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alternative splicing of MaMYB16L regulates starch degradation in banana fruit during ripening

Journal of Integrative Plant Biology 2021年第7期63卷 1341-1352页

作者： Guoxiang Jiang Dandan Zhang Zhiwei Li Hanzhi Liang Rufang Deng Xinguo Su Yueming Jiang Xuewu Duan South China Botanical Garden the Chinese Academy of SciencesGuangzhou510650 China Center of Economic Botany Core Botanical Gardensthe Chinese Academy of SciencesGuangzhou510650 China College of Advanced Agricultural Sciences University of Chinese Academy of SciencesBeijing100049 China Guangdong AIB Polytechnic Guangzhou510507 China

The alternative splicing of select genes is an important mechanism to regulate responses to endogenous and environmental signals in plants.However,the role of alternative splicing in regulating fruit ripening remains unclear.Here,we discovered that MaMYB16L,an R1-type MYB transcription factor,undergoes alternative splicing and generates two transcripts,the full-length isoform MaMYB16L and a truncated form MaMYB16S,in banana fruit.During banana fruit ripening,the alternative splicing process intensifies with downregulated MaMYB16L and upregulated MaMYB16S.Moreover,MaMYB16L is a transcriptional repressor that directly binds with the promoters of many genes associated with starch degradation and MaDREB2,a positive ripening regulator,and represses their expression.In contrast,MaMBY16S lacks a DNA-binding domain but competitively combines and forms non-functional heterodimers with functional MaMYB16L.MaMYB16L-MaMYB16S heterodimers decrease the binding capacity and transrepression activity of MaMYB16L.The downregulation of MaMYB16L and the upregulation of MaMYB16S,that is,a decreased ratio of active to non-active isoforms,facilitates the activation of ripening-related genes and thereby promotes fruit ripening.Furthermore,the transient overexpression of MaMYB16S promotes banana fruit ripening,whereas the overexpression of MaMYB16L delays this process.Therefore,the alternative splicing of MaMYB16L might generate a self-controlled regulatory loop to regulate banana fruit ripening.

关键词： alternative splicing fruit ripening Musa acuminata R1-MYB transcriptional regulation

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alternative splicing patterns of doublesex reveal a missing link between Nix and doublesex in the sex determination cascade of Aedes albopictus

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Insect Science 2021年第6期28卷 1601-1620页

作者： Binbin Jin Yijie Zhao Yunqiao Dong Peiwen Liu Yan Sun Xiaocong Li Xin Zhang Xiao-Guang Chen Jinbao Gu Department of Pathogen Biology Guangdong Provincial Key Laboratory of Tropical Disease ResearchSchool of Public HealthSouthern Medical UniversityGuangzhou510515China Reproductive Medical Centre of Guangdong Women and Children Hospital Guangzhou511442China

Sexual development in insects is regulated by a complicated hierarchical cascade of sex determination.The primary signals are diverse,whereas the central nexus doublesex(dsx)gene is relatively conserved within the pathway.Aedes(Stegomyia)albopictus is an important vector with an extensive worldwide distribution.We previously reported that Ae.albopictus dsx(Aalbdsx)yields one male-(Aalbdsx^(M))and three female-specific isoforms(Aalbdsx^(F1-3));however,the spatiotemporal expression profiles and mechanisms regulating sex-specific alternative splicing require further investigation.In this study,we demonstrated that the Aalbdsx^(M) messenger RNA(mRNA)represents the default pattern when analyzed in human foreskin fibroblasts and HeLa cells.We combined reverse transcription polymerase chain reaction with RNA immunoprecipitation using specific antibodies against tagged Ae.albopictus male-determining factor AalNix and confirmed that AalNix indirectly regulates dsx pre-mRNA and regulates its alternative splicing.During the early embryo stage(0-2 and 4-8 h),maternal dsx^(F) and default splicing dsx^(M) were detected in both sexes;the expression of dsx^(M) then decreased until sufficient AalNix transcripts accumulated in male embryos at 20-24 h.These findings suggest that one or more potential dsx splicing enhancers can shift dsx^(M) to dsx^(F) in both sexes;however,the presence of Nix influences the function of this unknown splicing enhancer and ultimately leads to the formation of dsx^(M) in males.Finally,our results provide important insight into the regulatory mechanism of dsx alternative splicing in the mosquito.

关键词： Aedes albopictus alternative splicing Doublesex\mosquito control sex determination

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alternative splicing of inner-ear-expressed genes

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Frontiers of Medicine 2016年第3期10卷 250-257页

作者： Yanfei Wang Yueyue Liu Hongyun Nie Xin Ma Zhigang Xu Shandong Provincial Key Laboratory of Animal Cells and Developmental Biology School of Life Sciences Shandong University Jinan 250100 China School of Control Science and Engineering Shandong University Jinan 250061 China

alternative splicing plays a fundamental role in the development and physiological function of the inner ear. Inner-ear-specific gene splicing is necessary to establish the identity and maintain the function of the inner ear. For example, exon 68 of Cadherin 23 （Cdh23） gene is subject to inner-ear-specific alternative splicing, and as a result, Cdh23（＋ 68） is only expressed in inner ear hair cells. alternative splicing along the tonotopic axis of the cochlea contributes to frequency tuning, particularly in lower vertebrates, such as chickens and turtles. Differential splicing of Kcnmal, which encodes for the a subunit of the Ca2＋-activated K~ channel （BK channel）, has been suggested to affect the channel gating properties and is important for frequency tuning. Consequently, deficits in alternative splicing have been shown to cause hearing loss, as we can observe in Bronx Waltzer Cov） mice and Sfswap mutant mice. Despite the advances in this field, the regulation of alternative splicing in the inner ear remains elusive. Further investigation is also needed to clarify the mechanism of hearing loss caused by alternative splicing deficits.

关键词： alternative splicing inner ear hearing loss hair cells

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alternative splicing of a carboxyl/choline esterase gene enhances the fenpropathrin tolerance of Tetranychus cinnabarinus

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Insect Science 2023年第5期30卷 1255-1266页

作者： Peng Wei Xinying Zeng Haonan Han Yiqing Yang Youjun Zhang Lin He Key Laboratory of Entomology and Pest Control Engineering College of Plant ProtectionSouthwest UniversityChongqingChina Academy of Agricultural Sciences Southwest UniversityChongqingChina National Citrus Engineering Research Center Southwest UniversityChongqingChina Department of Plant Protection Institute of Vegetables and FlowersChinese Academy of Agricultural SciencesBeijingChina

Detoxification plays a crucial role in agricultural pests to withstand pesticides,and cytochrome P450s,carboxyl/choline esterases(CCEs),and glutathione-S-transferases are the main proteins responsible for their detoxification ability.The activity of CCEs can be upregulated,downregulated,or modified by mutation.However,few studies have examined the role of alternative splicing in altering the properties of CCEs.We identified 2 variants of TcCCE23 in Tetranychus cinnabarinus:a long version(CCE23-V1)and a short version that is 18 nucleotides shorter than CCE23-V1(CCE23-V2).Whether splicing affects the activity of TcCCE23 remains unclear.Overexpression of CCE23-V2 in fenpropathrin-resistant T.cinnabarinus revealed that splicing affected the detoxification of fenpropathrin by CCE23-V2.The mortality of mites was significantly higher when the expression of CCE23-V2 was knocked down(43.2%±3.3%)via injection of CCE23-dsRNA(double-stranded RNA)compared with the control group injected with green fluorescent protein-dsRNA under fenpropathrin exposure;however,the downregulation of CCE23-V1(61.3%±6.3%)by CCE23-small interfering RNA had no such effect,indicating CCE23-V2 plays a greater role in xenobiotic metabolism than CCE23-V1.The tolerance of flies overexpressing CCE23-V2 to fenpropathrin(50%lethal dose[LD_(50)]=19.47μg/g)was significantly higher than that of Gal4/UAS-CCE23-V1 transgenic flies(LD_(50)=13.11μg/g).Molecular docking analysis showed that splicing opened a“gate”that enlarges the substrate binding cavity of CCE23-V2,might enhance the ability of CCE23-V2 to harbor fenpropathrin molecules.These findings suggest that splicing might enhance the detoxifying capability of TcCCE23.Generally,our data improve the understanding of the diversity and complexity of the mechanisms underlying the regulation of CCEs.

关键词： alternative splicing carboxyl/choline esterase(CCE) detoxification function pyrethroid pests

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alternative splicing of OsRAD1 Defines C-Terminal Domain Essential for Protein Function in Meiosis

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Rice science 2020年第4期27卷 289-301页

作者： YUAN Shuting XU Chunjue YAN Wei CHANG Zhenyi DENG Xingwang CHEN Zhufeng WU Jianxin TANG Xiaoyan Guangdong Provincial Key Laboratory of Biotechnology for Plant Development School of Life SciencesSouth China Normal UniversityGuangzhou 510631China Shenzhen Institute of Molecular Crop Design Shenzhen 518107China School of Life Sciences Southern University of Science and TechnologyShenzhen 518055China

alternative splicing can generate multiple mRNAs that differ in their untranslated regions or coding sequences,and these differences might affect mRNA stability or result in different protein isoforms with diverse functions and/or localizations.In this study,we isolated a sterile mutant in rice with abnormal meiosis of microspore mother cells and megaspore mother cells that carried a point mutation in OsRAD1 gene.Cloning of OsRAD1 cDNAs revealed three transcript variants,named as OsRAD1.1,OsRAD1.2 and OsRAD1.3,respectively,which were derived from alternative splicing of the last intron.Proteins derived from the three transcripts were mostly identical except the difference in the very C-terminal domain.The three transcripts exhibited similar expression patterns in various tissues,but the expression level of OsRAD1.1 was the highest.Specific knockout of OsRAD1.1 led to sterility,while knockout of OsRAD1.2 and OsRAD1.3 together did not change the plant fertility.Overexpression of OsRAD1.2 and OsRAD1.3 cDNAs in OsRAD1.1-specific mutant did not complement the plant fertility.Yeast two-hybrid assay showed that OsRAD1.1,but not OsRAD1.2 and OsRAD1.3,interacted with the three other meiosis proteins OsHUS1,OsRAD9 and OsRAD17,suggesting that the C-terminal domain of OsRAD1.1 is critical for the protein function.

关键词： Oryza sativa OsRAD1 alternative splicing meiosis protein interaction fertility sterility

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alternative splicing of POUM2 regulates embryonic cuticular formation and tanning in Bombyx mori

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Insect Science 2023年第5期30卷 1267-1281页

作者： Chunmei Tong Kang Zhang Zixia Rong Wanyu Mo Yuling Peng Sichun Zheng Qi-Li Feng Huimin Deng Guangdong Key Laboratory of Insect Developmental Biology and Applied Technology Guangzhou Key Laboratory of Insect Development Regulation and Application ResearchInstitute of Insect Science and Technology&School of Life SciencesSouth China Normal UniversityGuangzhouChina

Insect cuticle is an apical extracellular matrix produced by the epidermis,tracheal,hind-and foregut epithelia during embryogenesis and renewed during molting and metamorphosis.However,the underlying regulatory mechanism for embryonic cuticle formation remains largely unclear.Here,we investigate the function of the transcription factor POUM2 in the embryonic cuticular formation in Bombyx mori,a model lepidopteran insect.Clustered regularly interspaced palindromic repeats(CRISPR)/CRISPR-associated protein-9-mediated knockout of POUM2 resulted in the defect of cuticular deposition,pigmentation,and sclerotization in the embryos.Differentially expressed transcripts analysis of 7-d-old embryos identified 174 up-or downregulated cuticular protein transcripts,8 upregulated chitin degradation transcripts,2 downregulated chitin synthesis transcripts and 48 up-or downregulated transcription factor transcripts in the POUM2−/−embryos.The expression levels of the key factors of the tyrosine metabolic pathway,such as tyrosine hydroxylase(Th),Dopa decarboxylase(DDC),and arylalkylamine N-acetyltransferase(aaNAT),were significantly decreased in the POUM2−/−embryos.POUM2 isoform POUM2-L specifically bound the POU cis-regulatory element(CRE)in the Th promoter and increased the transcription of Th,whereas POUM2-S could not bind the POU CRE,although it also increased the transcription of Th.Heterogeneous nuclear ribonucleoprotein Squid-1 directly bound the POUM2 pre-mRNA(messenger RNA)and inhibited the alternative splicing of POUM2-L to POUM2-S mRNA.These results suggest that POUM2 participates in the cuticular formation by regulating the chitin and cuticular protein synthesis and metabolism,and the cuticular pigmentation and sclerotization by regulating tyrosine metabolism during embryogenesis.This study provides new insights into novel function of POUM2 in embryogenesis.

关键词： alternative splicing cuticular formation embryogenesis POUM2 tanning

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alternative splicing enhances transcriptome complexity in desiccating seeds

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Journal of Integrative Plant Biology 2016年第12期58卷 947-958页

作者： Arunkumar Srinivasan Jose M. Jimenez-Gomez Fabio Fornara Wim J. J. Soppe Vittoria Brambilla Department of Plant Breeding and Genetics Max Planck Institute for Plant Breeding Research Cologne Germany Open Analytics Antwerp Belgium Institut Jean-Pierre Bourgin Institut National de la Recherche Agronomique Versailles France University of Milan Department of Biosciences Milano 20133 Italy University of Milan Department of Agricultural and Environmental Sciences via Celoria 220133 Milano Italy

Before being dispersed in the environment, mature seeds need to be dehydrated. The survival of seeds after dispersal depends on their low hydration in combination with high desiccation tolerance. These characteristics are estab- lished during seed maturation. Some key seed maturation genes have been reported to be regulated by alternative splicing （AS）. However, so far AS was described only for single genes and a comprehensive analysis of AS during seed maturation has been lacking. We investigated gene expression and AS during Arabidopsis thaliana seed development at a global level, before and after desiccation. Bioinformatics tools were developed to identify differentially spliced regions within genes. Our data suggest the importance and shows the peculiar features of AS during seed desiccation. We identified AS in 347o of genes that are expressed at both timepoints before and after desiccation. Most of these AS transcript variants had not been found before in other tissues. Among the AS genes some seed master regulators could be found. Interestingly, 6% of all expressed transcripts were not transcriptionally regulated during desiccation, but only modified by AS. We propose that AS should be more routinely taken into account in the analysis of transcriptomic data to prevent overlooking potentially important regulators.

关键词： alternative splicing Arabidopsis thaliana seed desiccation

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两种富含甘氨酸的RNA结合蛋白参与选择性剪接实现对水稻耐热性的昼夜节律调控

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Science Bulletin 2024年第1期69卷 59-71页

作者：杨闯罗安妮芦海平 Seth Jon Davis 刘建祥 State Key Laboratory of Plant Environmental Resilience College of Life SciencesZhejiang UniversityHangzhou 310027China Department of Biology University of YorkYork YO105DDUK

Rice(Oryza sativa L.)production is threatened by global warming associated with extreme high temperatures,and rice heat sensitivity is differed when stress occurs between daytime and nighttime.However,the underlying molecular mechanism are largely unknown.We show here that two glycine-rich RNA binding proteins,Os GRP3 and Os GRP162,are required for thermotolerance in rice,especially at nighttime.The rhythmic expression of Os GRP3/Os GRP162 peaks at midnight,and at these coincident times,is increased by heat stress.This is largely dependent on the evening complex component Os ELF3-2.We next found that the double mutant of Os GRP3/Os GRP162 is strikingly more sensitive to heat stress in terms of survival rate and seed setting rate when comparing to the wild-type plants.Interestingly,the defect in thermotolerance is more evident when heat stress occurred in nighttime than that in daytime.Upon heat stress,the double mutant of Os GRP3/Os GRP162 displays globally reduced expression of heat-stress responsive genes,and increases of m RNA alternative splicing dominated by exon-skipping.This study thus reveals the important role of Os GRP3/Os GRP162 in thermotolerance in rice,and unravels the mechanism on how Os GRP3/Os GRP162 regulate thermotolerance in a diurnal manner.

关键词： alternative splicing Diurnal regulation Glycine-rich protein Oryza sativa L. Thermotolerance

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alternative splicing of the PECTINESTERASE gene encoding a cell wall-degrading enzyme affects postharvest softening in grape

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Journal of Integrative Agriculture 2024年第3期23卷 863-875页

作者： Hainan Liu Maosong Pei Charles Ampomah-Dwamena Yaxin Shang Yihe Yu Tonglu Wei Qiaofang Shi Dalong Guo College of Horticulture and Plant Protection Henan University of Science and TechnologyLuoyang 471000China Henan Engineering Technology Research Center of Quality Regulation and Control of Horticultural Plants Luoyang 471000China The New Zealand Institute for Plant&Food Research Limited(PFR) Auckland 1010New Zealand

The firmness of table grape berries is a crucial quality parameter. Despite extensive research on postharvest fruit softening, its precise molecular mechanisms remain elusive. To enhance our comprehension of the underlying molecular factors, we initially identified differentially expressed genes(DEGs) by comparing the transcriptomes of folic acid(FA)-treated and water-treated(CK) berries at different time points. We then analyzed the sequences to detect alternatively spliced(AS) genes associated with postharvest softening. A total of 2,559 DEGs were identified and categorized into four subclusters based on their expression patterns, with subcluster-4 genes exhibiting higher expression in the CK group compared with the FA treatment group. There were 1,045 AS-associated genes specific to FA-treated berries and 1,042 in the CK-treated berries, respectively. Gene Ontology(GO) annotation indicated that the AS-associated genes in CK-treated berries were predominantly enriched in cell wall metabolic processes,particularly cell wall degradation processes. Through a comparison between treatment-associated AS genes and subcluster-4 DEGs, we identified eight genes, including Pectinesterase 2(VvPE2, Vitvi15g00704), which encodes a cell wall-degrading enzyme and was predicted to undergo an A3SS event. The reverse transcription polymerase chain reaction further confirmed the presence of a truncated transcript variant of VvPE2 in the FA-treated berries.Our study provides a comprehensive analysis of AS events in postharvest grape berries using transcriptome sequencing and underscores the pivotal role of VvPE2 during the postharvest storage of grape berries.

关键词： grape postharvest softening folic acid alternative splicing Pectinesterase 2 alternative 3'splice site(A3SS)

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Temporal regulation of alternative splicing events in rice memory under drought stress

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Plant Diversity 2022年第1期44卷 116-125页

作者： Hong Yang Ping Li Guihua Jin Daping Gui Li Liu Chengjun Zhang Germplasm Bank of Wild Species Kunming Institute of BotanyChinese Academy of SciencesKunmingYunnan650201China State Key Laboratory of Biocatalysis and Enzyme Engineering Hubei Collaborative Innovation Center for Green Transformation of Bio-ResourcesHubei Key Laboratory of Industrial BiotechnologySchool of Life SciencesHubei UniversityWuhan430062China Haiyan Engineering&Technology Center Kunming Institute of BotanyChinese Academy of ScienceKunmingYunnan650201China University of Chinese Academy of Sciences Beijing100049China

Plant adaptation to drought stress is essential for plant survival and crop yield.Recently,harnessing drought memory,which is induced by repeated stress and recovery cycles,was suggested as a means to improve drought resistance at the transcriptional level.However,the genetic mechanism underlying drought memory is unclear.Here,we carried out a quantitative analysis of alternative splicing(AS)events in rice memory under drought stress,generating 12 transcriptome datasets.Notably,we identified exon skipping(ES)as the predominant AS type(>80%)in differential alternative splicing(DAS)in response to drought stress.Applying our analysis pipeline to investigate DAS events following drought stress in six other plant species revealed variable ES frequencies ranging from 9.94%to 60.70%depending on the species,suggesting that the relative frequency of DAS types in plants is likely to be speciesspecific.The dinucleotide sequence at AS splice sites in rice following drought stress was preferentially GC-AG and AT-AC.Since U12-type splicing uses the AFAC site,this suggests that drought stress may increase U12-type splicing,and thus increase ES frequency.We hypothesize that multiple isofbrms derived from exon skipping may be induced by drought stress in rice.We also identified 20 transcription factors and three highly connected hub genes with potential roles in drought memory that may be good targets for plant breeding.

关键词： alternative splicing Drought memory Exon skipping

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