Spatial Distribution of Cellular Function: The Partitioning of Proteins between Mitochondria and the Nucleus in MCF7 Breast Cancer Cells

Concurrent proteomics analysis of the nuclei and mitochondria of MCF7 breast cancer cells identified 985 proteins (40% of all detected proteins) present in both organelles. Numerous proteins from all five complexes involved in oxidative phosphorylation (e.g., NDUFA5, NDUFB10, NDUFS1, NDUF2, SDHA, UQRB, UQRC2, UQCRH, COX5A, COX5B, MT-CO2, ATP5A1, ATP5B, ATP5H, etc.), from the TCA-cycle (DLST, IDH2, IDH3A, OGDH, SUCLAG2, etc.), and from glycolysis (ALDOA, ENO1, FBP1, GPI, PGK1, TALDO1, etc.) were distributed to both the nucleus and mitochondria. In contrast, proteins involved in nuclear/mitochondrial RNA processing/translation and Ras/Rab signaling showed different partitioning patterns. The identity of the OxPhos, TCA-cycle, and glycolysis proteins distributed to both the nucleus and mitochondria provides evidence for spatio-functional integration of these processes over the two different subcellular organelles. We suggest that there are unrecognized aspects of functional coordination between the nucleus and mitochondria, that integration of core functional processes via wide subcellular distribution of constituent proteins is a common characteristic of cells, and that subcellular spatial integration of function may be a vital aspect of cancer

Spatial Distribution of Cellular Function: The Partitioning of Proteins between Mitochondria and the Nucleus in MCF7 Breast Cancer Cells

Concurrent proteomics analysis of the nuclei and mitochondria of MCF7 breast cancer cells identified 985 proteins (40% of all detected proteins) present in both organelles. Numerous proteins from all five complexes involved in oxidative phosphorylation (e.g., NDUFA5, NDUFB10, NDUFS1, NDUF2, SDHA, UQRB, UQRC2, UQCRH, COX5A, COX5B, MT-CO2, ATP5A1, ATP5B, ATP5H, etc.), from the TCA-cycle (DLST, IDH2, IDH3A, OGDH, SUCLAG2, etc.), and from glycolysis (ALDOA, ENO1, FBP1, GPI, PGK1, TALDO1, etc.) were distributed to both the nucleus and mitochondria. In contrast, proteins involved in nuclear/mitochondrial RNA processing/translation and Ras/Rab signaling showed different partitioning patterns. The identity of the OxPhos, TCA-cycle, and glycolysis proteins distributed to both the nucleus and mitochondria provides evidence for spatio-functional integration of these processes over the two different subcellular organelles. We suggest that there are unrecognized aspects of functional coordination between the nucleus and mitochondria, that integration of core functional processes via wide subcellular distribution of constituent proteins is a common characteristic of cells, and that subcellular spatial integration of function may be a vital aspect of cancer

Spatial Distribution of Cellular Function: The Partitioning of Proteins between Mitochondria and the Nucleus in MCF7 Breast Cancer Cells

Concurrent proteomics analysis of the nuclei and mitochondria of MCF7 breast cancer cells identified 985 proteins (40% of all detected proteins) present in both organelles. Numerous proteins from all five complexes involved in oxidative phosphorylation (e.g., NDUFA5, NDUFB10, NDUFS1, NDUF2, SDHA, UQRB, UQRC2, UQCRH, COX5A, COX5B, MT-CO2, ATP5A1, ATP5B, ATP5H, etc.), from the TCA-cycle (DLST, IDH2, IDH3A, OGDH, SUCLAG2, etc.), and from glycolysis (ALDOA, ENO1, FBP1, GPI, PGK1, TALDO1, etc.) were distributed to both the nucleus and mitochondria. In contrast, proteins involved in nuclear/mitochondrial RNA processing/translation and Ras/Rab signaling showed different partitioning patterns. The identity of the OxPhos, TCA-cycle, and glycolysis proteins distributed to both the nucleus and mitochondria provides evidence for spatio-functional integration of these processes over the two different subcellular organelles. We suggest that there are unrecognized aspects of functional coordination between the nucleus and mitochondria, that integration of core functional processes via wide subcellular distribution of constituent proteins is a common characteristic of cells, and that subcellular spatial integration of function may be a vital aspect of cancer

Spatial Distribution of Cellular Function: The Partitioning of Proteins between Mitochondria and the Nucleus in MCF7 Breast Cancer Cells

Concurrent proteomics analysis of the nuclei and mitochondria of MCF7 breast cancer cells identified 985 proteins (40% of all detected proteins) present in both organelles. Numerous proteins from all five complexes involved in oxidative phosphorylation (e.g., NDUFA5, NDUFB10, NDUFS1, NDUF2, SDHA, UQRB, UQRC2, UQCRH, COX5A, COX5B, MT-CO2, ATP5A1, ATP5B, ATP5H, etc.), from the TCA-cycle (DLST, IDH2, IDH3A, OGDH, SUCLAG2, etc.), and from glycolysis (ALDOA, ENO1, FBP1, GPI, PGK1, TALDO1, etc.) were distributed to both the nucleus and mitochondria. In contrast, proteins involved in nuclear/mitochondrial RNA processing/translation and Ras/Rab signaling showed different partitioning patterns. The identity of the OxPhos, TCA-cycle, and glycolysis proteins distributed to both the nucleus and mitochondria provides evidence for spatio-functional integration of these processes over the two different subcellular organelles. We suggest that there are unrecognized aspects of functional coordination between the nucleus and mitochondria, that integration of core functional processes via wide subcellular distribution of constituent proteins is a common characteristic of cells, and that subcellular spatial integration of function may be a vital aspect of cancer

Spatial Distribution of Cellular Function: The Partitioning of Proteins between Mitochondria and the Nucleus in MCF7 Breast Cancer Cells

Concurrent proteomics analysis of the nuclei and mitochondria of MCF7 breast cancer cells identified 985 proteins (40% of all detected proteins) present in both organelles. Numerous proteins from all five complexes involved in oxidative phosphorylation (e.g., NDUFA5, NDUFB10, NDUFS1, NDUF2, SDHA, UQRB, UQRC2, UQCRH, COX5A, COX5B, MT-CO2, ATP5A1, ATP5B, ATP5H, etc.), from the TCA-cycle (DLST, IDH2, IDH3A, OGDH, SUCLAG2, etc.), and from glycolysis (ALDOA, ENO1, FBP1, GPI, PGK1, TALDO1, etc.) were distributed to both the nucleus and mitochondria. In contrast, proteins involved in nuclear/mitochondrial RNA processing/translation and Ras/Rab signaling showed different partitioning patterns. The identity of the OxPhos, TCA-cycle, and glycolysis proteins distributed to both the nucleus and mitochondria provides evidence for spatio-functional integration of these processes over the two different subcellular organelles. We suggest that there are unrecognized aspects of functional coordination between the nucleus and mitochondria, that integration of core functional processes via wide subcellular distribution of constituent proteins is a common characteristic of cells, and that subcellular spatial integration of function may be a vital aspect of cancer

Nuclear Cytoplasmic Trafficking of Proteins is a Major Response of Human Fibroblasts to Oxidative Stress

We have used a subcellular spatial razor approach based on LC–MS/MS-based proteomics with SILAC isotope labeling to determine changes in protein abundances in the nuclear and cytoplasmic compartments of human IMR90 fibroblasts subjected to mild oxidative stress. We show that response to mild <i>tert</i>-butyl hydrogen peroxide treatment includes redistribution between the nucleus and cytoplasm of numerous proteins not previously associated with oxidative stress. The 121 proteins with the most significant changes encompass proteins with known functions in a wide variety of subcellular locations and of cellular functional processes (transcription, signal transduction, autophagy, iron metabolism, TCA cycle, ATP synthesis) and are consistent with functional networks that are spatially dispersed across the cell. Both nuclear respiratory factor 2 and the proline regulatory axis appear to contribute to the cellular metabolic response. Proteins involved in iron metabolism or with iron/heme as a cofactor as well as mitochondrial proteins are prominent in the response. Evidence suggesting that nuclear import/export and vesicle-mediated protein transport contribute to the cellular response was obtained. We suggest that measurements of global changes in total cellular protein abundances need to be complemented with measurements of the dynamic subcellular spatial redistribution of proteins to obtain comprehensive pictures of cellular function

Subcellular Proteomics Reveals a Role for Nucleo-cytoplasmic Trafficking at the DNA Replication Origin Activation Checkpoint

Depletion of DNA replication initiation factors such as CDC7 kinase triggers the origin activation checkpoint in healthy cells and leads to a protective cell cycle arrest at the G1 phase of the mitotic cell division cycle. This protective mechanism is thought to be defective in cancer cells. To investigate how this checkpoint is activated and maintained in healthy cells, we conducted a quantitative SILAC analysis of the nuclear- and cytoplasmic-enriched compartments of CDC7-depleted fibroblasts and compared them to a total cell lysate preparation. Substantial changes in total abundance and/or subcellular location were detected for 124 proteins, including many essential proteins associated with DNA replication/cell cycle. Similar changes in protein abundance and subcellular distribution were observed for various metabolic processes, including oxidative stress, iron metabolism, protein translation and the tricarboxylic acid cycle. This is accompanied by reduced abundance of two karyopherin proteins, suggestive of reduced nuclear import. We propose that altered nucleo-cytoplasmic trafficking plays a key role in the regulation of cell cycle arrest. The results increase understanding of the mechanisms underlying maintenance of the DNA replication origin activation checkpoint and are consistent with our proposal that cell cycle arrest is an actively maintained process that appears to be distributed over various subcellular locations

Subcellular Proteomics Reveals a Role for Nucleo-cytoplasmic Trafficking at the DNA Replication Origin Activation Checkpoint

Depletion of DNA replication initiation factors such as CDC7 kinase triggers the origin activation checkpoint in healthy cells and leads to a protective cell cycle arrest at the G1 phase of the mitotic cell division cycle. This protective mechanism is thought to be defective in cancer cells. To investigate how this checkpoint is activated and maintained in healthy cells, we conducted a quantitative SILAC analysis of the nuclear- and cytoplasmic-enriched compartments of CDC7-depleted fibroblasts and compared them to a total cell lysate preparation. Substantial changes in total abundance and/or subcellular location were detected for 124 proteins, including many essential proteins associated with DNA replication/cell cycle. Similar changes in protein abundance and subcellular distribution were observed for various metabolic processes, including oxidative stress, iron metabolism, protein translation and the tricarboxylic acid cycle. This is accompanied by reduced abundance of two karyopherin proteins, suggestive of reduced nuclear import. We propose that altered nucleo-cytoplasmic trafficking plays a key role in the regulation of cell cycle arrest. The results increase understanding of the mechanisms underlying maintenance of the DNA replication origin activation checkpoint and are consistent with our proposal that cell cycle arrest is an actively maintained process that appears to be distributed over various subcellular locations

Subcellular Proteomics Reveals a Role for Nucleo-cytoplasmic Trafficking at the DNA Replication Origin Activation Checkpoint

Depletion of DNA replication initiation factors such as CDC7 kinase triggers the origin activation checkpoint in healthy cells and leads to a protective cell cycle arrest at the G1 phase of the mitotic cell division cycle. This protective mechanism is thought to be defective in cancer cells. To investigate how this checkpoint is activated and maintained in healthy cells, we conducted a quantitative SILAC analysis of the nuclear- and cytoplasmic-enriched compartments of CDC7-depleted fibroblasts and compared them to a total cell lysate preparation. Substantial changes in total abundance and/or subcellular location were detected for 124 proteins, including many essential proteins associated with DNA replication/cell cycle. Similar changes in protein abundance and subcellular distribution were observed for various metabolic processes, including oxidative stress, iron metabolism, protein translation and the tricarboxylic acid cycle. This is accompanied by reduced abundance of two karyopherin proteins, suggestive of reduced nuclear import. We propose that altered nucleo-cytoplasmic trafficking plays a key role in the regulation of cell cycle arrest. The results increase understanding of the mechanisms underlying maintenance of the DNA replication origin activation checkpoint and are consistent with our proposal that cell cycle arrest is an actively maintained process that appears to be distributed over various subcellular locations

Subcellular Proteomics Reveals a Role for Nucleo-cytoplasmic Trafficking at the DNA Replication Origin Activation Checkpoint

Depletion of DNA replication initiation factors such as CDC7 kinase triggers the origin activation checkpoint in healthy cells and leads to a protective cell cycle arrest at the G1 phase of the mitotic cell division cycle. This protective mechanism is thought to be defective in cancer cells. To investigate how this checkpoint is activated and maintained in healthy cells, we conducted a quantitative SILAC analysis of the nuclear- and cytoplasmic-enriched compartments of CDC7-depleted fibroblasts and compared them to a total cell lysate preparation. Substantial changes in total abundance and/or subcellular location were detected for 124 proteins, including many essential proteins associated with DNA replication/cell cycle. Similar changes in protein abundance and subcellular distribution were observed for various metabolic processes, including oxidative stress, iron metabolism, protein translation and the tricarboxylic acid cycle. This is accompanied by reduced abundance of two karyopherin proteins, suggestive of reduced nuclear import. We propose that altered nucleo-cytoplasmic trafficking plays a key role in the regulation of cell cycle arrest. The results increase understanding of the mechanisms underlying maintenance of the DNA replication origin activation checkpoint and are consistent with our proposal that cell cycle arrest is an actively maintained process that appears to be distributed over various subcellular locations