Dear Editor,In plants, the floral transition is flexibly controlled by various environmental conditions and endogenous developmental cues. In Arabidopsis, six major flowering pathways respond to changes in these facto...Dear Editor,In plants, the floral transition is flexibly controlled by various environmental conditions and endogenous developmental cues. In Arabidopsis, six major flowering pathways respond to changes in these factors （Fornara et al., 2010）. The photoperiod, vernalization, and ambient pathways monitor exogenous signals from the environment such as day length, minimum winter temperature, and ambient temperature （Fornara et al., 2010）. By contrast, the autonomous, gibberellin, and age pathways respond to endogenous cues linked to developmental status （Fornara et al., 2010）. Accumulating evidence indicates that the six flowering pathways converge in a network to regulate floral integrator genes FLOWERING LOCUS T （FT）, TWIN SISTER OF FT （TSF）, and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1 （Fornara et al., 2010）.展开更多
Sesame is a labor intensive crop with limited mechanized harvesting mainly due to the seed shattering(SS)trait.In this study,we performed a genetic analysis of the seed-shattering resistance(SR)trait with a SR sesame ...Sesame is a labor intensive crop with limited mechanized harvesting mainly due to the seed shattering(SS)trait.In this study,we performed a genetic analysis of the seed-shattering resistance(SR)trait with a SR sesame mutant 12M07.Unlike the SS type,the parenchyma cells in the abscission zone of the 12M07 mutant are arranged loosely but adhere to the seed coat.Inheritance analysis of six generations derived from 12M07(SR)×Xiangcheng Dazibai(SS)showed that the SR trait is recessive and controlled by a single gene pair.Association mapping of the F2population with 888,619 variants(single-nucleotide polymorphisms(SNPs)and insertion-deletion(InDels))and 31,884 structural variations(SVs)determined that only SV12002 in the 5′upstream region of gene Sindi0765000(named SiHEC3)in Chr.3 was significantly associated with the SR trait.SiHEC3 encodes the bHLH transcription factor.A 1,049 bp deletion occurred in the 5′UTR of Sihec3 in 12M07.SiHEC3 is mainly expressed in developing placental tissues,with the expression peaking in capsules at 45 days after pollination.A dual-luciferase reporter assay in tobacco confirmed that the promoter activity of Sihec3 was reduced because of the deletion of the 1,049 bp promoter sequence.Protein–protein interaction network analysis showed that HEC3 is co-expressed with nine key proteins,such as SHATTERPROOF1(SHP1)and SEEDSTICK(STK)which participate in the secondary wall biosynthesis of the abscission layer in plants.The findings of this study show the important function of Sihec3corresponding with the SR trait and supply the genetic information for breeding new varieties that are amenable to mechanized harvesting in sesame and other crops.展开更多
Iron is an essential element for plant growth and development. Iron homeostasis in plants is tightly regulated at both transcriptional and posttranscriptional level. Several bHLH transcription factors involved in iron...Iron is an essential element for plant growth and development. Iron homeostasis in plants is tightly regulated at both transcriptional and posttranscriptional level. Several bHLH transcription factors involved in iron homeostasis have been identified recently. However, their regulatory mechanisms remain unknown. In this work, we demonstrate that the transcription factor FIT interacted with AtbHLH38 and AtbHLH39 and directly conferred the expression regulation of iron uptake genes for iron homeostasis in Arabidopsis. Yeast two-hybrid analysis and transient expression in Arabidopsis protoplasts showed that AtbHLH38 or AtbHLH39 interacted with FIT, a central transcription factor involved in iron homeostasis in Arabidopsis. Expression of FIT/AtbHLH38 or FIT/AtbHLH39 in yeast cells activated GUS expression driven by ferric chelate reductase （FRO2） and ferrous transporter （IRT1） promoters. Overexpression of FITwith either AtbHLH38 or AtbHLH39 in plants converted the expression of the iron uptake genes FRO2 and IRT1 from induced to constitutive. Further analysis revealed that FRO2 and IRT1 were not regulated at the posttranscriptional level in these plants because IRT1 protein accumulation and high ferric chelate reductase activity were detected in the overexpression plants under both iron deficiency and iron sufficiency. The double overexpression plants accumulated more iron in their shoots than wild type or the plants overexpressing either AtbHLH38, AtbHLH39 or FIT. Our data support that ferric-chelate reductase FRO2 and ferrous-transporter IRT1 are the targets of the three transcription factors and the transcription of FRO2 and IRT1 is directly regulated by a complex of FIT/AtbHLH38 or FIT/AtbHLH39.展开更多
The Betula genus contains pentacyclic triterpenoid betulin known for its environmental adaptation and medicinal properties.However,the mechanisms underlying betulin biosynthesis responding to climate change remain unc...The Betula genus contains pentacyclic triterpenoid betulin known for its environmental adaptation and medicinal properties.However,the mechanisms underlying betulin biosynthesis responding to climate change remain unclear.In this study,the role of epigenetic modification(DNA methylation) in betulin biosynthesis was examined and how climatic factors influence it.Whole-genome bisulfite sequencing was performed for greenhouse-grown Chinese white birch(Betula platyphylla Sukaczev) treated with DNA methylation inhibitor zebularine(ZEB) and a natural birch population in Northeast China.ZEB treatment significantly affected the CHH methylation level of transposable elements and betulin content in a hormesis dose-dependent manner.The methylation and expression of bHLH9,a key transcriptional factor controlling betulin biosynthesis,were also consistently affected by ZEB treatment as a hormetic dose-response.In the natural population,there was a positive correlation between promoter methylation of bHLH9 and summer precipitation,while winter temperature was negatively correlated.Thus climate-dependent methylation of bHLH9 regulates the expression of downstream genes involved in betulin biosynthesis.This study highlights the role of environmental signals to induce epigenetic changes that result in betulin production,possibly helping to develop resilient plants to combat ongoing climate change and enhance secondary metabolite production.展开更多
Chilling-induced accumulation of reactive oxygen species(ROS) is harmful to plants,which usually produce anthocyanins to scavenge ROS as protection from chilling stress.As a tropical crop,cassava is hypersensitive to ...Chilling-induced accumulation of reactive oxygen species(ROS) is harmful to plants,which usually produce anthocyanins to scavenge ROS as protection from chilling stress.As a tropical crop,cassava is hypersensitive to chilling,but the biochemical basis of this hypersensitivity remains unclear.We previously generated Me MYB2-RNAi transgenic cassava with increased chilling tolerance.Here we report that Me MYB2-RNAi transgenic cassava accumulated less ROS but more cyanidin-3-O-glucoside than the wild type under early chilling stress.Under this stress,the anthocyanin biosynthesis pathway was more active in Me MYB2-RNAi lines than in the wild type,and several genes involved in the pathway,including Me TT8,were up-regulated by Me MYB2-RNAi in the transgenic cassava.Me MYB2 bound to the Me TT8 promoter and blocked its expression under both normal and chilling conditions,thereby inhibiting anthocyanin accumulation.Me TT8 was shown to bind to the promoter of Dihydroflavonol 4-reductase(Me DFR-2)and increased Me DFR-2 expression.Me MYB2 appears to act as an inhibitor of chilling-induced anthocyanin accumulation during the rapid response of cassava to chilling stress.展开更多
Anthocyanins are secondary metabolites induced by environmental stimuli and developmental signals.The positive regulators of anthocyanin biosynthesis have been reported,whereas the anthocyanin repressors have been neg...Anthocyanins are secondary metabolites induced by environmental stimuli and developmental signals.The positive regulators of anthocyanin biosynthesis have been reported,whereas the anthocyanin repressors have been neglected.Although the signal transduction pathways of gibberellin(GA)and jasmonic acid(JA)and their regulation of anthocyanin biosynthesis have been investigated,the cross-talk between GA and JA and the antagonistic mechanism of regulating anthocyanin biosynthesis remain to be investigated.In this study,we identified the anthocyanin repressor MdbHLH162 in apple and revealed its molecular mechanism of regulating anthocyanin biosynthesis by integrating the GA and JA signals.MdbHLH162 exerted passive repression by interacting with MdbHLH3 and MdbHLH33,which are two recognized positive regulators of anthocyanin biosynthesis.MdbHLH162 negatively regulated anthocyanin biosynthesis by disrupting the formation of the anthocyanin-activated MdMYB1-MdbHLH3/33complexes and weakening transcriptional activation of the anthocyanin biosynthetic genes MdDFR and MdUF3GT by MdbHLH3 and MdbHLH33.The GA repressor MdRGL2a antagonized MdbHLH162-mediated inhibition of anthocyanins by sequestering MdbHLH162 from the MdbHLH162-MdbHLH3/33 complex.The JA repressors MdJAZ1 and MdJAZ2 interfered with the antagonistic regulation of MdbHLH162 by MdRGL2a by titrating the formation of the MdRGL2a-MdbHLH162 complex.Our findings reveal that MdbHLH162 integrates the GA and JA signals to negatively regulate anthocyanin biosynthesis.This study provides new information for discovering more anthocyanin biosynthesis repressors and explores the cross-talk between hormone signals.展开更多
Ginseng(Panax ginseng C.A. Meyer) is one of the best-selling herbal medicines, with ginsenosides as its main pharmacologically active constituents. Although extensive chemical and pharmaceutical studies of these compo...Ginseng(Panax ginseng C.A. Meyer) is one of the best-selling herbal medicines, with ginsenosides as its main pharmacologically active constituents. Although extensive chemical and pharmaceutical studies of these compounds have been performed, genome-wide studies of the basic helix-loop-helix(b HLH) transcription factors of ginseng are still limited. The b HLH transcription factor family is one of the largest transcription factor families found in eukaryotic organisms, and these proteins are involved in a myriad of regulatory processes. In our study, 169 bHLH transcription factor genes were identified in the genome of P. ginseng, and phylogenetic analysis indicated that these PGb HLHs could be classified into 24 subfamilies. A total of 21 RNA-seq data sets, including two sequencing libraries for jasmonate(JA)-responsive and 19 reported libraries for organ-specific expression analyses were constructed. Through a combination of gene-specific expression patterns and chemical contents,6 PGbHLH genes from 4 subfamilies were revealed to be potentially involved in the regulation of ginsenoside biosynthesis. These 6 PGbHLHs, which had distinct target genes, were further divided into two groups depending on the absence of MYC-N structure. Our results would provide a foundation for understanding the molecular basis and regulatory mechanisms of bHLH transcription factor action in P.ginseng.展开更多
A subfamily of four Phytochrome （phy）-Interacting bHLH transcription Factors （PIFs） collectively promote skotomorphogenic development in dark-grown seedlings. This activity is reversed upon exposure to light, by p...A subfamily of four Phytochrome （phy）-Interacting bHLH transcription Factors （PIFs） collectively promote skotomorphogenic development in dark-grown seedlings. This activity is reversed upon exposure to light, by photoacti- vated phy molecules that induce degradation of the PIFs, thereby triggering the transcriptional changes that drive a tran- sition to photomorphogenesis. The PIFs function both redundantly and partially differentially at the morphogenic level in this process, To identify the direct targets of PIF transcriptional regulation genome-wide, we analyzed the DNA-binding sites for all four PIFs by ChlP-seq analysis, and defined the genes transcriptionally regulated by each PIF, using RNA-seq analysis of pif mutants. Despite the absence of detectable differences in DNA-binding-motif recognition between the PIFs, the data show a spectrum of regulatory patterns, ranging from single PIF dominance to equal contributions by all four. Similarly, a broad array of promoter architectures was found, ranging from single PIF-binding sites, containing single sequence motifs, through multiple PIF-binding sites, each containing one or more motifs, with each site occupied prefer- entially by one to multiple PIFs. Quantitative analysis of the promoter occupancy and expression level induced by each PIF revealed an intriguing pattern. Although there is no robust correlation broadly across the target-gene population, examination of individual genes that are shared targets of multiple PIFs shows a gradation in correlation from strongly positive, through uncorrelated, to negative. This finding suggests a dual-layered mechanism of transcriptional regulation, comprising both a continuum of binding-site occupancy by each PIF and a superimposed layer of local regulation that acts differentially on each PIF, to modulate its intrinsic transcriptional activation capacity at each site, in a quantitative pattern that varies between the individual PIFs from gene to gene. These findings provide a framework for probing the mecha- nisms by which transcription factors with overlapping direct-target genes integrate and selectively transduce signals to their target networks.展开更多
A basic helix-loop-helix transcription factor,figα,is one of the earliest marker genes of oocyte differentiation in vertebrates.In the present study,we made figαknockout medaka by CRISPR/Cas9,expecting aborted progr...A basic helix-loop-helix transcription factor,figα,is one of the earliest marker genes of oocyte differentiation in vertebrates.In the present study,we made figαknockout medaka by CRISPR/Cas9,expecting aborted progress of oogenesis,to see if differentiation of somatic ovarian tissues is affected.Figαknockout male gonads differentiated normally into testes with functional sperm.The females,on the other hand,were sterile;there are oocytes only up to pachytene.No growing oocytes in diplotene were found.The phenotype was already apparent at 10 days after hatching,when diplotene oocytes start to develop in the control.Furthermore,several putative target genes of figαwere not expressed in the mutant female gonads.Previous studies showed that medaka lacking germ cells have morphologically abnormal gonads and female to male sex reversal occurred.Figαknockout female gonads differentiated morphologically into ovaries,suggesting that in medaka,figαknockout prevents oogenesis progress into diplotene stage,and ovarian differentiation do not need developed oocytes beyond pachytene stage.This ovarian phenotype reminded us of"pachytene checkpoint",which blocks meiotic progress into diplotene when chromosomal abnormalities are present.Therefore,we further analyzed phenotype of oocytes in the mutant ovaries.The number of oocytes at each meiotic stage suggested that oogenesis normally proceeded up to pachytene stage.The number of apoptotic oocytes in the knockout ovaries was small.Finally,we did not detect any abnormal pairings or unrepaired double-strand breaks by immunostaining.Taken together,figαknockout likely stop oocyte growth at or around pachytene via unknown mechanisms other than"pachytene checkpoint"and arrested oocytes remained in the ovary.This is in contrast to figαknockout mice where all oocytes underwent apoptosis quickly after birth.展开更多
文摘Dear Editor,In plants, the floral transition is flexibly controlled by various environmental conditions and endogenous developmental cues. In Arabidopsis, six major flowering pathways respond to changes in these factors （Fornara et al., 2010）. The photoperiod, vernalization, and ambient pathways monitor exogenous signals from the environment such as day length, minimum winter temperature, and ambient temperature （Fornara et al., 2010）. By contrast, the autonomous, gibberellin, and age pathways respond to endogenous cues linked to developmental status （Fornara et al., 2010）. Accumulating evidence indicates that the six flowering pathways converge in a network to regulate floral integrator genes FLOWERING LOCUS T （FT）, TWIN SISTER OF FT （TSF）, and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1 （Fornara et al., 2010）.
基金financially supported by the earmarked funding for the China Agriculture Research System of MOF and MARA(CARS-14)the Key Project of Science and Technology of Henan Province,China(201300110600)+6 种基金the Henan Province Specific Professor Position Program,China(SPPP2022)the Zhongyuan Scientist Workshop Construction,China(ZSWC2019 and 214400510026)the Innovation Scientist and Technician Troop Construction Project of Henan Province,China(ISTTCPHP2016)the Shennong Laboratory First Class Program,China(SN01-2022-04)the Key Research and Development Project of Henan Province,China(221111520400)the Innovation Scientists and Technicians Troop Construction Project of Henan Academy of Agricultural Sciences,China(2023TD04)the Henan Province Science and Technology Research and Development Plan Joint Fund,China(232301420108)。
文摘Sesame is a labor intensive crop with limited mechanized harvesting mainly due to the seed shattering(SS)trait.In this study,we performed a genetic analysis of the seed-shattering resistance(SR)trait with a SR sesame mutant 12M07.Unlike the SS type,the parenchyma cells in the abscission zone of the 12M07 mutant are arranged loosely but adhere to the seed coat.Inheritance analysis of six generations derived from 12M07(SR)×Xiangcheng Dazibai(SS)showed that the SR trait is recessive and controlled by a single gene pair.Association mapping of the F2population with 888,619 variants(single-nucleotide polymorphisms(SNPs)and insertion-deletion(InDels))and 31,884 structural variations(SVs)determined that only SV12002 in the 5′upstream region of gene Sindi0765000(named SiHEC3)in Chr.3 was significantly associated with the SR trait.SiHEC3 encodes the bHLH transcription factor.A 1,049 bp deletion occurred in the 5′UTR of Sihec3 in 12M07.SiHEC3 is mainly expressed in developing placental tissues,with the expression peaking in capsules at 45 days after pollination.A dual-luciferase reporter assay in tobacco confirmed that the promoter activity of Sihec3 was reduced because of the deletion of the 1,049 bp promoter sequence.Protein–protein interaction network analysis showed that HEC3 is co-expressed with nine key proteins,such as SHATTERPROOF1(SHP1)and SEEDSTICK(STK)which participate in the secondary wall biosynthesis of the abscission layer in plants.The findings of this study show the important function of Sihec3corresponding with the SR trait and supply the genetic information for breeding new varieties that are amenable to mechanized harvesting in sesame and other crops.
基金The authors thank ProfMary Lou Guerinot （Department of Biological Sciences, Dartmouth College, Hanover, New Hampshire） for providing IRT1 peptide antibody and for the critical reading of the manuscript. We are also grateful to Drs Zhentao Lin and Yongfu Fu （Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing） for providing the BiFC assay system and technical supporting. This work was supported by the National Natural Science Foundation of China （Grant nos, 30530460 and 30521001） and the Ministry of Science and Technology of China （Grant nos, 2005cb20904 and 2006AA 10A 105） and Chinese Academy of Sciences （Grant no. KSCX2-YW-N- 001） as well as by the Harvest Plus-China Program.
文摘Iron is an essential element for plant growth and development. Iron homeostasis in plants is tightly regulated at both transcriptional and posttranscriptional level. Several bHLH transcription factors involved in iron homeostasis have been identified recently. However, their regulatory mechanisms remain unknown. In this work, we demonstrate that the transcription factor FIT interacted with AtbHLH38 and AtbHLH39 and directly conferred the expression regulation of iron uptake genes for iron homeostasis in Arabidopsis. Yeast two-hybrid analysis and transient expression in Arabidopsis protoplasts showed that AtbHLH38 or AtbHLH39 interacted with FIT, a central transcription factor involved in iron homeostasis in Arabidopsis. Expression of FIT/AtbHLH38 or FIT/AtbHLH39 in yeast cells activated GUS expression driven by ferric chelate reductase （FRO2） and ferrous transporter （IRT1） promoters. Overexpression of FITwith either AtbHLH38 or AtbHLH39 in plants converted the expression of the iron uptake genes FRO2 and IRT1 from induced to constitutive. Further analysis revealed that FRO2 and IRT1 were not regulated at the posttranscriptional level in these plants because IRT1 protein accumulation and high ferric chelate reductase activity were detected in the overexpression plants under both iron deficiency and iron sufficiency. The double overexpression plants accumulated more iron in their shoots than wild type or the plants overexpressing either AtbHLH38, AtbHLH39 or FIT. Our data support that ferric-chelate reductase FRO2 and ferrous-transporter IRT1 are the targets of the three transcription factors and the transcription of FRO2 and IRT1 is directly regulated by a complex of FIT/AtbHLH38 or FIT/AtbHLH39.
基金the National Non-profi t Institute Research Grant of the Chinese Academy of Forestry(CAFYBB2019ZY003)the National Natural Science Foundation of China(31871220 and 31801444)+2 种基金the Innovation Project of State Key Laboratory of Tree Genetics andBreeding(Northeast Forestry University)(2013A06)the Fundamental Research Funds for the Central Universities(2572017DA06 and 2572020DP01)Heilongjiang Provincial Natural Science Foundation of China(LH2021C005).
文摘The Betula genus contains pentacyclic triterpenoid betulin known for its environmental adaptation and medicinal properties.However,the mechanisms underlying betulin biosynthesis responding to climate change remain unclear.In this study,the role of epigenetic modification(DNA methylation) in betulin biosynthesis was examined and how climatic factors influence it.Whole-genome bisulfite sequencing was performed for greenhouse-grown Chinese white birch(Betula platyphylla Sukaczev) treated with DNA methylation inhibitor zebularine(ZEB) and a natural birch population in Northeast China.ZEB treatment significantly affected the CHH methylation level of transposable elements and betulin content in a hormesis dose-dependent manner.The methylation and expression of bHLH9,a key transcriptional factor controlling betulin biosynthesis,were also consistently affected by ZEB treatment as a hormetic dose-response.In the natural population,there was a positive correlation between promoter methylation of bHLH9 and summer precipitation,while winter temperature was negatively correlated.Thus climate-dependent methylation of bHLH9 regulates the expression of downstream genes involved in betulin biosynthesis.This study highlights the role of environmental signals to induce epigenetic changes that result in betulin production,possibly helping to develop resilient plants to combat ongoing climate change and enhance secondary metabolite production.
基金supported by the Hainan Provincial Natural Science Foundation of China (322RC798)the National Key Research and Development Program of China (2018YFD1000501)+1 种基金the Major Science and Technology Plan of Hainan Province (ZDKJ2021012)the China Agriculture Research System (CARS11-HNCX)。
文摘Chilling-induced accumulation of reactive oxygen species(ROS) is harmful to plants,which usually produce anthocyanins to scavenge ROS as protection from chilling stress.As a tropical crop,cassava is hypersensitive to chilling,but the biochemical basis of this hypersensitivity remains unclear.We previously generated Me MYB2-RNAi transgenic cassava with increased chilling tolerance.Here we report that Me MYB2-RNAi transgenic cassava accumulated less ROS but more cyanidin-3-O-glucoside than the wild type under early chilling stress.Under this stress,the anthocyanin biosynthesis pathway was more active in Me MYB2-RNAi lines than in the wild type,and several genes involved in the pathway,including Me TT8,were up-regulated by Me MYB2-RNAi in the transgenic cassava.Me MYB2 bound to the Me TT8 promoter and blocked its expression under both normal and chilling conditions,thereby inhibiting anthocyanin accumulation.Me TT8 was shown to bind to the promoter of Dihydroflavonol 4-reductase(Me DFR-2)and increased Me DFR-2 expression.Me MYB2 appears to act as an inhibitor of chilling-induced anthocyanin accumulation during the rapid response of cassava to chilling stress.
基金financially supported by grants from the Natural Science Foundation of Shandong Province(ZR2022YQ24)the Development Plan of the Youth Innovation Team of the Higher Education Institutions in Shandong Province(2022KJ326)Wuhan Botanical Garden Scientific Research Support Project(E3559901)。
文摘Anthocyanins are secondary metabolites induced by environmental stimuli and developmental signals.The positive regulators of anthocyanin biosynthesis have been reported,whereas the anthocyanin repressors have been neglected.Although the signal transduction pathways of gibberellin(GA)and jasmonic acid(JA)and their regulation of anthocyanin biosynthesis have been investigated,the cross-talk between GA and JA and the antagonistic mechanism of regulating anthocyanin biosynthesis remain to be investigated.In this study,we identified the anthocyanin repressor MdbHLH162 in apple and revealed its molecular mechanism of regulating anthocyanin biosynthesis by integrating the GA and JA signals.MdbHLH162 exerted passive repression by interacting with MdbHLH3 and MdbHLH33,which are two recognized positive regulators of anthocyanin biosynthesis.MdbHLH162 negatively regulated anthocyanin biosynthesis by disrupting the formation of the anthocyanin-activated MdMYB1-MdbHLH3/33complexes and weakening transcriptional activation of the anthocyanin biosynthetic genes MdDFR and MdUF3GT by MdbHLH3 and MdbHLH33.The GA repressor MdRGL2a antagonized MdbHLH162-mediated inhibition of anthocyanins by sequestering MdbHLH162 from the MdbHLH162-MdbHLH3/33 complex.The JA repressors MdJAZ1 and MdJAZ2 interfered with the antagonistic regulation of MdbHLH162 by MdRGL2a by titrating the formation of the MdRGL2a-MdbHLH162 complex.Our findings reveal that MdbHLH162 integrates the GA and JA signals to negatively regulate anthocyanin biosynthesis.This study provides new information for discovering more anthocyanin biosynthesis repressors and explores the cross-talk between hormone signals.
文摘Ginseng(Panax ginseng C.A. Meyer) is one of the best-selling herbal medicines, with ginsenosides as its main pharmacologically active constituents. Although extensive chemical and pharmaceutical studies of these compounds have been performed, genome-wide studies of the basic helix-loop-helix(b HLH) transcription factors of ginseng are still limited. The b HLH transcription factor family is one of the largest transcription factor families found in eukaryotic organisms, and these proteins are involved in a myriad of regulatory processes. In our study, 169 bHLH transcription factor genes were identified in the genome of P. ginseng, and phylogenetic analysis indicated that these PGb HLHs could be classified into 24 subfamilies. A total of 21 RNA-seq data sets, including two sequencing libraries for jasmonate(JA)-responsive and 19 reported libraries for organ-specific expression analyses were constructed. Through a combination of gene-specific expression patterns and chemical contents,6 PGbHLH genes from 4 subfamilies were revealed to be potentially involved in the regulation of ginsenoside biosynthesis. These 6 PGbHLHs, which had distinct target genes, were further divided into two groups depending on the absence of MYC-N structure. Our results would provide a foundation for understanding the molecular basis and regulatory mechanisms of bHLH transcription factor action in P.ginseng.
文摘A subfamily of four Phytochrome （phy）-Interacting bHLH transcription Factors （PIFs） collectively promote skotomorphogenic development in dark-grown seedlings. This activity is reversed upon exposure to light, by photoacti- vated phy molecules that induce degradation of the PIFs, thereby triggering the transcriptional changes that drive a tran- sition to photomorphogenesis. The PIFs function both redundantly and partially differentially at the morphogenic level in this process, To identify the direct targets of PIF transcriptional regulation genome-wide, we analyzed the DNA-binding sites for all four PIFs by ChlP-seq analysis, and defined the genes transcriptionally regulated by each PIF, using RNA-seq analysis of pif mutants. Despite the absence of detectable differences in DNA-binding-motif recognition between the PIFs, the data show a spectrum of regulatory patterns, ranging from single PIF dominance to equal contributions by all four. Similarly, a broad array of promoter architectures was found, ranging from single PIF-binding sites, containing single sequence motifs, through multiple PIF-binding sites, each containing one or more motifs, with each site occupied prefer- entially by one to multiple PIFs. Quantitative analysis of the promoter occupancy and expression level induced by each PIF revealed an intriguing pattern. Although there is no robust correlation broadly across the target-gene population, examination of individual genes that are shared targets of multiple PIFs shows a gradation in correlation from strongly positive, through uncorrelated, to negative. This finding suggests a dual-layered mechanism of transcriptional regulation, comprising both a continuum of binding-site occupancy by each PIF and a superimposed layer of local regulation that acts differentially on each PIF, to modulate its intrinsic transcriptional activation capacity at each site, in a quantitative pattern that varies between the individual PIFs from gene to gene. These findings provide a framework for probing the mecha- nisms by which transcription factors with overlapping direct-target genes integrate and selectively transduce signals to their target networks.
基金supported in part by a Grant-in-Aid for Scientific Research from JSPS(15K07127 to AK and MK).
文摘A basic helix-loop-helix transcription factor,figα,is one of the earliest marker genes of oocyte differentiation in vertebrates.In the present study,we made figαknockout medaka by CRISPR/Cas9,expecting aborted progress of oogenesis,to see if differentiation of somatic ovarian tissues is affected.Figαknockout male gonads differentiated normally into testes with functional sperm.The females,on the other hand,were sterile;there are oocytes only up to pachytene.No growing oocytes in diplotene were found.The phenotype was already apparent at 10 days after hatching,when diplotene oocytes start to develop in the control.Furthermore,several putative target genes of figαwere not expressed in the mutant female gonads.Previous studies showed that medaka lacking germ cells have morphologically abnormal gonads and female to male sex reversal occurred.Figαknockout female gonads differentiated morphologically into ovaries,suggesting that in medaka,figαknockout prevents oogenesis progress into diplotene stage,and ovarian differentiation do not need developed oocytes beyond pachytene stage.This ovarian phenotype reminded us of"pachytene checkpoint",which blocks meiotic progress into diplotene when chromosomal abnormalities are present.Therefore,we further analyzed phenotype of oocytes in the mutant ovaries.The number of oocytes at each meiotic stage suggested that oogenesis normally proceeded up to pachytene stage.The number of apoptotic oocytes in the knockout ovaries was small.Finally,we did not detect any abnormal pairings or unrepaired double-strand breaks by immunostaining.Taken together,figαknockout likely stop oocyte growth at or around pachytene via unknown mechanisms other than"pachytene checkpoint"and arrested oocytes remained in the ovary.This is in contrast to figαknockout mice where all oocytes underwent apoptosis quickly after birth.
