Limits...
Nuclear envelope remnants: fluid membranes enriched in sterols and polyphosphoinositides.

Garnier-Lhomme M, Byrne RD, Hobday TM, Gschmeissner S, Woscholski R, Poccia DL, Dufourc EJ, Larijani B - PLoS ONE (2009)

Bottom Line: Although most of the sperm nuclear envelope disassembles, remnants of the envelope at the acrosomal and centriolar fossae do not and are subsequently incorporated into the newly forming male pronuclear envelope.Using lipid mass spectrometry and a new application of deuterium solid-state NMR spectroscopy we have characterized the lipid composition and membrane dynamics of the sperm nuclear envelope remnants in isolated sperm nuclei.The localization of the polybasic effector domain of MARCKS illustrates the non-nuclear aspect of the polyphosphoinositides.

View Article: PubMed Central - PubMed

Affiliation: Cell Biophysics Laboratory, Lincoln's Inn Fields Laboratories, Cancer Research UK, London, United Kingdom.

ABSTRACT

Background: The cytoplasm of eukaryotic cells is a highly dynamic compartment where membranes readily undergo fission and fusion to reorganize the cytoplasmic architecture, and to import, export and transport various cargos within the cell. The double membrane of the nuclear envelope that surrounds the nucleus, segregates the chromosomes from cytoplasm and regulates nucleocytoplasmic transport through pores. Many details of its formation are still unclear. At fertilization the sperm devoid of nuclear envelope pores enters the egg. Although most of the sperm nuclear envelope disassembles, remnants of the envelope at the acrosomal and centriolar fossae do not and are subsequently incorporated into the newly forming male pronuclear envelope. Remnants are conserved from annelid to mammalian sperm.

Methodology/principal findings: Using lipid mass spectrometry and a new application of deuterium solid-state NMR spectroscopy we have characterized the lipid composition and membrane dynamics of the sperm nuclear envelope remnants in isolated sperm nuclei.

Conclusions/significance: We report nuclear envelope remnants are relatively fluid membranes rich in sterols, devoid of sphingomyelin, and highly enriched in polyphosphoinositides and polyunsaturated phospholipids. The localization of the polybasic effector domain of MARCKS illustrates the non-nuclear aspect of the polyphosphoinositides. Based on their atypical biophysical characteristics and phospholipid composition, we suggest a possible role for nuclear envelope remnants in membrane fusion leading to nuclear envelope assembly.

Show MeSH
Poly-phosphoinositides in 0.1% nuclei and whole sperm are enriched in the acrosomal and centriolar fossae.L. pictus 0.1% nuclei (left) and whole live sperm (right) were incubated with the Texas Red labelled MARCKS peptide and visualised by fluorescence microscopy. The punctate staining of the acrosomal and centriolar fossae is typical of the majority of nuclei observed in experiments on two independent sperm and 0.1% nuclei preparations.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2626249&req=5

pone-0004255-g004: Poly-phosphoinositides in 0.1% nuclei and whole sperm are enriched in the acrosomal and centriolar fossae.L. pictus 0.1% nuclei (left) and whole live sperm (right) were incubated with the Texas Red labelled MARCKS peptide and visualised by fluorescence microscopy. The punctate staining of the acrosomal and centriolar fossae is typical of the majority of nuclei observed in experiments on two independent sperm and 0.1% nuclei preparations.

Mentions: To corroborate our mass spectrometry lipid data we determined the localization of poly-phosphoinositide lipids on 0.1% nuclei. This was undertaken with a Texas Red MARCKS peptide containing a polybasic series of residues that form non-specific electrostatic interactions with all negatively charged phospholipids, but preferentially with the more highly phosphorylated phosphoinositides. The MARCKS peptide will sequester PtdIns(4,5)P2 preferentially over PtdSer, even when the latter is 300-fold in excess [25]. Figure 4 shows the probe bound specifically to the regions of 0.1% nuclei corresponding to the NERs. This indicated that the poly-phosphoinositides detected in the ESI-MS/MS lipid analysis were localized in the NERs, and not within the chromatin region. The nuclei of eukaryoates are known to contain pools of phosphoinositides distinct from the lipids of the nuclear envelope [46], [47]. To rule out that the NER phosphoinositide enrichment was not an artifact created by a preferential retention of nuclear matrix phosphoinositides during Triton extraction, we stained live whole sperm with Texas Red MARCKs. The peptide bound preferentially to the region of the NERs in over 80% of stained sperm examined. These data together indicate that NERs are enriched in poly-phosphoinositides in both live whole sperm and 0.1% nuclei. Furthermore, the lipid composition of NERs is not artificially created by detergent extraction.


Nuclear envelope remnants: fluid membranes enriched in sterols and polyphosphoinositides.

Garnier-Lhomme M, Byrne RD, Hobday TM, Gschmeissner S, Woscholski R, Poccia DL, Dufourc EJ, Larijani B - PLoS ONE (2009)

Poly-phosphoinositides in 0.1% nuclei and whole sperm are enriched in the acrosomal and centriolar fossae.L. pictus 0.1% nuclei (left) and whole live sperm (right) were incubated with the Texas Red labelled MARCKS peptide and visualised by fluorescence microscopy. The punctate staining of the acrosomal and centriolar fossae is typical of the majority of nuclei observed in experiments on two independent sperm and 0.1% nuclei preparations.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2626249&req=5

pone-0004255-g004: Poly-phosphoinositides in 0.1% nuclei and whole sperm are enriched in the acrosomal and centriolar fossae.L. pictus 0.1% nuclei (left) and whole live sperm (right) were incubated with the Texas Red labelled MARCKS peptide and visualised by fluorescence microscopy. The punctate staining of the acrosomal and centriolar fossae is typical of the majority of nuclei observed in experiments on two independent sperm and 0.1% nuclei preparations.
Mentions: To corroborate our mass spectrometry lipid data we determined the localization of poly-phosphoinositide lipids on 0.1% nuclei. This was undertaken with a Texas Red MARCKS peptide containing a polybasic series of residues that form non-specific electrostatic interactions with all negatively charged phospholipids, but preferentially with the more highly phosphorylated phosphoinositides. The MARCKS peptide will sequester PtdIns(4,5)P2 preferentially over PtdSer, even when the latter is 300-fold in excess [25]. Figure 4 shows the probe bound specifically to the regions of 0.1% nuclei corresponding to the NERs. This indicated that the poly-phosphoinositides detected in the ESI-MS/MS lipid analysis were localized in the NERs, and not within the chromatin region. The nuclei of eukaryoates are known to contain pools of phosphoinositides distinct from the lipids of the nuclear envelope [46], [47]. To rule out that the NER phosphoinositide enrichment was not an artifact created by a preferential retention of nuclear matrix phosphoinositides during Triton extraction, we stained live whole sperm with Texas Red MARCKs. The peptide bound preferentially to the region of the NERs in over 80% of stained sperm examined. These data together indicate that NERs are enriched in poly-phosphoinositides in both live whole sperm and 0.1% nuclei. Furthermore, the lipid composition of NERs is not artificially created by detergent extraction.

Bottom Line: Although most of the sperm nuclear envelope disassembles, remnants of the envelope at the acrosomal and centriolar fossae do not and are subsequently incorporated into the newly forming male pronuclear envelope.Using lipid mass spectrometry and a new application of deuterium solid-state NMR spectroscopy we have characterized the lipid composition and membrane dynamics of the sperm nuclear envelope remnants in isolated sperm nuclei.The localization of the polybasic effector domain of MARCKS illustrates the non-nuclear aspect of the polyphosphoinositides.

View Article: PubMed Central - PubMed

Affiliation: Cell Biophysics Laboratory, Lincoln's Inn Fields Laboratories, Cancer Research UK, London, United Kingdom.

ABSTRACT

Background: The cytoplasm of eukaryotic cells is a highly dynamic compartment where membranes readily undergo fission and fusion to reorganize the cytoplasmic architecture, and to import, export and transport various cargos within the cell. The double membrane of the nuclear envelope that surrounds the nucleus, segregates the chromosomes from cytoplasm and regulates nucleocytoplasmic transport through pores. Many details of its formation are still unclear. At fertilization the sperm devoid of nuclear envelope pores enters the egg. Although most of the sperm nuclear envelope disassembles, remnants of the envelope at the acrosomal and centriolar fossae do not and are subsequently incorporated into the newly forming male pronuclear envelope. Remnants are conserved from annelid to mammalian sperm.

Methodology/principal findings: Using lipid mass spectrometry and a new application of deuterium solid-state NMR spectroscopy we have characterized the lipid composition and membrane dynamics of the sperm nuclear envelope remnants in isolated sperm nuclei.

Conclusions/significance: We report nuclear envelope remnants are relatively fluid membranes rich in sterols, devoid of sphingomyelin, and highly enriched in polyphosphoinositides and polyunsaturated phospholipids. The localization of the polybasic effector domain of MARCKS illustrates the non-nuclear aspect of the polyphosphoinositides. Based on their atypical biophysical characteristics and phospholipid composition, we suggest a possible role for nuclear envelope remnants in membrane fusion leading to nuclear envelope assembly.

Show MeSH