Integration of mechanical properties with the biomolecular network is fundamental in understanding various developmental and resilience signaling in plants.The mechanical properties of the cell wall-plasma membrane-cy...Integration of mechanical properties with the biomolecular network is fundamental in understanding various developmental and resilience signaling in plants.The mechanical properties of the cell wall-plasma membrane-cytoskeleton continuum and interconnected endomembrane system can regulate plant growth signaling and plant-microbiome interactions that unlock new opportunities for enhancing crop yield and defense,thereby promoting sustainable agriculture and food security.展开更多
Arabidopsis plastid antiporters KEA1 and KEA2are critical for plastid development, photosynthetic efficiency, and plant development.Here, we show that KEA1 and KEA2 are involved in vacuolar protein trafficking. Geneti...Arabidopsis plastid antiporters KEA1 and KEA2are critical for plastid development, photosynthetic efficiency, and plant development.Here, we show that KEA1 and KEA2 are involved in vacuolar protein trafficking. Genetic analyses found that the kea1 kea2 mutants had short siliques, small seeds, and short seedlings. Molecular and biochemical assays showed that seed storage proteins were missorted out of the cell and the precursor proteins were accumulated in kea1 kea2. Protein storage vacuoles(PSVs) were smaller in kea1 kea2. Further analyses showed that endosomal trafficking in kea1 kea2 was compromised. Vacuolar sorting receptor 1(VSR1) subcellular localizations, VSR–cargo interactions, and p24 distribution on the endoplasmic reticulum(ER) and Golgi apparatus were affected in kea1 kea2. Moreover, plastid stromule growth was reduced and plastid association with the endomembrane compartments was disrupted in kea1 kea2. Stromule growth was regulated by the cellular pH and K+homeostasis maintained by KEA1 and KEA2. The organellar pH along the trafficking pathway was altered in kea1 kea2. Overall, KEA1 and KEA2 regulate vacuolar trafficking by controlling the function of plastid stromules via adjusting pH and K+homeostasis.展开更多
The pH of intracellular compartments is essential for the viability of cells. Despite its relevance, little is known about the pH of these compartments. To measure pH in vivo, we have first generated two pH sensors by...The pH of intracellular compartments is essential for the viability of cells. Despite its relevance, little is known about the pH of these compartments. To measure pH in vivo, we have first generated two pH sensors by combining the improved-solubility feature of solubility-modified green fluorescent protein （GFP） （smGFP） with the pH-sensing capabil- ity of the pHluorins and codon optimized for expression in Arabidopsis. PEpHluorin （plant-solubility-modified ecliptic pHluorin） gradually loses fluorescence as pH is lowered with fluorescence vanishing at pH 6.2 and PRpHluorin （plant- solubility-modified ratiomatric pHluorin）, a dual-excitation sensor, allowing for precise measurements. Compartment- specific sensors were generated by further fusing specific sorting signals to PEpHluorin and PRpHluorin. Our results show that the pH of cytosol and nucleus is similar （pH 7.3 and 7.2）, while peroxisomes, mitochondrial matrix, and plastidial stroma have alkaline pH. Compartments of the secretory pathway reveal a gradual acidification, spanning from pH 7.1 in the endoplasmic reticulum （ER） to pH 5.2 in the vacuole. Surprisingly, pH in the trans-Golgi network （TGN） and mul- tivesicular body （MVB） is, with pH 6.3 and 6.2, quite similar. The inhibition of vacuolar-type H＋-ATPase （V-ATPase） with concanamycin A （ConcA） caused drastic increase in pH in TGN and vacuole. Overall, the PEpHluorin and PRpHluorin are excellent pH sensors for visualization and quantification of pH in vivo, respectively.展开更多
Plant prevacuolar compartments （PVCs）, or multivesicular bodies （MVBs）, are single membrane-bound organelles that play important roles in mediating protein trafficking to vacuoles in the secretory pathway. PVC/MVB...Plant prevacuolar compartments （PVCs）, or multivesicular bodies （MVBs）, are single membrane-bound organelles that play important roles in mediating protein trafficking to vacuoles in the secretory pathway. PVC/MVB also serves as a late endosome in the endocytic pathway in plants. Since the plant PVC was iden- tified as an MVB more than 10 years ago,-great progress has been made toward the understanding of PVC/ MVB function and biogenesis in plants. In this review, we first summarize previous research into the iden- tification and characterization of plant PVCs/MVBs, and then highlight recent advances on the mechanisms underlying intraluminal vesicle formation and maturation of plant PVCs/MVBs. In addition, we discuss the possible crosstalk that appears to occur between PVCs/MVBs and autophagosomes during autophagy in plants. Finally, we list some open questions and present future perspectives in this field.展开更多
In multicellular and even single-celled organisms,individual components are interconnected at multiscale levels to produce enormously complex biological networks that help these systems maintain homeostasis for develo...In multicellular and even single-celled organisms,individual components are interconnected at multiscale levels to produce enormously complex biological networks that help these systems maintain homeostasis for development and environmental adaptation.Systems biology studies initially adopted network analysis to explore how relationships between individual components give rise to complex biological processes.Network analysis has been applied to dissect the complex connectivity of mammalian brains across different scales in time and space in The Human Brain Project.In plant science,network analysis has similarly been applied to study the connectivity of plant components at the molecular,subcellular,cellular,organic,and organism levels.Analysis of these multiscale networks contributes to our understanding of how genotype determines phenotype.In this review,we summarized the theoretical framework of plant multiscale networks and introduced studies investigating plant networks by various experimental and computational modalities.We next discussed the currently available analytic methodologies and multi-level imaging techniques used to map multiscale networks in plants.Finally,we highlighted some of the technical challenges and key questions remaining to be addressed in this emerging field.展开更多
Eukaryotic cells consist of numerous membrane-bound organelles,which compartmentalize cellular materials to fulfil a variety of vital functions.In the post-genomic era,it is widely recognized that identification of th...Eukaryotic cells consist of numerous membrane-bound organelles,which compartmentalize cellular materials to fulfil a variety of vital functions.In the post-genomic era,it is widely recognized that identification of the subcellular organelle localization and transport mechanisms of the encoded proteins are necessary for a fundamental understanding of their biological functions and theorganization of cellular activity.Multiple experimental approaches are now available to determine the subcellular localizations and dynamics of proteins.In this review,we provide an overview of the current methods and organelle markers for protein subcellular localization and trafficking studies in plants,with a focus on the organelles of the endomembrane system.We also discuss the limitations of each method in terms of protein colocalization studies.展开更多
Membrane trafficking is essential for plant growth and responses to external signals.The plant unique FYVE domain-containing protein FREE1 is a component of the ESCRT complex（endosomal sorting complex required for tr...Membrane trafficking is essential for plant growth and responses to external signals.The plant unique FYVE domain-containing protein FREE1 is a component of the ESCRT complex（endosomal sorting complex required for transport）.FREE1 plays multiple roles in regulating protein trafficking and organelle biogenesis including the formation of intraluminal vesicles of multivesicular body（MVB）,vacuolar protein transport and vacuole biogenesis,and autophagic degradation.FREE1 knockout plants show defective MVB formation,abnormal vacuolar transport,fragmented vacuoles,accumulated autophagosomes,and seedling lethality.To further uncover the underlying mechanisms of FREE1 function in plants,we performed a forward genetic screen for mutants that suppressed the seedling lethal phenotype of FREE1-RNAi transgenic plants.The obtained mutants are termed as suppressors of free1（sof）.To date,229 putative sof mutants have been identified.Barely detecting of FREE1 protein with M3 plants further identified 84 FREE1-related suppressors.Also145 mutants showing no reduction of FREE1 protein were termed as RNAi-related mutants.Through next-generation sequencing（NGS）of bulked DNA from F2 mapping population of two RNAi-related sof mutants,FREE1-RNAi T-DNA inserted on chromosome 1 was identified and the causal mutation of putative sof mutant is being identified similarly.These FREE1-and RNAi-related sof mutants will be useful tools and resources for illustrating the underlying mechanisms of FREE1 function in intracellular trafficking and organelle biogenesis,as well as for uncovering the new components involved in the regulation of silencing pathways in plants.展开更多
Dear Editor Membrane proteins destined for degradation in eukaryotic cells are tagged by ubiquitin （Ub） for further sorting by the endosomat sorting complex required for transport （ESCRT） machinery into the intral...Dear Editor Membrane proteins destined for degradation in eukaryotic cells are tagged by ubiquitin （Ub） for further sorting by the endosomat sorting complex required for transport （ESCRT） machinery into the intraluminal vesicles （tLVs） of multivesicular bodies （MVBs） or prevacuolar compartments （PVCs） for subsequent lysosomal/vacuolar degradation （Henne et al., 2011; Schmidt and Teis, 2012）. The ESCRT machinery comprises a cascade of several distinct complexes （ESCRTs -0, -Ⅰ, -Ⅱ, and -Ⅲ, and Vps4 complex）.展开更多
The roles of Rho family guanosine triphosphatases(GTPases)of plants(ROPs)in modulating plant growth and development have been well characterized.However,little is known about the roles of ROP signaling pathways in reg...The roles of Rho family guanosine triphosphatases(GTPases)of plants(ROPs)in modulating plant growth and development have been well characterized.However,little is known about the roles of ROP signaling pathways in regulating plant autophagy and autophagosome formation.In this study,we identify a unique ROP signaling mechanism,which mediates developmental to autophagic transition under stress conditions in the model plant Arabidopsis.Loss-of-function mutants of ROP8 showed stress-induced hypersensitive phenotypes and compromised autophagic flux.Similar to other ROPs in the ROP/RAC family,ROP8 exhibits both plasma membrane and cytosolic punctate localization patterns.Upon autophagic induction,active ROP8 puncta colocalize with autophagosomal markers and are degraded inside the vacuole.In human cells,RalB,an RAS subfamily GTPase,engages its effector Exo84 for autophagosome assembly.However,a RalB counterpart is missing in the plant lineage.Intriguingly,we discovered that plant ROP8 promotes autophagy via its downstream effector Sec5.Live-cell super-resolution imaging showed that ROP8 and Sec5 reside on phagophores for autophagosome formation.Taken together,our findings highlight a previously unappreciated role of an ROP8-Sec5 signaling axis in autophagy promotion,providing new insights into how plants utilize versatile ROP signaling networks to coordinate developmental and autophagic responses depending on environmental changes.展开更多
基金the Zhejiang A&F University and School of Biological Sciences,Nanyang Technological University.J.S.was supported by the National Natural Science Foundation of China(32170342)the Fundamental Research Funds for the Provincial Universities of Zhejiang(2020KJ001)+1 种基金the Zhejiang A&F University Starting Fundings(2024LFR053).Y.M.was supportedby MOETier 2(MOET2EP30122-0021)Tier 3(MOE2019-T3-1-012)of the National Research Foundation,Singapore(NRF-NRFI08-2022-0012).
文摘Integration of mechanical properties with the biomolecular network is fundamental in understanding various developmental and resilience signaling in plants.The mechanical properties of the cell wall-plasma membrane-cytoskeleton continuum and interconnected endomembrane system can regulate plant growth signaling and plant-microbiome interactions that unlock new opportunities for enhancing crop yield and defense,thereby promoting sustainable agriculture and food security.
基金supported by the National Natural Science Foundation of China (NSFC)(31571464, 31371438, 31070222 to Quan-Sheng Qiu)the National Basic Research Program of China (973)project, 2013CB429904 to Quan-Sheng Qiu)+5 种基金the Research Fund for the Doctoral Program of Higher Education of China(RFDP)(20130211110001 to Quan-Sheng Qiu)Research Team of Stress Tolerance Mechanisms and Molecular Breeding of Plateau PlantsQinghai Province “Kunlun Talents·Advanced Innovative and Entrepreneurial Talents” Program (2022 to QuanSheng Qiu)the Qinghai Provincial Department of Science and Technology Qinghai basic research program (2022-ZJ-724 to Quan-Sheng Qiu)the Independent Research and Development Project of State Key Laboratory of Herbage Improvement and Grassland Agro-ecosystems (202202 to Quan-Sheng Qiu)the Core Facility of School of Life Sciences,Lanzhou University。
文摘Arabidopsis plastid antiporters KEA1 and KEA2are critical for plastid development, photosynthetic efficiency, and plant development.Here, we show that KEA1 and KEA2 are involved in vacuolar protein trafficking. Genetic analyses found that the kea1 kea2 mutants had short siliques, small seeds, and short seedlings. Molecular and biochemical assays showed that seed storage proteins were missorted out of the cell and the precursor proteins were accumulated in kea1 kea2. Protein storage vacuoles(PSVs) were smaller in kea1 kea2. Further analyses showed that endosomal trafficking in kea1 kea2 was compromised. Vacuolar sorting receptor 1(VSR1) subcellular localizations, VSR–cargo interactions, and p24 distribution on the endoplasmic reticulum(ER) and Golgi apparatus were affected in kea1 kea2. Moreover, plastid stromule growth was reduced and plastid association with the endomembrane compartments was disrupted in kea1 kea2. Stromule growth was regulated by the cellular pH and K+homeostasis maintained by KEA1 and KEA2. The organellar pH along the trafficking pathway was altered in kea1 kea2. Overall, KEA1 and KEA2 regulate vacuolar trafficking by controlling the function of plastid stromules via adjusting pH and K+homeostasis.
文摘The pH of intracellular compartments is essential for the viability of cells. Despite its relevance, little is known about the pH of these compartments. To measure pH in vivo, we have first generated two pH sensors by combining the improved-solubility feature of solubility-modified green fluorescent protein （GFP） （smGFP） with the pH-sensing capabil- ity of the pHluorins and codon optimized for expression in Arabidopsis. PEpHluorin （plant-solubility-modified ecliptic pHluorin） gradually loses fluorescence as pH is lowered with fluorescence vanishing at pH 6.2 and PRpHluorin （plant- solubility-modified ratiomatric pHluorin）, a dual-excitation sensor, allowing for precise measurements. Compartment- specific sensors were generated by further fusing specific sorting signals to PEpHluorin and PRpHluorin. Our results show that the pH of cytosol and nucleus is similar （pH 7.3 and 7.2）, while peroxisomes, mitochondrial matrix, and plastidial stroma have alkaline pH. Compartments of the secretory pathway reveal a gradual acidification, spanning from pH 7.1 in the endoplasmic reticulum （ER） to pH 5.2 in the vacuole. Surprisingly, pH in the trans-Golgi network （TGN） and mul- tivesicular body （MVB） is, with pH 6.3 and 6.2, quite similar. The inhibition of vacuolar-type H＋-ATPase （V-ATPase） with concanamycin A （ConcA） caused drastic increase in pH in TGN and vacuole. Overall, the PEpHluorin and PRpHluorin are excellent pH sensors for visualization and quantification of pH in vivo, respectively.
文摘Plant prevacuolar compartments （PVCs）, or multivesicular bodies （MVBs）, are single membrane-bound organelles that play important roles in mediating protein trafficking to vacuoles in the secretory pathway. PVC/MVB also serves as a late endosome in the endocytic pathway in plants. Since the plant PVC was iden- tified as an MVB more than 10 years ago,-great progress has been made toward the understanding of PVC/ MVB function and biogenesis in plants. In this review, we first summarize previous research into the iden- tification and characterization of plant PVCs/MVBs, and then highlight recent advances on the mechanisms underlying intraluminal vesicle formation and maturation of plant PVCs/MVBs. In addition, we discuss the possible crosstalk that appears to occur between PVCs/MVBs and autophagosomes during autophagy in plants. Finally, we list some open questions and present future perspectives in this field.
基金supported by the National Natural Science Foundation of China(31530084,32000558,32000483,and31800504)the Programme of Introducing Talents of Discipline to Universities(111 project,B13007)the China Postdoctoral Science Foundation Grant(2019M660494)。
文摘In multicellular and even single-celled organisms,individual components are interconnected at multiscale levels to produce enormously complex biological networks that help these systems maintain homeostasis for development and environmental adaptation.Systems biology studies initially adopted network analysis to explore how relationships between individual components give rise to complex biological processes.Network analysis has been applied to dissect the complex connectivity of mammalian brains across different scales in time and space in The Human Brain Project.In plant science,network analysis has similarly been applied to study the connectivity of plant components at the molecular,subcellular,cellular,organic,and organism levels.Analysis of these multiscale networks contributes to our understanding of how genotype determines phenotype.In this review,we summarized the theoretical framework of plant multiscale networks and introduced studies investigating plant networks by various experimental and computational modalities.We next discussed the currently available analytic methodologies and multi-level imaging techniques used to map multiscale networks in plants.Finally,we highlighted some of the technical challenges and key questions remaining to be addressed in this emerging field.
基金This work was supported by the National Natural Science Foundation of China(31970181)the Zhejiang Provincial Natural Science Foundation of China(R20C020001)+1 种基金the National Key Research and Development Program of China(2018YFD1000604)the Zhejiang Agricultural and Forestry University Starting Funding(2018FR029).
文摘Eukaryotic cells consist of numerous membrane-bound organelles,which compartmentalize cellular materials to fulfil a variety of vital functions.In the post-genomic era,it is widely recognized that identification of the subcellular organelle localization and transport mechanisms of the encoded proteins are necessary for a fundamental understanding of their biological functions and theorganization of cellular activity.Multiple experimental approaches are now available to determine the subcellular localizations and dynamics of proteins.In this review,we provide an overview of the current methods and organelle markers for protein subcellular localization and trafficking studies in plants,with a focus on the organelles of the endomembrane system.We also discuss the limitations of each method in terms of protein colocalization studies.
基金supported by grants from the NIH GM114660 to Y.Zhaothe Research Grants Council of Hong Kong (CUHK466011,465112,466613,CUHK2/CRF/11G,C401114R and Ao E/M-05/12)+2 种基金NSFC/RGC (N_CUHK406/12)NSFC (31270226 and 31470294)Shenzhen Peacock Project (KQTD201101) to L.Jiang
文摘Membrane trafficking is essential for plant growth and responses to external signals.The plant unique FYVE domain-containing protein FREE1 is a component of the ESCRT complex（endosomal sorting complex required for transport）.FREE1 plays multiple roles in regulating protein trafficking and organelle biogenesis including the formation of intraluminal vesicles of multivesicular body（MVB）,vacuolar protein transport and vacuole biogenesis,and autophagic degradation.FREE1 knockout plants show defective MVB formation,abnormal vacuolar transport,fragmented vacuoles,accumulated autophagosomes,and seedling lethality.To further uncover the underlying mechanisms of FREE1 function in plants,we performed a forward genetic screen for mutants that suppressed the seedling lethal phenotype of FREE1-RNAi transgenic plants.The obtained mutants are termed as suppressors of free1（sof）.To date,229 putative sof mutants have been identified.Barely detecting of FREE1 protein with M3 plants further identified 84 FREE1-related suppressors.Also145 mutants showing no reduction of FREE1 protein were termed as RNAi-related mutants.Through next-generation sequencing（NGS）of bulked DNA from F2 mapping population of two RNAi-related sof mutants,FREE1-RNAi T-DNA inserted on chromosome 1 was identified and the causal mutation of putative sof mutant is being identified similarly.These FREE1-and RNAi-related sof mutants will be useful tools and resources for illustrating the underlying mechanisms of FREE1 function in intracellular trafficking and organelle biogenesis,as well as for uncovering the new components involved in the regulation of silencing pathways in plants.
文摘Dear Editor Membrane proteins destined for degradation in eukaryotic cells are tagged by ubiquitin （Ub） for further sorting by the endosomat sorting complex required for transport （ESCRT） machinery into the intraluminal vesicles （tLVs） of multivesicular bodies （MVBs） or prevacuolar compartments （PVCs） for subsequent lysosomal/vacuolar degradation （Henne et al., 2011; Schmidt and Teis, 2012）. The ESCRT machinery comprises a cascade of several distinct complexes （ESCRTs -0, -Ⅰ, -Ⅱ, and -Ⅲ, and Vps4 complex）.
基金supported by grants from the National Natural Science Foundation of China(31670179 and 91854201)the Research Grants Council of Hong Kong(CUHK14130716,14104716,14102417,14100818,14101219,C4012-16E,C4033-19E,C4002-17G,C4002-20W,R4005-18,and AOE/M-05/12)。
文摘The roles of Rho family guanosine triphosphatases(GTPases)of plants(ROPs)in modulating plant growth and development have been well characterized.However,little is known about the roles of ROP signaling pathways in regulating plant autophagy and autophagosome formation.In this study,we identify a unique ROP signaling mechanism,which mediates developmental to autophagic transition under stress conditions in the model plant Arabidopsis.Loss-of-function mutants of ROP8 showed stress-induced hypersensitive phenotypes and compromised autophagic flux.Similar to other ROPs in the ROP/RAC family,ROP8 exhibits both plasma membrane and cytosolic punctate localization patterns.Upon autophagic induction,active ROP8 puncta colocalize with autophagosomal markers and are degraded inside the vacuole.In human cells,RalB,an RAS subfamily GTPase,engages its effector Exo84 for autophagosome assembly.However,a RalB counterpart is missing in the plant lineage.Intriguingly,we discovered that plant ROP8 promotes autophagy via its downstream effector Sec5.Live-cell super-resolution imaging showed that ROP8 and Sec5 reside on phagophores for autophagosome formation.Taken together,our findings highlight a previously unappreciated role of an ROP8-Sec5 signaling axis in autophagy promotion,providing new insights into how plants utilize versatile ROP signaling networks to coordinate developmental and autophagic responses depending on environmental changes.
