Abstract
The basic-helix-loop-helix (bHLH) transcription factors superfamily is present in eukaryotes and has been widely characterized in several species. The study of these proteins has focused on processes involved in plant growth and development. However, it has also been shown that bHLH plays a relevant role in processes of response to abiotic stress. In this study, the bHLH family in Eucalyptus was systematically characterized for the first time, using the Eucalyptus grandis genome. 153 bHLH genes in the E. grandis genome were identified, which were characterized phylogenetically and structurally. Then, a cold-tolerant hybrid of Eucalyptus (Eucalyptus nitens × Eucalyptus globulus) to generate a reference assembly was used to study the alternative splicing events produced in the cold response. 44 bHLH candidate genes to generate alternative splicing events in the hybrid were identified. These genes had exons differentially expressed in cold acclimation conditions. The splicing event prediction was further validated by qRT-PCR in three bHLH genes. Through this experiment was possible to determine that two of them were differentially expressed, so that new isoforms of these genes were being produced under cold acclimation conditions.
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Acknowledgements
The computing capacity was provided by Facultad de Ciencias Biológicas, Universidad de Concepción. The authors would like to acknowledge plant biotechnologists, Catalina Lagos, Mariana Arias, and Stephanie Riquelme, for their support in the cold chamber assay.
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Financial support came from Fondecyt Iniciación 11121559 and Fondecyt 1161063, Genómica Forestal S.A.
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VE provided tested and selected individuals of E. nitens and hybrids. MF performed growth chamber experiments. HA performed the annotation and characterization of bHLH genes, alternative splice event identification, expression analysis, and validation by qRT-PCR. DC performed GO analysis. MS and AF performed the transcriptome de novo assembly. JC performed the transcriptome annotation. SV provided cDNA samples, reagents for qRT-PCR analysis, and support in laboratory procedures. ETS provided technical support for bioinformatics analysis and contributed in the bioinformatics procedures. HA, MS, and MF drafted the manuscript. ETS and SV collaborated in experimental design of the project. MF and HA designed the project. MF initiated and led the project. All authors contributed to manuscript preparation and editing. All authors read and approved the final manuscript.
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• This work reported that changes in alternative splicing (AS) in bHLH transcription factors (TFs) of a cold-tolerant hybrid genotype of Eucalyptus nitens × Eucalyptus globulus may contribute to rapid changes in gene expression during cold acclimation, suggesting that AS events may play an important regulatory role in this process
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Apablaza, H., Solís, M., Conejera, D. et al. bHLH Transcription Factors Undergo Alternative Splicing During Cold Acclimation in a Eucalyptus hybrid. Plant Mol Biol Rep 40, 310–326 (2022). https://doi.org/10.1007/s11105-021-01313-7
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Issue Date:
DOI: https://doi.org/10.1007/s11105-021-01313-7