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The Inner Nuclear Membrane Protein Src1 Is Required for Stable Post-Mitotic Progression into G1 in Aspergillus nidulans.

Liu HL, Osmani AH, Osmani SA - PLoS ONE (2015)

Bottom Line: How membranes and associated proteins of the nuclear envelope (NE) are assembled specifically and inclusively around segregated genomes during exit from mitosis is incompletely understood.We suggest the term "reboot regulation" to define this mode of cell cycle regulation.The findings are discussed in relationship to recent studies showing the Cdk1 master oscillator can entrain subservient oscillators that when uncoupled cause cell cycle transitions to be repeated.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics, Ohio State University, Columbus, Ohio 43210, United States of America.

ABSTRACT
How membranes and associated proteins of the nuclear envelope (NE) are assembled specifically and inclusively around segregated genomes during exit from mitosis is incompletely understood. Inner nuclear membrane (INM) proteins play key roles by providing links between DNA and the NE. In this study we have investigated the highly conserved INM protein Src1 in Aspergillus nidulans and have uncovered a novel cell cycle response during post mitotic formation of G1 nuclei. Live cell imaging indicates Src1 could have roles during mitotic exit as it preferentially locates to the NE abscission points during nucleokinesis and to the NE surrounding forming daughter G1 nuclei. Deletion analysis further supported this idea revealing that although Src1 is not required for interphase progression or mitosis it is required for stable post-mitotic G1 nuclear formation. This conclusion is based upon the observation that in the absence of Src1 newly formed G1 nuclei are structurally unstable and immediately undergo architectural modifications typical of mitosis. These changes include NPC modifications that stop nuclear transport as well as disassembly of nucleoli. More intriguingly, the newly generated G1 nuclei then cycle between mitotic- and interphase-like states. The findings indicate that defects in post-mitotic G1 nuclear formation caused by lack of Src1 promote repeated failed attempts to generate stable G1 nuclei. To explain this unexpected phenotype we suggest a type of regulation that promotes repetition of defective cell cycle transitions rather than preventing progression past the defective cell cycle transition. We suggest the term "reboot regulation" to define this mode of cell cycle regulation. The findings are discussed in relationship to recent studies showing the Cdk1 master oscillator can entrain subservient oscillators that when uncoupled cause cell cycle transitions to be repeated.

No MeSH data available.


Related in: MedlinePlus

Nuclei oscillate between mitotic-like and interphase-like states in Δsrc1 cells.(A) Time course imaging over 34 minutes of NLS-DsRed in a Δsrc1 cell indicates its nuclei oscillate between nuclear transport active and inactive states. (B) To more clearly track individual nuclei within Δsrc1 cells the distribution of nuclear NLS-DsRed was monitored in benomyl treated cells to stop nuclear movements. Nuclei can be seen to oscillate between transport active and inactive states through the kymograph representing the intensity of NLS-DsRed across the yellow line with time; as also plotted as nuclear fluorescence intensity. The vertical red line indicates the first ten minutes of the live cell imaging. (C) Analysis of the data revealed nuclei spent on average an equal time between nuclear transport active and inactive states with no discernable pattern between each state between nuclei within individual cells.
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pone.0132489.g006: Nuclei oscillate between mitotic-like and interphase-like states in Δsrc1 cells.(A) Time course imaging over 34 minutes of NLS-DsRed in a Δsrc1 cell indicates its nuclei oscillate between nuclear transport active and inactive states. (B) To more clearly track individual nuclei within Δsrc1 cells the distribution of nuclear NLS-DsRed was monitored in benomyl treated cells to stop nuclear movements. Nuclei can be seen to oscillate between transport active and inactive states through the kymograph representing the intensity of NLS-DsRed across the yellow line with time; as also plotted as nuclear fluorescence intensity. The vertical red line indicates the first ten minutes of the live cell imaging. (C) Analysis of the data revealed nuclei spent on average an equal time between nuclear transport active and inactive states with no discernable pattern between each state between nuclei within individual cells.

Mentions: Time lapse microscopy of NLS-DsRed in Δsrc1 cells undergoing their first cell cycle found their single nuclei constitutively imported NLS-DsRed. However, analysis of post-mitotic Δsrc1 cells containing at least two nuclei showed that nuclei of cells lacking Src1 could oscillate between active and inactive nuclear transport states (Fig 6). These studies (Fig 6A) were complicated by the movement of nuclei within cells which made identification of individual nuclei difficult. Because nuclear movement is dependent on microtubules, we depolymerized microtubules using benomyl treatment then followed NLS-DsRed in post mitotic Δsrc1 nuclei that were now more stationary (Fig 6B). The analysis revealed that nuclei behaved in a nuclear-autonomous manner and that, although variable, on average switched from a transport competent state to a transport incompetent state every 3–4 minutes without a preferential stay in either one of the states (Fig 6C). We have never seen such oscillatory behavior in wildtype cells. Because nuclei were able to oscillate between transport active and inactive states in the presence of benomyl it suggests these transitions are not regulated by the SAC or, more likely, that such nuclei have passed the SAC arrest point.


The Inner Nuclear Membrane Protein Src1 Is Required for Stable Post-Mitotic Progression into G1 in Aspergillus nidulans.

Liu HL, Osmani AH, Osmani SA - PLoS ONE (2015)

Nuclei oscillate between mitotic-like and interphase-like states in Δsrc1 cells.(A) Time course imaging over 34 minutes of NLS-DsRed in a Δsrc1 cell indicates its nuclei oscillate between nuclear transport active and inactive states. (B) To more clearly track individual nuclei within Δsrc1 cells the distribution of nuclear NLS-DsRed was monitored in benomyl treated cells to stop nuclear movements. Nuclei can be seen to oscillate between transport active and inactive states through the kymograph representing the intensity of NLS-DsRed across the yellow line with time; as also plotted as nuclear fluorescence intensity. The vertical red line indicates the first ten minutes of the live cell imaging. (C) Analysis of the data revealed nuclei spent on average an equal time between nuclear transport active and inactive states with no discernable pattern between each state between nuclei within individual cells.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4492595&req=5

pone.0132489.g006: Nuclei oscillate between mitotic-like and interphase-like states in Δsrc1 cells.(A) Time course imaging over 34 minutes of NLS-DsRed in a Δsrc1 cell indicates its nuclei oscillate between nuclear transport active and inactive states. (B) To more clearly track individual nuclei within Δsrc1 cells the distribution of nuclear NLS-DsRed was monitored in benomyl treated cells to stop nuclear movements. Nuclei can be seen to oscillate between transport active and inactive states through the kymograph representing the intensity of NLS-DsRed across the yellow line with time; as also plotted as nuclear fluorescence intensity. The vertical red line indicates the first ten minutes of the live cell imaging. (C) Analysis of the data revealed nuclei spent on average an equal time between nuclear transport active and inactive states with no discernable pattern between each state between nuclei within individual cells.
Mentions: Time lapse microscopy of NLS-DsRed in Δsrc1 cells undergoing their first cell cycle found their single nuclei constitutively imported NLS-DsRed. However, analysis of post-mitotic Δsrc1 cells containing at least two nuclei showed that nuclei of cells lacking Src1 could oscillate between active and inactive nuclear transport states (Fig 6). These studies (Fig 6A) were complicated by the movement of nuclei within cells which made identification of individual nuclei difficult. Because nuclear movement is dependent on microtubules, we depolymerized microtubules using benomyl treatment then followed NLS-DsRed in post mitotic Δsrc1 nuclei that were now more stationary (Fig 6B). The analysis revealed that nuclei behaved in a nuclear-autonomous manner and that, although variable, on average switched from a transport competent state to a transport incompetent state every 3–4 minutes without a preferential stay in either one of the states (Fig 6C). We have never seen such oscillatory behavior in wildtype cells. Because nuclei were able to oscillate between transport active and inactive states in the presence of benomyl it suggests these transitions are not regulated by the SAC or, more likely, that such nuclei have passed the SAC arrest point.

Bottom Line: How membranes and associated proteins of the nuclear envelope (NE) are assembled specifically and inclusively around segregated genomes during exit from mitosis is incompletely understood.We suggest the term "reboot regulation" to define this mode of cell cycle regulation.The findings are discussed in relationship to recent studies showing the Cdk1 master oscillator can entrain subservient oscillators that when uncoupled cause cell cycle transitions to be repeated.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics, Ohio State University, Columbus, Ohio 43210, United States of America.

ABSTRACT
How membranes and associated proteins of the nuclear envelope (NE) are assembled specifically and inclusively around segregated genomes during exit from mitosis is incompletely understood. Inner nuclear membrane (INM) proteins play key roles by providing links between DNA and the NE. In this study we have investigated the highly conserved INM protein Src1 in Aspergillus nidulans and have uncovered a novel cell cycle response during post mitotic formation of G1 nuclei. Live cell imaging indicates Src1 could have roles during mitotic exit as it preferentially locates to the NE abscission points during nucleokinesis and to the NE surrounding forming daughter G1 nuclei. Deletion analysis further supported this idea revealing that although Src1 is not required for interphase progression or mitosis it is required for stable post-mitotic G1 nuclear formation. This conclusion is based upon the observation that in the absence of Src1 newly formed G1 nuclei are structurally unstable and immediately undergo architectural modifications typical of mitosis. These changes include NPC modifications that stop nuclear transport as well as disassembly of nucleoli. More intriguingly, the newly generated G1 nuclei then cycle between mitotic- and interphase-like states. The findings indicate that defects in post-mitotic G1 nuclear formation caused by lack of Src1 promote repeated failed attempts to generate stable G1 nuclei. To explain this unexpected phenotype we suggest a type of regulation that promotes repetition of defective cell cycle transitions rather than preventing progression past the defective cell cycle transition. We suggest the term "reboot regulation" to define this mode of cell cycle regulation. The findings are discussed in relationship to recent studies showing the Cdk1 master oscillator can entrain subservient oscillators that when uncoupled cause cell cycle transitions to be repeated.

No MeSH data available.


Related in: MedlinePlus