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Title: The stringent response regulates adaptation to darkness in the cyanobacterium Synechococcus elongatus

Authors:
; ; ; ; ORCiD logo
Publication Date:
Sponsoring Org.:
USDOE
OSTI Identifier:
1281325
Grant/Contract Number:
SC0006394
Resource Type:
Journal Article: Published Article
Journal Name:
Proceedings of the National Academy of Sciences of the United States of America
Additional Journal Information:
Journal Volume: 113; Journal Issue: 33; Related Information: CHORUS Timestamp: 2017-06-24 15:48:28; Journal ID: ISSN 0027-8424
Publisher:
Proceedings of the National Academy of Sciences
Country of Publication:
United States
Language:
English

Citation Formats

Hood, Rachel D., Higgins, Sean A., Flamholz, Avi, Nichols, Robert J., and Savage, David F.. The stringent response regulates adaptation to darkness in the cyanobacterium Synechococcus elongatus. United States: N. p., 2016. Web. doi:10.1073/pnas.1524915113.
Hood, Rachel D., Higgins, Sean A., Flamholz, Avi, Nichols, Robert J., & Savage, David F.. The stringent response regulates adaptation to darkness in the cyanobacterium Synechococcus elongatus. United States. doi:10.1073/pnas.1524915113.
Hood, Rachel D., Higgins, Sean A., Flamholz, Avi, Nichols, Robert J., and Savage, David F.. 2016. "The stringent response regulates adaptation to darkness in the cyanobacterium Synechococcus elongatus". United States. doi:10.1073/pnas.1524915113.
@article{osti_1281325,
title = {The stringent response regulates adaptation to darkness in the cyanobacterium Synechococcus elongatus},
author = {Hood, Rachel D. and Higgins, Sean A. and Flamholz, Avi and Nichols, Robert J. and Savage, David F.},
abstractNote = {},
doi = {10.1073/pnas.1524915113},
journal = {Proceedings of the National Academy of Sciences of the United States of America},
number = 33,
volume = 113,
place = {United States},
year = 2016,
month = 8
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record at 10.1073/pnas.1524915113

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  • An active photosystem I (PSI) complex was isolated from the thermophilic cyanobacterium Synechococcus elongatus by a procedure consisting of three steps: First, extraction of photosystem II from the thylakoids by a sulfobetaine detergent yields PSI-enriched membranes. Second, the latter are treated with Triton X-100 to extract PSI particles, which are further purified by preparative isoelectric focusing. Third, anion-exchange chromatography is used to remove contaminating phycobilisome polypeptides. The purified particles show three major bands in sodium dodecyl sulfate gel electrophoresis of apparent molecular mass of 110, 15, and 10 kDa. Charge separation was monitored by the kinetics of flash-induced absorption changesmore » at 820 nm. A chlorophyll/P700 ratio of 60 was found. When the particles are stored at 4 degrees C, charge separation was stable for weeks. The molecular mass of the PSI particles, determined by measurement of zero-angle neutron scattering intensity, was 217,000 Da. The PSI particles thus consist of one heterodimer of the 60-80-kDa polypeptides and presumably one copy of the 15- and 10-kDa polypeptides, respectively.« less
  • As autotrophic prokaryotes, cyanobacteria are ideal chassis organisms for sustainable production of various useful compounds. The newly characterized cyanobacterium Synechococcus elongatus UTEX 2973 is a promising candidate for serving as a microbial cell factory because of its unusually rapid growth rate. Here, we seek to develop a genetic toolkit that enables extensive genomic engineering of Synechococcus 2973 by implementing a CRISPR/Cas9 editing system. We targeted the nblA gene because of its important role in biological response to nitrogen deprivation conditions. First, we determined that the Streptococcus pyogenes Cas9 enzyme is toxic in cyanobacteria, and conjugational transfer of stable, replicating constructsmore » containing the cas9 gene resulted in lethality. However, after switching to a vector that permitted transient expression of the cas9 gene, we achieved markerless editing in 100 % of cyanobacterial exconjugants after the first patch. Moreover, we could readily cure the organisms of antibiotic resistance, resulting in a markerless deletion strain. In conclusion, high expression levels of the Cas9 protein in Synechococcus 2973 appear to be toxic and result in cell death. However, introduction of a CRISPR/Cas9 genome editing system on a plasmid backbone that leads to transient cas9 expression allowed for efficient markerless genome editing in a wild type genetic background.« less
  • In this investigation, changes were characterized in cell structure and cytoplasmic membrane organization that occur when the freshwater cyanobacterium Synechococcus 6311 is transferred from low salt (0.03 molar NaCl) to high salt (0.5 molar NaCl) media (i.e. sea water concentration). Cells were examined at several time points after the imposition of the salt stress and compared to control cells, in thin sections and freeze fracture electron microscopy, and by flow cytometry. One minute after exposure to high salt, i.e. salt shock, virtually all intracellular granules disappeared, the density of the cytoplasm decreased, and the appearance of DNA material was changed.more » Glycogen and other granules, however, reappeared by 4 hours after salt exposure. The organization of the cytoplasmic membrane undergoes major reorganization following salt shock. Freeze-fracture electron microscopy showed that small intramembrane particles (diameters 7.5 and 8.5 nanometers) are reduced in number by two- to fivefold, whereas large particles, (diameters 14.5 and 17.5 nanometers) increase two- to fourfold in frequency, compared to control cells grown in low salt medium. The changes in particle size distribution suggest synthesis of new membrane proteins, in agreement with the known increases in respiration, cytochrome oxidase, and sodium proton exchange activity of the cytoplasmic membrane.« less
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