Rice tiller angle is a key agronomic trait that contributes to ideal plant architecture and grain production.LAZY1 (LA1) was previously shown to control tiller angle via affecting shoot gravitropism,but the underlying...Rice tiller angle is a key agronomic trait that contributes to ideal plant architecture and grain production.LAZY1 (LA1) was previously shown to control tiller angle via affecting shoot gravitropism,but the underlying molecular mechanism remains largely unknown.In this study,we identified an LA1-interacting protein named Brevis Radix Like 4 (OsBRXL4).We showed that the interaction between OsBRXL4 and LA1 occurs at the plasma membrane and that their interaction determines nuclear localization of LA1.We found that nuclear localization of LA1 is essential for its function,which is different from AtLA1,its Arabidopsis ortho.log.Overexpression of OsBRXL4 leads to a prostrate growth phenotype,whereas OsBRXLs RNAi plants,in which the expression levels of OsBRXLI,OsBRXL4,and OsBRXL5 were decreased,display a compact phenotype.Further genetic analysis also supported that OsBRXL4 controls rice tiller angle by affecting nuclear localization of LA1.Consistently,we demonstrated that OsBRXL4 regulates the shoot gravitropism through affecting polar auxin transport as did LA1.Taken together,our study not only identifies OsBRXL4 as a regulatory component of rice tiller angle but also provides new insights into genetic regulation of rice plant architecture.展开更多
AIM: To characterize the nuclear import of hepatitis B virus (HBV) polymerase (P) and its relevance for the viral life cycle.METHODS: Sequence analysis was performed to predict functional motives within P. Phosphoryla...AIM: To characterize the nuclear import of hepatitis B virus (HBV) polymerase (P) and its relevance for the viral life cycle.METHODS: Sequence analysis was performed to predict functional motives within P. Phosphorylation of P was analyzed by in vitro phosphorylation. Phosphorylation site and nuclear localization signal (NLS) were destroyed by site directed mutagenesis. Functionality of the identified NLS was analyzed by confocal fluorescence microscopy and characterizing the karyopherin binding. Relevance of the structural motives for viral life cycle was studied by infection of primary Tupaia hepatocytes with HBV.RESULTS: We identified by sequence alignment and functional experiments a conserved bipartite NLS containing a casein kinase II (CKII) phosphorylation site located within the terminal protein domain (TP) of the HBV polymerase. Inhibition of CKII impairs the functionality of this NLS and thereby prevents the nuclear import of the polymerase. Binding of the import factor karyopherin-α2 to the polymerase depends on its CKII-mediated phosphorylation of the bipartite NLS. In HBV-infected primary Tupaia hepatocytes CKII inhibition in the early phase (post entry phase) of the infection process prevents the establishment of the infection.CONCLUSION: Based on these data it is suggested that during HBV infection the final import of the genome complex into the nucleus is mediated by a novel bipartite NLS localized in the TP domain of HBV polymerase.展开更多
For genome mulUplication hepadnaviruses use the transcriptional machinery of the cell that is found within the nucleus. Thus the viral genome has to be transported through the cytoplasm and nuclear pore. The intracyto...For genome mulUplication hepadnaviruses use the transcriptional machinery of the cell that is found within the nucleus. Thus the viral genome has to be transported through the cytoplasm and nuclear pore. The intracytosolic translocation is facilitated by the viral capsid that surrounds the genome and that interacts with cellular microtubules. The subsequent passage through the nuclear pore complexes （NPC） is mediated by the nuclear transport receptors importin α andβ. Importin α binds to the C-terminus of the capsid protein that comprises a nuclear localization signal （NLS）. The exposure of the NLS is regulated and depends upon genome maturation and/or phosphorylation of the capsid protein. As for other karyophilic cargos using this pathway importin α interacts with importin β that facilitates docking of the import complex to the NPC and the passage through the pore. Being a unique strategy, the import of the viral capsid is incomplete in that it becomes arrested inside the nuclear basket, which is a cage-like structure on the karyoplasmic face of the NPC. Presumably only this compartment provides the factors that are required for capsid disassembly and genome release that is restricted to those capsids comprising a mature viral DNA genome.展开更多
Homeodomains,a 60-amino acid sequence encoded by 180 nucleotides,are highly conserved DNA-binding motifs that are present in a variety of transcription factors in species ranging from yeast to humans.The NKX proteins ...Homeodomains,a 60-amino acid sequence encoded by 180 nucleotides,are highly conserved DNA-binding motifs that are present in a variety of transcription factors in species ranging from yeast to humans.The NKX proteins belong to the homeodomain(HD)-containing transcription factor family.They play vital roles in the regulation of morphogenesis.NKX1-2 is one member of the NKX subfamily.At present,information about its nuclear localization signal(NLS)sequence is limited.We studied the NLS sequence of zebrafish Nkx1.2 by introducing sequence changes such as deletion,mutation,and truncation,and identified an NLS motif(QNRRTKWKKQ)that is localized at the C-terminus of the homeodomain.Moreover,the deletion of two amino acid residues(RR)in this NLS motif prevents Nkx1.2 from entering the nucleus,indicating that the two amino acids are essential for Nkx1.2 nuclear localization.However,the NLS motif alone is unable to target cytoplasmic protein glutathione S-transferase(GST)to the nucleus.An intact homeodomain is necessary for mediating the complete nuclear transport of cytoplasmic protein.Unlike most nuclear import proteins with short NLS sequences,a long NLS is present in zebrafish Nkx1.2.We also demonstrated that the sequences of homeodomain of NKX1.2 are well conserved among different species.This study is informative to verify the function of the NKX1.2 protein.展开更多
基金supported by grants from the National Natural Science Foundation of China (9163530191535204+2 种基金31601276)the Ministry of Agriculture of China (2016ZX08009-003)the Strategic Priority Research Program "Molecular Mechanism of Plant Growth and Development" of CAS (XDPB0401).
文摘Rice tiller angle is a key agronomic trait that contributes to ideal plant architecture and grain production.LAZY1 (LA1) was previously shown to control tiller angle via affecting shoot gravitropism,but the underlying molecular mechanism remains largely unknown.In this study,we identified an LA1-interacting protein named Brevis Radix Like 4 (OsBRXL4).We showed that the interaction between OsBRXL4 and LA1 occurs at the plasma membrane and that their interaction determines nuclear localization of LA1.We found that nuclear localization of LA1 is essential for its function,which is different from AtLA1,its Arabidopsis ortho.log.Overexpression of OsBRXL4 leads to a prostrate growth phenotype,whereas OsBRXLs RNAi plants,in which the expression levels of OsBRXLI,OsBRXL4,and OsBRXL5 were decreased,display a compact phenotype.Further genetic analysis also supported that OsBRXL4 controls rice tiller angle by affecting nuclear localization of LA1.Consistently,we demonstrated that OsBRXL4 regulates the shoot gravitropism through affecting polar auxin transport as did LA1.Taken together,our study not only identifies OsBRXL4 as a regulatory component of rice tiller angle but also provides new insights into genetic regulation of rice plant architecture.
文摘AIM: To characterize the nuclear import of hepatitis B virus (HBV) polymerase (P) and its relevance for the viral life cycle.METHODS: Sequence analysis was performed to predict functional motives within P. Phosphorylation of P was analyzed by in vitro phosphorylation. Phosphorylation site and nuclear localization signal (NLS) were destroyed by site directed mutagenesis. Functionality of the identified NLS was analyzed by confocal fluorescence microscopy and characterizing the karyopherin binding. Relevance of the structural motives for viral life cycle was studied by infection of primary Tupaia hepatocytes with HBV.RESULTS: We identified by sequence alignment and functional experiments a conserved bipartite NLS containing a casein kinase II (CKII) phosphorylation site located within the terminal protein domain (TP) of the HBV polymerase. Inhibition of CKII impairs the functionality of this NLS and thereby prevents the nuclear import of the polymerase. Binding of the import factor karyopherin-α2 to the polymerase depends on its CKII-mediated phosphorylation of the bipartite NLS. In HBV-infected primary Tupaia hepatocytes CKII inhibition in the early phase (post entry phase) of the infection process prevents the establishment of the infection.CONCLUSION: Based on these data it is suggested that during HBV infection the final import of the genome complex into the nucleus is mediated by a novel bipartite NLS localized in the TP domain of HBV polymerase.
文摘For genome mulUplication hepadnaviruses use the transcriptional machinery of the cell that is found within the nucleus. Thus the viral genome has to be transported through the cytoplasm and nuclear pore. The intracytosolic translocation is facilitated by the viral capsid that surrounds the genome and that interacts with cellular microtubules. The subsequent passage through the nuclear pore complexes （NPC） is mediated by the nuclear transport receptors importin α andβ. Importin α binds to the C-terminus of the capsid protein that comprises a nuclear localization signal （NLS）. The exposure of the NLS is regulated and depends upon genome maturation and/or phosphorylation of the capsid protein. As for other karyophilic cargos using this pathway importin α interacts with importin β that facilitates docking of the import complex to the NPC and the passage through the pore. Being a unique strategy, the import of the viral capsid is incomplete in that it becomes arrested inside the nuclear basket, which is a cage-like structure on the karyoplasmic face of the NPC. Presumably only this compartment provides the factors that are required for capsid disassembly and genome release that is restricted to those capsids comprising a mature viral DNA genome.
基金Supported by the National Natural Science Foundation of China(No.31970429)the Shandong Provincial Natural Science Foundation(No.ZR 2022 MC 032)。
文摘Homeodomains,a 60-amino acid sequence encoded by 180 nucleotides,are highly conserved DNA-binding motifs that are present in a variety of transcription factors in species ranging from yeast to humans.The NKX proteins belong to the homeodomain(HD)-containing transcription factor family.They play vital roles in the regulation of morphogenesis.NKX1-2 is one member of the NKX subfamily.At present,information about its nuclear localization signal(NLS)sequence is limited.We studied the NLS sequence of zebrafish Nkx1.2 by introducing sequence changes such as deletion,mutation,and truncation,and identified an NLS motif(QNRRTKWKKQ)that is localized at the C-terminus of the homeodomain.Moreover,the deletion of two amino acid residues(RR)in this NLS motif prevents Nkx1.2 from entering the nucleus,indicating that the two amino acids are essential for Nkx1.2 nuclear localization.However,the NLS motif alone is unable to target cytoplasmic protein glutathione S-transferase(GST)to the nucleus.An intact homeodomain is necessary for mediating the complete nuclear transport of cytoplasmic protein.Unlike most nuclear import proteins with short NLS sequences,a long NLS is present in zebrafish Nkx1.2.We also demonstrated that the sequences of homeodomain of NKX1.2 are well conserved among different species.This study is informative to verify the function of the NKX1.2 protein.
