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Light dramatically affects alternative splicing[6,47]. Critical genes of light signaling pathways in plants, including PHYB, PHYA, CRY2, and CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1), act similarly to SFs in determining AS (Fig. 3 & Table 1). Among them, PHYB regulates photomorphogenesis, thermomorphogenesis, and freezing tolerance[48,49]. PHYB could also directly interact with SPLICING FACTOR FOR PHYTOCHROME SIGNALING (SFPS) and red-light responses in CRY1CRY2 background1 (RRC1) in nuclear speckles to synergistically regulate pre-mRNA splicing of a group of genes involved in light signaling and circadian clock pathways[12,35,50−52]. Furthermore, PHYA regulates AS via interaction with the Arabidopsis homologue of human CNOT9 (NOT9B)[53]. During photomorphogenesis, COP1 can regulate AS of U2 small nuclear ribonucleoprotein auxiliary factor 65A (U2AF65A) and SR30 by interacting with U2AF65-associated protein (UAP56)[54]. Hence, we can conclude that these proteins in light signaling pathways can directly regulate SFs in order to modulate AS in response to changes in the light environment.
Figure 3.
The major pathway for regulating pre-mRNA AS under light and temperature stresses. At first, some SFs could act as sensors to perceive the changes in light and temperatures. Secondly, those sensors will phosphorylate or interact with other SFs and change their location and activity. These modifications of SFs will affect AS by controlling spliceosome function. Finally, plant growth, development, flower, fertility, and tolerance are regulated under adverse light and temperature conditions.
Table 1. Splicing factors involved in light and temperature stress adaption.
Annotation Name Species Gene ID Function Reference DEAD-box RNA heatlicases RH42 Rice Os08G0159900 Cold Tolerance Lu et al.[65] U1 snRNP Prp39a Arabidopsis AT1G04080 Temperature-induced flowering time Chang et al.[73] hnRNPs RBP45d Arabidopsis AT5G19350 Temperature-induced flowering time Chang et al.[73] Phytochrome CRY2 Arabidopsis AT1G04400 Blue-light regulation of thermosensory flowering Zhao et al.[74] mRNA binding SF1/BBP Arabidopsis AT5G51300 Temperature-induced flowering time Lee et al.[71] hnRNP A/B family UBA2c Arabidopsis AT3G15010 Light and temperature-induced flowering time Zhao et al.[72] Core proteins (Sm) SmE-b/PCP/SME1 Arabidopsis AT2G18740 Arrest of growth and male sterile at
low temperatureCapovilla et al.[17] U1 snRNP Luc7a Arabidopsis AT3G03340 Low temperature resulted in arrest
of growthAmorim et al.[63] U1 snRNP Luc7b Arabidopsis AT5G17440 Low temperature resulted in arrest
of growthAmorim et al.[63] U1 snRNP Luc7-rl Arabidopsis AT5G51410 Low temperature resulted in arrest
of growthAmorim et al.[63] Glycine rich protein GRPs-7 Arabidopsis AT2G30560 Cold tolerance Kim et al.[62] DEAD box RNA helicase Prp5-1b/RCF1 Arabidopsis AT1G20920 Cold tolerance and cold acclimion Guan et al.[59] U5 snRNP U5-200-2b/Brr2b Arabidopsis AT2G42270 Cold tolerance and cold acclimion Guan et al.[59] DEAD-box RNA helicase U2B"a/U2B"-LIKE Arabidopsis AT1G06960 Cold tolerance and cold acclimion Calixto et al.[64] SR protein kinase AFC3/AME3 Arabidopsis AT4G32660 Cold tolerance and cold acclimion L. Savitch, unpublisheatd data 17S U2 snRNP GEMIN2 Arabidopsis AT1G54380 Cold tolerance and cold acclimion Schlaen et al.[61] U5 snRNP U5-102KD/STA1 Arabidopsis AT4G03430 Cold tolerance and cold acclimion;
heat tolerance and heat acclimionKim et al.[66];
Guan et al.[59]Inositol-requiring enzyme-1 IRE1b Arabidopsis AT5G24360 Heat stress responses Deng et al.[39]; Moreno et al.[38] Inositol-requiring enzyme-1 IRE1a Arabidopsis AT2G17520 Heat stress responses Deng et al.[39]; Moreno et al.[38] hnRNP HOS5/RCF3 Arabidopsis AT5G53060 Heat stress tolerance Guan et al.[60] HSP70-3 HSP70-3 Arabidopsis AT3G09440 Heat stress tolerance Song et al.[56] HSP70-4 HSP70-4 Arabidopsis AT3G12580 Heat stress tolerance Wang et al.[57] HSC70-1 HSC70-1 Arabidopsis AT5G02500 Heat stress tolerance Tiwari et al.[58] Cyclin-dependent kinase CDCL2p110-1 Arabidopsis AT1G67580 High temperature-induced male fertility defects Cavallari et al.[70]; Zheng et al.[69]; Nibau et al.[68] Cyclin-dependent kinase CDCL2p110-2/CDKG1 Arabidopsis AT5G63370 High temperature induce fertility defects Cavallari et al.[70]; Zheng et al.[69]; Nibau et al.[68] SR protein Kinase AFC2 Arabidopsis AT4G24740 Thermosensors and thermomorphogenesis Lin et al.[88] Phytochrome PHYB Arabidopsis AT2G18790 Thermosensors and thermomorphogenesis;
photoreceptors and photomorphogenesisJung et al.[48]; Shikata et al.[12,51]; Hartmann et al.[52]; Xin et al.[35,50] Phytochrome PHYA Arabidopsis AT1G09570 Photomorphogenesis Shikata et al.[12]; Schwenk et al.[53] E3 ubiquitin ligase COP1 Arabidopsis AT2G32950 Photomorphogenesis Li et al.[54] 17S U2 associated proteins SR140-1/RRC1 Arabidopsis AT5G25060 Photomorphogenesis Shikata et al.[12,51]; Hartmann et al.[52]; Xin et al.[35,50] SFs are also discovered to play a critical role in regulating plant temperature stress tolerance. Heat shock proteins (HSP70s), as a spliceosome component, are involved in the repair of AS during heat stress[55]. In Arabidopsis, HSP70-3, HSP70-4, and heat shock cognate protein 70–1 (HSC70–1) are all required for heat stress tolerance[56−58]. Apart from heat shock protein, a nuclear-localized SF with a K homology (KH) domain, REGULATOR OF CBF GENE EXPRESSION 3 (RCF3) serves as a negative regulator in heat stress tolerance[59,60]. For cold stress, SFs including RCF1, U5 small nuclear ribonucleoprotein helicase (Brr2b)[59],Gemin2[61], GRP7[62], the LAMMER kinase 3 (AME3), LETHAL UNLESS CBC 7 (LUC7)[63], and U2 small nuclear ribonucleoprotein particle (snRNP)-specific proteins (U2B”-LIKE)[64] play important roles in cold acclimation and tolerance in Arabidopsis. In rice, DEAD-box RNA Helicase42 (OsRH42) modulates pre-mRNA splicing at low temperatures by directly interacting with U2 small nuclear RNA. OsRH42-knockdown and OsRH42 overexpression dramatically affect the pre-mRNA splicing pattern, results in retarded plants growth and impaired cold tolerance[65]. Interestingly, the U5-snRNP-interacting protein homolog STABILIZED1 (STA1) is necessary for both cold and heat stress tolerance (Fig. 3 & Table 1). STA1 acts as a high temperature-induced SF for controlling AS of HSFA3[66]. Cold stress has also been shown to enhance STA1 gene expression, and the sta1-1 mutant is defective in splicing the cold-induce gene COR15[67]. Thus, the preceding studies indicate that the majority of SFs positively influence temperature stress tolerance.
Under adverse temperature conditions, SFs can also have a significant impact on plant development, flowering, and fertility. Sm protein E1 (SME1), for instance, is a temperature-specific SF in Arabidopsis, and mis-splicing WUSCHEL and CLAVATA3 rendered sme1 mutants growth and male sterility[17]. Similarly, CDKG1 regulates fertility under temperature stress and can interact with splicing factor Arginine/serine-rich zinc knuckle-containing protein33 (RSZ33), U2AF65A, and RS-containing zinc finger protein22 (SRZ22) to facilitate the splicing of the sixth intron of CalS5, consequently modifying pollen wall development (Fig. 3)[68,69]. CDKG1 can also be spliced into two transcript variants (CDKG1L and CDKG1S), and the ratio between long and short forms is regulated by temperature and CDKG2[46,70]. Furthermore, SFs are primarily engaged in flowering time control. In response to temperature stress, AtSF1[71], UBIQUITIN-SPECIFIC PROTEASE 1-ASSOCIATED PROTEIN 2C (UBA2c)[72], RNA binding protein 45d (RBP45d)[73], CRY2[74], U2AF65A[74], CRY2 INTERACTING SPLICING FACTOR 1 (CIS1)[74], PRP39a, CYCLYN L1 (CYCL1), and CDKG2[68], all of them can govern the AS of FLM to fine-tune flowering time. Among them, CDKG2 together with its cognate CYCL1 controls the AS of FLM. CRY2 can promote CIS1 binding to pre-mRNA of FLM, then CIS1 interact with SF1 and U2AF65A to recruit U2-snRNP to control AS of FLM. Thus, the CRY2–CIS1–U2AF65A/SF1–FLM signaling pathway shows the specific roles of AS under light and temperature stresses (Fig. 3).
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Traditionally, most studies only examine at one major splicing variant, and it is also presumed that the splicing variant of interest can be translated into proteins. One gene can produce several or even dozens of splicing variants through AS. Yet, only some splicing variants can be translated into distinct proteins. Other splicing variants without coding capacities maybe act as active non-coding RNA (Fig. 1). It is not enough to explain the gene function by analyzing one or two splicing variants with coding capacities. In plants, splicing factors and splicing codes are two major intrinsic factors in the control of AS. Many studies have proven that DNA polymorphism in splicing code regions could lead to changes in AS and phenotype[82−86]. SFs are another factor controlling AS, and there is a complex hierarchical regulatory relationship between SFs (Fig. 3). SFs including receptor-like kinase receptor FERONIA (FER) and LAMMER kinase 2(AFC2) could act as sensors to detect changes in suboptimal light and temperature and subsequently phosphorylate or interact with other SFs (GRP7 and RSZ21) to alter their location or activity in regulation of AS of Abscisic acid response element binding factor 1 (ABF1) and PIF4[87−89]. Furthermore, the interaction between SFs and splicing codes cannot be neglected since SFs can also influence splicing fidelity by restricting the usage of novel splicing code[23]. Consequently, a thorough investigation of splicing codes, splicing factors, and their effects on AS under stress will allow us to better understand and exploit it in the future for vegetable crops.
CRISPR-Cas9 is the most advanced genome editing approach, and it has been extensively used in precision breeding of important cereals and vegetable crops[90,91]. AS modulates many thermomorphogenesis and photomorphogenesis genes in Arabidopsis, including CRY1, PHYB, and EARLY FLOWERING 3 (ELF3). They are also important in modulating seedling elongation under low light or high temperature stresses[92]. Yet, knockout of these genes by CRISPR/Cas9 will make plants extremely susceptible to low light and high temperature. They will also cause undesirable alterations in plant growth and development, including flowering timing, fruit set, seed germination, and so on[93]. Whether we can edit their splicing code (intron or UTR region) or splicing factors to manipulate AS of these genes by CRISPR/Cas9 to produce critical splicing variants for inhibiting excessive elongation but without other undesired effects? Thus, we propose an application potential of this AS mechanism for vegetable crops in (Fig. 5) and hope that this research strategy will promote studies on AS regulation in combination with gene editing technology under light and temperature stresses, which can be used in the future for the genetic improvement of vegetable crops.
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Cite this article
Cao H, Wu T, Shi L, Li Y, Zhang C. 2023. Alternative splicing control of light and temperature stress responses and its prospects in vegetable crops. Vegetable Research 3:17 doi: 10.48130/VR-2023-0017
Alternative splicing control of light and temperature stress responses and its prospects in vegetable crops
- Received: 20 December 2022
- Accepted: 17 March 2023
- Published online: 02 June 2023
Abstract: Vegetable crop quality and productivity are impaired by adverse light and temperature stresses, such as low light, cold, and heat stress. Alternative splicing is a powerful strategy for coping with these abiotic stresses by producing multiple mRNA splicing variants with different subcellular locations, translation efficiency, and coding sequences. Pre-mRNA is alternatively spliced by the spliceosome consisting of different splicing-related proteins termed splicing factors. Recent studies have shown that splicing factors and splicing variants have diverse effects on the development, flowering, and fertility of model plants under light and temperature stresses, but there is very little information available for vegetable crops. Therefore, we conducted a comprehensive literature review on the roles of alternative splicing in regulating plants' responses to light and temperature stress. Several suggestions for future studies and implementation in the field of vegetable crops are presented. We can manipulate alternative splicing using CRISPR/Cas9 technology to produce splicing variants with diverse roles for improving performance of vegetable crop under stress environments.
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Key words:
- Alternative splicing /
- Splicing factor /
- Light and temperature stresses /
- Spliceosome /
- CRISPR/Cas9