casein kinase I-like 2 and 7 confer specific high-temperature response in anther. Plant Mol. Biol. 98, 33-49 (2018).

[41] Liu, W. et al. Roles of OsCKI1, a rice casein kinase I, in root development and plant hormone sensitivity. Plant J. Cell Mol. Biol. 36, 189-202 (2010).

[42] min, L. et al. Cotton GhCKI disrupts normal male reproduction by delaying tapetum programmed cell death via inactivating starch synthase. Plant J. 75, 823-835 (2013).

[43] Cheng, Y. F. et al. Cytological and comparative proteomic analyses on male sterility in Brassica napus L. induced by the chemical hybridization agent monosulphuron ester sodium. Plos One 8, e80191 (2013).

[44] Li, H. & Durbin, R. Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics 25, 1754-1760 (2009).

[45] Li, H. et al. The sequence alignment/map format and SAMtools. Bioinformatics 25, 2078-2079 (2009).

[46] McKenna, A. et al. The Genome Analysis Toolkit: a MapReduce framework for analyzing next-generation DNA sequencing data. Genome Res. 20, 1297-1303 (2010).

[47] Wang, K., Li, M. & Hakonarson, H. ANNOVAR: functional annotation of genetic variants from high-throughput sequencing data. Nucleic acids Res. 38, e164 (2010).

[48] Takagi, H. et al. QTL-seq: rapid mapping of quantitative trait loci in rice by whole genome resequencing of DNA from two bulked populations. Plant J. 74, 174-183 (2013).

[49] Livak, K. J. & Schmittgen, T. D. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods 25, 402-408 (2001).