Xenopus laevis egg extracts cell-free nuclear assembly system was used as an experimental model to study the process of nuclear lamina assembly in nuclear reconstitution in vitro. The experimental results showed that ...Xenopus laevis egg extracts cell-free nuclear assembly system was used as an experimental model to study the process of nuclear lamina assembly in nuclear reconstitution in vitro. The experimental results showed that lamin was involved in the nuclear assembly in vitro. The assembly of nuclear lamina was preceded by the assembly of nuclear matrix, and probably, inner nuclear matrix assembly provided the basis for nuclear lamina assembly. Inhibition of normal assembly of nuclear lamina, by preincubating egg extracts cell-free system with anti-lamin antibodies, resulted in abnormal assembly of nuclear envelope, suggesting that nuclear envelope assembly is closely associated with nuclear lamina assembly.展开更多
1 Introduction Nuclear assembly occurs universally in cell life, such as postmitotic or postmeiotic nuclear assembly. It is a good model for studying the organization and function of nucleus. The organism or even sing...1 Introduction Nuclear assembly occurs universally in cell life, such as postmitotic or postmeiotic nuclear assembly. It is a good model for studying the organization and function of nucleus. The organism or even single cell, however, is a complex system in which many factors interact on each other, which is difficult to manipulate, in vitro cell-free nuclear assembly system can provide a good tool. Nuclear matrix is an insoluble proteinaceous filament network in nucleus. Much research work has demonstrated that many imoortant life actiyities such as展开更多
Accumulating evidence suggests that unicellular Archezoa are the most primitive eukaryotes and their nuclei are of significance to the study of evolution of the eukaryotic nucleus. Nuclear matrix is an ubiquitous impo...Accumulating evidence suggests that unicellular Archezoa are the most primitive eukaryotes and their nuclei are of significance to the study of evolution of the eukaryotic nucleus. Nuclear matrix is an ubiquitous important structure of eukaryotic nucleus; its evolution is certainly one of the most important parts of the evolution of nucleus. To study the evolution of nuclear matrix, nuclear matrices of Archezoa are investigated. Giardia lamblia cells are extracted sequentially. Both embedment free section EM and whole mount cell EM of the extracted cells show that, like higher eukaryotes, this species has a residual nuclear matrix in its nucleus and rich intermediate filaments in its cytoplasm, and the two networks connect with each other to form a united network. But its nuclear matrix does not have nucleolar matrix and its lamina is not as typical as that of higher eukaryotes; Western blotting shows that lamina of Giardia and two other Archezoa Entamoeba invadens and Trichomonas vaginali all contain only one polypeptide each which reacts with a mammalia anti lamin polyclonal serum and is similar to lamin B (67 ku) of mammlia in molecular weight. According to the results and references, it is suggested that nuclear matrix is an early acquisition of the eukaryotic nucleus, and it and the "eukaryotic chromatin" as a whole must have originated very early in the process of evolution of eukaryotic cell, and their origin should be an important prerequisite of the origin of eukaryotic nucleus; in the lamin (gene) family, B type lamins (gene) should be the ancestral type and that A type lamins (gene) might derive therefrom.展开更多
Using Electron Spectroscopic Imaging (ESI), we visualized the in situ binding of nucleic acids to nuclear matrix and 3H-thymidine incorporation which indicates that a small partial DNA bound to nuclear matrix tightly....Using Electron Spectroscopic Imaging (ESI), we visualized the in situ binding of nucleic acids to nuclear matrix and 3H-thymidine incorporation which indicates that a small partial DNA bound to nuclear matrix tightly. Furthermore we found that chromosomal telomere DNA could bind to nuclear matrix specifically by the dot and Southern hybridization. The result of the Southwestern blot suggests that telomere DNA has high affinity to lamin B, vimentin and some nuclear matrix proteins. Therefore, the nuclear matrix and lamina of HeLa cell are possibly associated with spatial organization and action of chromosome.展开更多
The fine structure of the nuclear lamina （NL） in sperm cells of \%Ginkgo biloba\% was visualised using high resolution low\|voltage scanning electron microscopy （LVSEM）. It was shown that the nuclear lamina was co...The fine structure of the nuclear lamina （NL） in sperm cells of \%Ginkgo biloba\% was visualised using high resolution low\|voltage scanning electron microscopy （LVSEM）. It was shown that the nuclear lamina was composed of 10 nm filaments which formed a fine network. Lamins were purified from cultured carrot suspension cells and assembled \%in vitro.\% Long 8\12 nm diameter filaments were seen and sometimes subfilaments could be distinguished. Western blot of filament preparations showed that these contained the 66 and 84 ku lamins. These data demonstrate that plant lamins are capable of assembling into filaments \%in vitro.\%展开更多
The eukaryotic genome is packaged as chromatin within the three-dimensional nuclear space. Decades of cytological studies have revealed that chromosomes and genes are non-randomly localized within the nucleus and such...The eukaryotic genome is packaged as chromatin within the three-dimensional nuclear space. Decades of cytological studies have revealed that chromosomes and genes are non-randomly localized within the nucleus and such organizations have important roles on genome function. However, several fundamental questions remain to be resolved. For example, what is required for the preferential localization ofa gene to a nuclear landmark？ What is the mechanism underlying gene repositioning in the nucleus？ How does subnuclear gene positioning regulate gene transcription？ Recent studies have revealed that several factors such as DNA sequence composition, specific regulatory sequences, epigenetic modifications, chromatin remodelers, post-transcriptional regulators and nuclear architectural proteins can influence chromatin dynamics and gene positioning in a gene-specific manner among organisms from yeast to human. In this review, we discuss some recent findings as well as experimental tools to investigate subnuclear gene positioning and to explore its implications in genome functions.展开更多
The nuclear matrix of diplomonad Giardia lamblia was detected for the first time with DGD embedmentsectioning- embedment free electron microscopy after a series of specific extractions. The result showed that archaezo...The nuclear matrix of diplomonad Giardia lamblia was detected for the first time with DGD embedmentsectioning- embedment free electron microscopy after a series of specific extractions. The result showed that archaezoa Giardia lamblia already possessed nuclear matrix within its two nuclei. The finest fibrils of the nuclear matrix of Giardia lamblia were measured to be about 11 to 13 nm in thickness. However, the nuclear lamina and nucleolus have never been observed. These results seem to suggest that nuclear matrix is an indispensable intranuclear structural component even in the primitive nucleus.展开更多
In this study, freeze - fractured specimens of Allium cepa root tip meristems were examined under the scanning electron microscope (SEM). This technique permitted the visualization of the outer membrane of the nuclear...In this study, freeze - fractured specimens of Allium cepa root tip meristems were examined under the scanning electron microscope (SEM). This technique permitted the visualization of the outer membrane of the nuclear envelope with nuclear pore complexes and polyribosomes. Some of the cell nuclei prepared with this procedure had fissures of various widths on their nuclear envelopes through which the nuclear lamina-like filaments (LLF) underneath the nucleoplasmic side of the envelopes were clearly visible. The diameters of these filaments varied between 25 and 125 nm. Many of the LLFs showed granular thickenings at places, and were attached to the inner surface of nuclear envelope in some regions. Similar LLFs were also seen at the peripheries of the freeze -fractured faces of nuclei. Meanwhile,the spatial relation between the nuclear matrix filaments (NMF) and other nuclear structures (nucleoli, chromatin and peripheral lamina - like filaments) was revealed in these fractured preparations. In addition, the methods and techniques in studying the nuclear lamina morphology and the roles played by NMFs in activities of various nuclear structures were discussed in brief.展开更多
Objective:The Hutchinson-Gilford progeria syndrome (HGPS or progeria) is a childhood disorder with features of premature aging and is caused by mutations in the lamin A gene resulting in the production of an abnormal ...Objective:The Hutchinson-Gilford progeria syndrome (HGPS or progeria) is a childhood disorder with features of premature aging and is caused by mutations in the lamin A gene resulting in the production of an abnormal protein,termed progerin. To investigate the underlying pathogenic mechanism,we studied the nuclear co-localization and association of progerin interactive partner proteins (PIPPs) with lamina proteins. Methods:Both wild-type (WT) and progeria fibroblasts were studied by various methods including confocal microscopy,immunoprecipitation and Western blot.Results:All PIPPs discovered so-far co-localized with lamin A/C. In addition,the PIPPs were selectively associated with lamina proteins. An increased immunofluorescent staining signal was found for Mel18 in HGPS as compared to WT cells. An association of Mel18 with emerin was observed in HGPS,but not in WT cells. Conclusion:Based on these findings,we propose that PIPPs,along with associated lamina proteins may form a pathogenic progerin-containing protein complex.展开更多
A cell-free preparation obtained from extracts of activated Xenopus laevis eggs induced chromatin decondensation and nuclear formation from demembranated Xenopus sperm nuclei. Electron microscopy revealed that the rea...A cell-free preparation obtained from extracts of activated Xenopus laevis eggs induced chromatin decondensation and nuclear formation from demembranated Xenopus sperm nuclei. Electron microscopy revealed that the reassembled nucleus had a double-layered nuclear membrane, nuclear pore complexes, and decondensed chromatin etc. Indirect immunofluorescence analysis demonstrated the presence of lamina in newly assembled nuclei. Western-blotting results showed that lamin LII was present in egg extracts and in lamina of the reassembled nuclei which were previously reported to contain only eggderived lamin LIII.展开更多
After lamins A, B and C were isolated and purified from rat liver, their assembly properties were examined by electron microscopy and scanning tunneling microscopy using negative staining and the glycerol coating meth...After lamins A, B and C were isolated and purified from rat liver, their assembly properties were examined by electron microscopy and scanning tunneling microscopy using negative staining and the glycerol coating method,respectively By varying the assembly time or the ionic conditions under which polymerization takes place, we have observed different stages of lamin assembly, which may provide clues on the structure of the 10 nm lamin filaments. At the first level of structural organization, two lamin polypeptides associate laterally into dimers with the two domains being parallel and in register. At the second level of structural organization, two dimers associate in a half-staggered and atiparallel fashion to form a tetramer 75 nm in length. At the third level of structural organization, 4-10 lamin tetramers associate laterally in register to form 75 nm long 10nm filaments, which in turn combine head to head into long, fully assembled lamin filaments.The assembled lamin filaments are nonpolar.展开更多
Herpesviruses assemble and fill their capsids in the infected cell nucleus, and must then move this enormous macromolecular assembly across the nuclear membrane and into the cytoplasm. Doing so is a complex, multi-ste...Herpesviruses assemble and fill their capsids in the infected cell nucleus, and must then move this enormous macromolecular assembly across the nuclear membrane and into the cytoplasm. Doing so is a complex, multi-step process that involves envelopment of the capsid at the inner nuclear membrane and de-envelopment by fusion with the outer nuclear membrane. This process is orchestrated by viral proteins, but requires the modification of cellular structures and mechanisms including the nuclear lamina. In this review I summarize recent research on the mechanism of nuclear envelopment and the viral and cellular systems involved in its execution.展开更多
基金Project supported by the National Natural Science Foundation of China.
文摘Xenopus laevis egg extracts cell-free nuclear assembly system was used as an experimental model to study the process of nuclear lamina assembly in nuclear reconstitution in vitro. The experimental results showed that lamin was involved in the nuclear assembly in vitro. The assembly of nuclear lamina was preceded by the assembly of nuclear matrix, and probably, inner nuclear matrix assembly provided the basis for nuclear lamina assembly. Inhibition of normal assembly of nuclear lamina, by preincubating egg extracts cell-free system with anti-lamin antibodies, resulted in abnormal assembly of nuclear envelope, suggesting that nuclear envelope assembly is closely associated with nuclear lamina assembly.
基金Project supported by the National Natural Science Foundation of China.
文摘1 Introduction Nuclear assembly occurs universally in cell life, such as postmitotic or postmeiotic nuclear assembly. It is a good model for studying the organization and function of nucleus. The organism or even single cell, however, is a complex system in which many factors interact on each other, which is difficult to manipulate, in vitro cell-free nuclear assembly system can provide a good tool. Nuclear matrix is an insoluble proteinaceous filament network in nucleus. Much research work has demonstrated that many imoortant life actiyities such as
文摘Accumulating evidence suggests that unicellular Archezoa are the most primitive eukaryotes and their nuclei are of significance to the study of evolution of the eukaryotic nucleus. Nuclear matrix is an ubiquitous important structure of eukaryotic nucleus; its evolution is certainly one of the most important parts of the evolution of nucleus. To study the evolution of nuclear matrix, nuclear matrices of Archezoa are investigated. Giardia lamblia cells are extracted sequentially. Both embedment free section EM and whole mount cell EM of the extracted cells show that, like higher eukaryotes, this species has a residual nuclear matrix in its nucleus and rich intermediate filaments in its cytoplasm, and the two networks connect with each other to form a united network. But its nuclear matrix does not have nucleolar matrix and its lamina is not as typical as that of higher eukaryotes; Western blotting shows that lamina of Giardia and two other Archezoa Entamoeba invadens and Trichomonas vaginali all contain only one polypeptide each which reacts with a mammalia anti lamin polyclonal serum and is similar to lamin B (67 ku) of mammlia in molecular weight. According to the results and references, it is suggested that nuclear matrix is an early acquisition of the eukaryotic nucleus, and it and the "eukaryotic chromatin" as a whole must have originated very early in the process of evolution of eukaryotic cell, and their origin should be an important prerequisite of the origin of eukaryotic nucleus; in the lamin (gene) family, B type lamins (gene) should be the ancestral type and that A type lamins (gene) might derive therefrom.
文摘Using Electron Spectroscopic Imaging (ESI), we visualized the in situ binding of nucleic acids to nuclear matrix and 3H-thymidine incorporation which indicates that a small partial DNA bound to nuclear matrix tightly. Furthermore we found that chromosomal telomere DNA could bind to nuclear matrix specifically by the dot and Southern hybridization. The result of the Southwestern blot suggests that telomere DNA has high affinity to lamin B, vimentin and some nuclear matrix proteins. Therefore, the nuclear matrix and lamina of HeLa cell are possibly associated with spatial organization and action of chromosome.
文摘The fine structure of the nuclear lamina （NL） in sperm cells of \%Ginkgo biloba\% was visualised using high resolution low\|voltage scanning electron microscopy （LVSEM）. It was shown that the nuclear lamina was composed of 10 nm filaments which formed a fine network. Lamins were purified from cultured carrot suspension cells and assembled \%in vitro.\% Long 8\12 nm diameter filaments were seen and sometimes subfilaments could be distinguished. Western blot of filament preparations showed that these contained the 66 and 84 ku lamins. These data demonstrate that plant lamins are capable of assembling into filaments \%in vitro.\%
文摘The eukaryotic genome is packaged as chromatin within the three-dimensional nuclear space. Decades of cytological studies have revealed that chromosomes and genes are non-randomly localized within the nucleus and such organizations have important roles on genome function. However, several fundamental questions remain to be resolved. For example, what is required for the preferential localization ofa gene to a nuclear landmark？ What is the mechanism underlying gene repositioning in the nucleus？ How does subnuclear gene positioning regulate gene transcription？ Recent studies have revealed that several factors such as DNA sequence composition, specific regulatory sequences, epigenetic modifications, chromatin remodelers, post-transcriptional regulators and nuclear architectural proteins can influence chromatin dynamics and gene positioning in a gene-specific manner among organisms from yeast to human. In this review, we discuss some recent findings as well as experimental tools to investigate subnuclear gene positioning and to explore its implications in genome functions.
文摘The nuclear matrix of diplomonad Giardia lamblia was detected for the first time with DGD embedmentsectioning- embedment free electron microscopy after a series of specific extractions. The result showed that archaezoa Giardia lamblia already possessed nuclear matrix within its two nuclei. The finest fibrils of the nuclear matrix of Giardia lamblia were measured to be about 11 to 13 nm in thickness. However, the nuclear lamina and nucleolus have never been observed. These results seem to suggest that nuclear matrix is an indispensable intranuclear structural component even in the primitive nucleus.
文摘In this study, freeze - fractured specimens of Allium cepa root tip meristems were examined under the scanning electron microscope (SEM). This technique permitted the visualization of the outer membrane of the nuclear envelope with nuclear pore complexes and polyribosomes. Some of the cell nuclei prepared with this procedure had fissures of various widths on their nuclear envelopes through which the nuclear lamina-like filaments (LLF) underneath the nucleoplasmic side of the envelopes were clearly visible. The diameters of these filaments varied between 25 and 125 nm. Many of the LLFs showed granular thickenings at places, and were attached to the inner surface of nuclear envelope in some regions. Similar LLFs were also seen at the peripheries of the freeze -fractured faces of nuclei. Meanwhile,the spatial relation between the nuclear matrix filaments (NMF) and other nuclear structures (nucleoli, chromatin and peripheral lamina - like filaments) was revealed in these fractured preparations. In addition, the methods and techniques in studying the nuclear lamina morphology and the roles played by NMFs in activities of various nuclear structures were discussed in brief.
文摘Objective:The Hutchinson-Gilford progeria syndrome (HGPS or progeria) is a childhood disorder with features of premature aging and is caused by mutations in the lamin A gene resulting in the production of an abnormal protein,termed progerin. To investigate the underlying pathogenic mechanism,we studied the nuclear co-localization and association of progerin interactive partner proteins (PIPPs) with lamina proteins. Methods:Both wild-type (WT) and progeria fibroblasts were studied by various methods including confocal microscopy,immunoprecipitation and Western blot.Results:All PIPPs discovered so-far co-localized with lamin A/C. In addition,the PIPPs were selectively associated with lamina proteins. An increased immunofluorescent staining signal was found for Mel18 in HGPS as compared to WT cells. An association of Mel18 with emerin was observed in HGPS,but not in WT cells. Conclusion:Based on these findings,we propose that PIPPs,along with associated lamina proteins may form a pathogenic progerin-containing protein complex.
文摘A cell-free preparation obtained from extracts of activated Xenopus laevis eggs induced chromatin decondensation and nuclear formation from demembranated Xenopus sperm nuclei. Electron microscopy revealed that the reassembled nucleus had a double-layered nuclear membrane, nuclear pore complexes, and decondensed chromatin etc. Indirect immunofluorescence analysis demonstrated the presence of lamina in newly assembled nuclei. Western-blotting results showed that lamin LII was present in egg extracts and in lamina of the reassembled nuclei which were previously reported to contain only eggderived lamin LIII.
文摘After lamins A, B and C were isolated and purified from rat liver, their assembly properties were examined by electron microscopy and scanning tunneling microscopy using negative staining and the glycerol coating method,respectively By varying the assembly time or the ionic conditions under which polymerization takes place, we have observed different stages of lamin assembly, which may provide clues on the structure of the 10 nm lamin filaments. At the first level of structural organization, two lamin polypeptides associate laterally into dimers with the two domains being parallel and in register. At the second level of structural organization, two dimers associate in a half-staggered and atiparallel fashion to form a tetramer 75 nm in length. At the third level of structural organization, 4-10 lamin tetramers associate laterally in register to form 75 nm long 10nm filaments, which in turn combine head to head into long, fully assembled lamin filaments.The assembled lamin filaments are nonpolar.
基金supported by US Public health Service award AI 41478
文摘Herpesviruses assemble and fill their capsids in the infected cell nucleus, and must then move this enormous macromolecular assembly across the nuclear membrane and into the cytoplasm. Doing so is a complex, multi-step process that involves envelopment of the capsid at the inner nuclear membrane and de-envelopment by fusion with the outer nuclear membrane. This process is orchestrated by viral proteins, but requires the modification of cellular structures and mechanisms including the nuclear lamina. In this review I summarize recent research on the mechanism of nuclear envelopment and the viral and cellular systems involved in its execution.
