Abstract
In eukaryotes, alternative splicing (AS) is a crucial regulatory mechanism that modulates mRNA diversity and stability. The contribution of AS to stress is known in many species related to stress, but the posttranscriptional mechanism in poplar under cold stress is still unclear. Recent studies have utilized the advantages of single molecular real-time (SMRT) sequencing technology from Pacific Bioscience (PacBio) to identify full-length transcripts. We, therefore, used a combination of single-molecule long-read sequencing and Illumina RNA sequencing (RNA-Seq) for a global analysis of AS in two poplar species (Populus trichocarpa and P. ussuriensis) under cold stress. We further identified 1,261 AS events in P. trichocarpa and 2,101 in P. ussuriensis among which intron retention, with a frequency of more than 30%, was the most prominent type under cold stress. RNA-Seq data analysis and annotation revealed the importance of calcium, abscisic acid, and reactive oxygen species signaling in cold stress response. Besides, the low temperature rapidly induced multiple splicing factors, transcription factors, and differentially expressed genes through AS. In P. ussuriensis, there was a rapid occurrence of AS events, which provided a new insight into the complexity and regulation of AS during cold stress response in different poplar species for the first time.
Highlights
In eukaryotes, precursor messenger RNAs with multiple introns undergo alternative splicing (AS) to generate two or more mature mRNA isoforms, encoding structurally and functionally different proteins (Palusa et al, 2007)
We found that several differentially expressed genes (DEGs) related to cold stress, such as cold-regulated inner membrane protein 1 (CORIMP1, Potri.001G353500) at 25◦C, low-temperature- and salt-responsive proteins (LTI6A, Potri.013G001600 and CORIMP1Potri.001G353500) at 3◦C, and late elongated hypocotyl (LHY, Potri.014G106800), lowtemperature-induced integral membrane protein (LTI6A, Potri.013G001600), and calcium-binding protein (CML, Potri.002G182500, Potri.014G108200) at −3◦C in P. ussuriensis (Table 4) had more isoforms based on P. trichocarpa genome
We found that calcium signaling-related genes, such as CAM, calcium-dependent protein kinase (CPK), and CBL-interacting protein kinase (CIPK), and many mitogen-activated protein kinase (MAPK) cascades were upregulated in P. ussuriensis at 25, 3, and −3◦C compared with P. trichocarpa
Summary
Precursor messenger RNAs (pre-mRNAs) with multiple introns undergo alternative splicing (AS) to generate two or more mature mRNA isoforms, encoding structurally and functionally different proteins (Palusa et al, 2007). AS is a crucial regulatory mechanism that contributes to cellular and functional complexity and has been extensively studied in animals and plants (Wang et al, 2018; Li et al, 2019). Genome-wide investigation of AS has been performed on development or in response to stresses in multiple plants. Studies have revealed that an estimated 60% of Arabidopsis, 50% of soybean, 40% of cotton, and 40% of maize intron-containing genes undergo AS (Marquez et al, 2012; Syed et al, 2012; Li et al, 2019).
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