scholarly journals Proximity-based proteomics reveals the thylakoid lumen proteome in the cyanobacterium Synechococcus sp. PCC 7002

2020 ◽  
Author(s):  
Kelsey K. Dahlgren ◽  
Colin Gates ◽  
Thomas Lee ◽  
Jeffrey C. Cameron
Keyword(s):  
Photosystem Ii ◽  
Cell Biology ◽  
Region Of Interest ◽  
Protein Labeling ◽  
Spatiotemporal Resolution ◽  
Thylakoid Lumen ◽  
Conventional Fractionation ◽  
Great Progress ◽  
Biochemical Fractionation ◽  
Synechococcus Sp

AbstractCyanobacteria possess unique intracellular organization. Many proteomic studies have examined different features of cyanobacteria to learn about the intracellular structures and their respective functions. While these studies have made great progress in understanding cyanobacterial physiology, the conventional fractionation methods used to purify cellular structures have limitations; specifically, certain regions of cells cannot be purified with existing fractionation methods. Proximity-based proteomics techniques were developed to overcome the limitations of biochemical fractionation for proteomics. Proximity-based proteomics relies on spatiotemporal protein labeling followed by mass spectrometry of the labeled proteins to determine the proteome of the region of interest. We performed proximity-based proteomics in the cyanobacterium Synechococcus sp. PCC 7002 with the APEX2 enzyme, an engineered ascorbate peroxidase. We determined the proteome of the thylakoid lumen, a region of the cell that has remained challenging to study with existing methods, using a translational fusion between APEX2 and PsbU, a lumenal subunit of photosystem II. Our results demonstrate the power of APEX2 as a tool to study the cell biology of intracellular features and processes, including photosystem II assembly in cyanobacteria, with enhanced spatiotemporal resolution.

scholarly journals Proximity-based proteomics reveals the thylakoid lumen proteome in the cyanobacterium Synechococcus sp. PCC 7002

2020 ◽  
Author(s):  
Kelsey K. Dahlgren ◽  
Colin Gates ◽  
Thomas Lee ◽  
Jeffrey C. Cameron
Keyword(s):  
Cell Biology ◽  
Gene Fusion ◽  
Region Of Interest ◽  
Protein Labeling ◽  
Cellular Structures ◽  
Spatiotemporal Resolution ◽  
Thylakoid Lumen ◽  
Great Progress ◽  
Biochemical Fractionation ◽  
Synechococcus Sp

AbstractCyanobacteria possess unique intracellular organization. Many proteomic studies have examined different features of cyanobacteria to learn about the structure-function relationships between the intracellular structures of cyanobacteria and their roles in cells. While these studies have made great progress in understanding cyanobacterial physiology, the previous fractionation methods used to purify cellular structures have limitations; specifically, certain regions of cells cannot be purified with existing fractionation methods. Proximity-based proteomics techniques were developed to overcome the limitations of biochemical fractionation for proteomics. Proximity-based proteomics relies on spatiotemporal protein labeling followed by mass spectrometry of the labeled proteins to determine the proteome of the region of interest. We have performed proximity-based proteomics in the cyanobacterium Synechococcus sp. PCC 7002 with the APEX2 enzyme, an engineered ascorbate peroxidase. We determined the proteome of the thylakoid lumen, a region of the cell that has remained challenging to study with existing methods, using a PsbU-APEX2 gene fusion. This study demonstrates the power of APEX2 as a tool to study the cell biology of intracellular features of cyanobacteria with enhanced spatiotemporal resolution.

Positional Correlative Anatomy of Invertebrate Model Organisms Increases Efficiency of TEM Data Production

2014 ◽  
Vol 20 (5) ◽  
pp. 1392-1403 ◽  
Author(s):  
Irina Kolotuev
Keyword(s):  
Developmental Biology ◽  
Cell Biology ◽  
Sampling Rate ◽  
Unmet Need ◽  
Region Of Interest ◽  
Model Organisms ◽  
Data Generation ◽  
Step Method ◽  
Efficient Data ◽  
Research Questions

AbstractTransmission electron microscopy (TEM) is an important tool for studies in cell biology, and is essential to address research questions from bacteria to animals. Recent technological innovations have advanced the entire field of TEM, yet classical techniques still prevail for most present-day studies. Indeed, the majority of cell and developmental biology studies that use TEM do not require cutting-edge methodologies, but rather fast and efficient data generation. Although access to state-of-the-art equipment is frequently problematic, standard TEM microscopes are typically available, even in modest research facilities. However, a major unmet need in standard TEM is the ability to quickly prepare and orient a sample to identify a region of interest. Here, I provide a detailed step-by-step method for a positional correlative anatomy approach to flat-embedded samples. These modifications make the TEM preparation and analytic procedures faster and more straightforward, supporting a higher sampling rate. To illustrate the modified procedures, I provide numerous examples addressing research questions in Caenorhabditis elegans and Drosophila. This method can be equally applied to address questions of cell and developmental biology in other small multicellular model organisms.

Stabilization of the oxygen-evolving complex of photosystem II bybicarbonate and glycinebetaine in thylakoid and subthylakoidpreparations

2003 ◽  
Vol 30 (7) ◽  
pp. 797 ◽  
Author(s):  
Vyacheslav V. Klimov ◽  
Suleyman I. Allakhverdiev ◽  
Yoshitaka Nishiyama ◽  
AndreiA. Khorobrykh ◽  
Norio Murata
Keyword(s):  
Photosystem Ii ◽  
Protective Effect ◽  
Thermal Inactivation ◽  
Oxygen Evolving Complex ◽  
Photosynthetic Oxygen Evolution ◽  
Protective Effects ◽  
Synechococcus Sp ◽  
Oxygen Evolving ◽  
Triton X ◽  
Water Oxidizing Complex

The protective effect of 1 M glycinebetaine on thermal inactivation of photosynthetic oxygen evolution in isolated photosystem II membrane fragments from spinach is observed in CO2-free medium in both the presence and absence of added 2 mM bicarbonate. Conversely, the protective effect of 2 mM bicarbonate against thermoinactivation is seen in the absence as well as in the presence of 1 M glycinebetaine. The stabilizing effect of bicarbonate is also observed in thylakoid membranes from Synechococcus sp. PCC 7002 treated with 0.1% Triton X-100, and in unbroken spinach thylakoids. It is shown for the first time that bicarbonate protects the water-oxidizing complex against inactivation induced by pre-incubation of photosystem II membrane fragments (25°C) and thylakoids (40°C) at low pH (5.0–5.5) in non-bicarbonate-depleted medium. We conclude that the protective effects of glycinebetaine and bicarbonate are of a different nature; glycinebetaine acts as a non-specific, compatible, zwitterionic osmolyte while bicarbonate is considered an essential constituent of the water-oxidizing complex of photosystem II, important for its functioning and stabilization.

scholarly journals The Thylakoid Lumen Protease Deg1 Is Involved in the Repair of Photosystem II from Photoinhibition in Arabidopsis

2007 ◽  
Vol 19 (3) ◽  
pp. 1039-1047 ◽  
Author(s):  
Einat Kapri-Pardes ◽  
Leah Naveh ◽  
Zach Adam
Keyword(s):  
Photosystem Ii ◽  
Thylakoid Lumen

scholarly journals The autophagy protein Ambra1 regulates gene expression by supporting novel transcriptional complexes

2020 ◽  
Vol 295 (34) ◽  
pp. 12045-12057
Author(s):  
Christina Schoenherr ◽  
Adam Byron ◽  
Billie Griffith ◽  
Alexander Loftus ◽  
Jimi C. Wills ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cancer Cell ◽  
Cell Invasion ◽  
Cell Biology ◽  
Protein Complexes ◽  
Transcriptional Regulators ◽  
Adaptor Proteins ◽  
Nuclear Pore ◽  
Biochemical Fractionation

Ambra1 is considered an autophagy and trafficking protein with roles in neurogenesis and cancer cell invasion. Here, we report that Ambra1 also localizes to the nucleus of cancer cells, where it has a novel nuclear scaffolding function that controls gene expression. Using biochemical fractionation and proteomics, we found that Ambra1 binds to multiple classes of proteins in the nucleus, including nuclear pore proteins, adaptor proteins such as FAK and Akap8, chromatin-modifying proteins, and transcriptional regulators like Brg1 and Atf2. We identified biologically important genes, such as Angpt1, Tgfb2, Tgfb3, Itga8, and Itgb7, whose transcription is regulated by Ambra1-scaffolded complexes, likely by altering histone modifications and Atf2 activity. Therefore, in addition to its recognized roles in autophagy and trafficking, Ambra1 scaffolds protein complexes at chromatin, regulating transcriptional signaling in the nucleus. This novel function for Ambra1, and the specific genes impacted, may help to explain the wider role of Ambra1 in cancer cell biology.

scholarly journals Digital Spindle: A New Way to Explore Mitotic Functions by Whole Cell Data Collection and a Computational Approach

Cells ◽  
2020 ◽  
Vol 9 (5) ◽  
pp. 1255
Author(s):  
Norio Yamashita ◽  
Masahiko Morita ◽  
Hideo Yokota ◽  
Yuko Mimori-Kiyosue
Keyword(s):  
Cell Biology ◽  
Information Science ◽  
Three Dimensional ◽  
Light Sheet ◽  
Live Imaging ◽  
Three Dimensions ◽  
Complex Data ◽  
Spatiotemporal Resolution ◽  
Whole Cell ◽  
Light Sheet Microscopy

From cells to organisms, every living system is three-dimensional (3D), but the performance of fluorescence microscopy has been largely limited when attempting to obtain an overview of systems’ dynamic processes in three dimensions. Recently, advanced light-sheet illumination technologies, allowing drastic improvement in spatial discrimination, volumetric imaging times, and phototoxicity/photobleaching, have been making live imaging to collect precise and reliable 3D information increasingly feasible. In particular, lattice light-sheet microscopy (LLSM), using an ultrathin light-sheet, enables whole-cell 3D live imaging of cellular processes, including mitosis, at unprecedented spatiotemporal resolution for extended periods of time. This technology produces immense and complex data, including a significant amount of information, raising new challenges for big image data analysis and new possibilities for data utilization. Once the data are digitally archived in a computer, the data can be reused for various purposes by anyone at any time. Such an information science approach has the potential to revolutionize the use of bioimage data, and provides an alternative method for cell biology research in a data-driven manner. In this article, we introduce examples of analyzing digital mitotic spindles and discuss future perspectives in cell biology.

scholarly journals Mobilization of Photosystem II Induced by Intense Red Light in the Cyanobacterium Synechococcus sp PCC7942

2005 ◽  
Vol 18 (2) ◽  
pp. 457-464 ◽  
Author(s):  
Mary Sarcina ◽  
Nikolaos Bouzovitis ◽  
Conrad W. Mullineaux
Keyword(s):  
Photosystem Ii ◽  
Red Light ◽  
Synechococcus Sp

scholarly journals Immunological studies on the D-1 and D-2 proteins of photosystem II preparations from the thermophilic cyanobacterium, Synechococcus sp

FEBS Letters ◽  
1988 ◽  
Vol 233 (1) ◽  
pp. 1-6 ◽  
Author(s):  
Kazuhiko Satoh ◽  
Ralf Dostatni ◽  
Udo Johanningmeier ◽  
Walter Oettmeier
Keyword(s):  
Photosystem Ii ◽  
Thermophilic Cyanobacterium ◽  
Synechococcus Sp
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