Nanoscale mapping of nuclear phosphatidylinositol phosphate landscape by dual-color dSTORM

Hoboth, Peter; Sztacho, Martin; Šebesta, Ondřej; Schätz, Martin; Castaño, Enrique; Hozák, Pavel

doi:10.1016/j.bbalip.2021.158890

Cited by 17 publications

(26 citation statements)

References 67 publications

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“…Consistent with the idea that specificity of interaction may be due to the presence of subnuclear pools of different PPIns, PARP1 colocalized with PtdIns(3,4,5) P 3 in the nucleolus and with PtdIns(3,4) P 2 in nucleoplasmic foci, and not with PtdIns(4,5) P 2 . The localization of PtdIns(3,4) P 2 in nucleoplasmic foci is consistent with recent studies ( 13 , 27 ). The identity of these foci has not been investigated but appear to be distinct to PtdIns(4,5) P 2 -positive sites that localize to nuclear speckles ( 21 ) but not with PARP1.…”

Section: Discussionsupporting

confidence: 93%

“…SHIP2 ( 107 ) or in its phosphorylated form on serine 132 ( 108 ) was found in nuclear speckles in different cells. Alternatively, the class II PI3K, PI3KC2α, known to produce PtdIns(3,4) P 2 by phosphorylating PtdIns4 P , was also reported to localize in nuclear foci ( 109 ) as well as its substrate PtdIns4 P ( 26 , 27 ), thus suggesting a potential synthesis route for PtdIns(3,4) P 2 in nuclear speckles. A key question is whether PtdIns(3,4) P 2 and PtdIns(3,4,5) P 3 bind to or even recruit PARP1 to their nuclear sites.…”

Section: Discussionmentioning

confidence: 99%

“…The concept of PPIn metabolism and signaling occurring in the nucleus independently of the cytoplasm was reported shortly after in several studies ( 10 , 11 , 12 ). Consequently, with the exception of phosphatidylinositol 3,5-bisphosphate, the remaining six PPIns have been detected and/or quantified in the nucleus ( 13 , 14 , 15 , 16 , 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 , 25 , 26 , 27 , 28 ). The intranuclear biophysico-chemical state of PPIns is still unclear, but several possibilities are emerging to explain how the acyl chains can be shielded from the nuclear aqueous environment.…”

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confidence: 99%

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Nuclear Phosphatidylinositol 3,4,5-Trisphosphate Interactome Uncovers an Enrichment in Nucleolar Proteins

Gavgani

Slinning

Morovicz

et al. 2021

Molecular & Cellular Proteomics

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Polyphosphoinositides (PPIns) play essential roles as lipid signaling molecules, and many of their functions have been elucidated in the cytoplasm. However, PPIns are also intranuclear where they contribute to chromatin remodeling, transcription, and mRNA splicing. The PPIn, phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5) P 3 ), has been mapped to the nucleus and nucleoli, but its role remains unclear in this subcellular compartment. To gain further insights into the nuclear functions of PtdIns(3,4,5) P 3 , we applied a previously developed quantitative MS-based approach to identify the targets of PtdIns(3,4,5) P 3 from isolated nuclei. We identified 179 potential PtdIns(3,4,5) P 3 -interacting partners, and gene ontology analysis for the biological functions of this dataset revealed an enrichment in RNA processing/splicing, cytokinesis, protein folding, and DNA repair. Interestingly, about half of these interactors were common to nucleolar protein datasets, some of which had dual functions in rRNA processes and DNA repair, including poly(ADP-ribose) polymerase 1 (PARP1, now referred as ADP-ribosyltransferase 1). PARP1 was found to interact directly with PPIn via three polybasic regions in the DNA-binding domain and the linker located N-terminal of the catalytic region. PARP1 was shown to bind to PtdIns(3,4,5) P 3 as well as phosphatidylinositol 3,4-bisphosphate in vitro and to colocalize with PtdIns(3,4,5) P 3 in the nucleolus and with phosphatidylinositol 3,4-bisphosphate in nucleoplasmic foci. In conclusion, the PtdIns(3,4,5) P 3 interactome reported here will serve as a resource to further investigate the molecular mechanisms underlying PtdIns(3,4,5) P 3 -mediated interactions in the nucleus and nucleolus.

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Section: Discussionsupporting

confidence: 93%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Nuclear Phosphatidylinositol 3,4,5-Trisphosphate Interactome Uncovers an Enrichment in Nucleolar Proteins

Gavgani

Slinning

Morovicz

et al. 2021

Molecular & Cellular Proteomics

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“…3), localized to the specific sub-nuclear compartments and linked with the crucial nuclear processes [110][111][112][113][114][115][116][117][118]. So far, we started to paint with nanometer precision the static pictures of nuclear PIP distributions [119]. However, as we learned from the cytoplasmic membrane, intracellular membrane compartments and from the nuclear envelope, lipids are highly mobile entities [52].…”

Section: Discussionmentioning

confidence: 99%

How Single-Molecule Localization Microscopy Expanded Our Mechanistic Understanding of RNA Polymerase II Transcription

Hoboth

Šebesta

Hozák

2021

Preprint

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Classical models of gene expression were built using genetics and biochemistry. Although these approaches are powerful, they have very limited consideration of the spatial and temporal organization of gene expression. Although the spatial organization and dynamics of RNA polymerase II (RNAPII) transcription machinery has fundamental functional consequences for gene expression, its detailed studies have been for long time abrogated by the limits of classical light microscopy. The advent of super-resolution microscopy (SRM) techniques allowed for the visualization of the RNAPII transcription machinery with nanometer resolution and millisecond precision. In this review, we summarize the recent methodological advances in SRM, focus on its application for studies of the nanoscale organization in space and time of RNAPII transcription, and discuss its consequences for the mechanistic understanding of gene expression.

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“…The precise localization approach that preserves the sub-nuclear architecture and the detailed quantitative analysis of the nuclear PIP2 distribution will expand the set of standard biochemical and lipidomic analyses, which are powerful but in their nature cumulative and thus neglect the spatial context of the nuclear architecture. This experimental pipeline is applicable for the functional studies of the role of nuclear PIP2, other nuclear PIPs and lipids in the establishment of nuclear architecture and regulation of gene expression [5] .…”

Section: Motivationmentioning

confidence: 99%

Dual-color dSTORM imaging and ThunderSTORM image reconstruction and analysis to study the spatial organization of the nuclear phosphatidylinositol phosphates

et al. 2021

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Single molecule localization microscopy (SMLM) provided an unprecedented insight into the sub-nuclear organization of proteins and nucleic acids but apart from the nuclear envelope the role of the nuclear lipids in the functional organization of the cell nucleus was less studied. Nevertheless, nuclear lipids and specifically phosphatidylinositol phosphates (PIPs) play increasingly evident roles in gene expression. Therefore, here we provide the SMLM-based approach for the quantitative evaluation of the nuclear PIPs distribution while preserving the context of nuclear architecture. Specifically, on the example of phosphatidylinositol 4,5-bisphosphate (PIP2) we have: Implemented and optimized the dual-color dSTORM imaging of nuclear PIP2. Customized the Nearest Neighbor Distance analysis using ImageJ2 plug-in ThunderSTORM to quantitatively evaluate the spatial distribution of nuclear PIP2. Developed an ImageJ2 tool for the visualization of the Nearest Neighbor Distance analysis results in cellulo . Our customization of the dual-color dSTORM imaging and quantitative analysis provide a tool that is independent of but complementary to the biochemical and lipidomic analyses of the nuclear PIPs. Contrary to the biochemical and lipidomic analyses, the advantage of our analysis is that it preserves the spatial context of the nuclear PIP distribution.

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Nanoscale mapping of nuclear phosphatidylinositol phosphate landscape by dual-color dSTORM

Cited by 17 publications

References 67 publications

Nuclear Phosphatidylinositol 3,4,5-Trisphosphate Interactome Uncovers an Enrichment in Nucleolar Proteins

Nuclear Phosphatidylinositol 3,4,5-Trisphosphate Interactome Uncovers an Enrichment in Nucleolar Proteins

How Single-Molecule Localization Microscopy Expanded Our Mechanistic Understanding of RNA Polymerase II Transcription

Dual-color dSTORM imaging and ThunderSTORM image reconstruction and analysis to study the spatial organization of the nuclear phosphatidylinositol phosphates

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