Dataset Information

Reversible oxidative signaling in Chlamydomonas following nitrogen deprivation

ABSTRACT: As global temperatures climb to historic highs, the far-reaching effects of climate change have impacted agricultural nutrient availability. This has extended to low latitude oceans, where a deficit in both nitrogen and phosphorus stores has led to dramatic decreases in carbon sequestration in oceanic phytoplankton. Although Chlamydomonas reinhardtii, a freshwater model green alga, has shown drastic systems-level alterations following nitrogen deprivation, the mechanism(s) through which these alterations are triggered and regulated are unclear. This study examined the role of reversible oxidative signaling in the nitrogen stress response of C. reinhardtii. Using oxidized cysteine resin-assisted capture enrichment coupled with label-free quantitative proteomics, 7,889 unique oxidized cysteine thiol identifiers were quantified, with 231 significantly changing peptides from 184 proteins following 2 h of nitrogen deprivation. These results demonstrate proteins affected by oxidation involved in nitrogen assimilation, amino acid metabolism, photosynthesis, and other metabolic processes. An enhanced role of non-damaging oxidative pathways is observed throughout the photosynthetic apparatus that provides a framework for further analysis in phototrophs.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Chlamydomonas Reinhardtii

TISSUE(S): Photosynthetic Cell

SUBMITTER: Leslie Hicks

LAB HEAD: Leslie M. Hicks

PROVIDER: PXD019491 | Pride | 2020-06-28

REPOSITORIES: Pride

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Publications

Photosynthetic Metabolism and Nitrogen Reshuffling Are Regulated by Reversible Cysteine Thiol Oxidation Following Nitrogen Deprivation in Chlamydomonas.

Smythers Amanda L AL McConnell Evan W EW Lewis Hailey C HC Mubarek Saher N SN Hicks Leslie M LM

Plants (Basel, Switzerland) 20200623 6

As global temperatures climb to historic highs, the far-reaching effects of climate change have impacted agricultural nutrient availability. This has extended to low latitude oceans, where a deficit in both nitrogen and phosphorus stores has led to dramatic decreases in carbon sequestration in oceanic phytoplankton. Although <i>Chlamydomonas reinhardtii</i>, a freshwater model green alga, has shown drastic systems-level alterations following nitrogen deprivation, the mechanisms through which the ...[more]

PMID: 32585825

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Project description:We performed chromatin immunoprecipitation on microarray (ChIP-chip) transcriptional profiling of the SsrB regulator of Salmonella enterica Typhimurium to better characterize its regulon and to identify the DNA-recognition element coordinating its specific interaction at cis-regulatory sites. SsrB, the response regulator of the two component regulatory system SsrA-SsrB encoded within the Salmonella Pathogenicity Island (SPI-2), directs transcriptional activation of the closely associated type three secretion system (T3SS) also encoded at this locus. Immunoprecipitaiton of SsrB translationally fused to a C-terminal FLAG-tag from formaldehyde cross-linked genomic DNA was performed under SPI-2 activating and non-activating conditions. Downstream analysis and experimental work identified specific interactions within SPI-2 at the previously identified ssrA, ssaB, sseA, ssaG, ssaM promoters and identified an additional cryptic promoter driving expression upstream of ssaR. Additionally, a previously unknown SsrB interaction site within ssaE was shown to be the major contributor driving the expression of the downstream genes and not the previously identified interaction site within the intergenic region of ssaE-sseA. Interactions were also identified outside of SPI-2 upstream of other T3SS-associated genes encoded within other genomic islands, and for other uncharacterized genes. Integration of this data with transcriptional microarray work, previously published DNase I footprinting data, bacteria 1-hybrid investigations and comparative genomics analyses of cis-regulatory regions within the orthologous Sodalis symbiosis region 3 (SSR-3) of Sodalis glossinidius enabled identification of a conserved 18bp palindrome which was experimentally validated as being required for transcriptional activation of SsrB dependant genes. SsrB-FLAG immunoprecipitations were performed under SsrB activating and non-activating conditions from formaldehyde cross-linked bacterial lysates. Nine immunoprecipitation reactions were pooled into three replicate samples for the activating condition and three reactions were pooled into one sample for the non-activating control condition. The immunoprecipitated DNA in addition to the non-immunoprecipitated control DNA for each of the four samples were labeled with Cy3 or Cy5 fluorophores respectively and were concurrently hybridized to four arrays on a single slide.

Dataset Information

Reversible oxidative signaling in Chlamydomonas following nitrogen deprivation

Publications

Photosynthetic Metabolism and Nitrogen Reshuffling Are Regulated by Reversible Cysteine Thiol Oxidation Following Nitrogen Deprivation in Chlamydomonas.

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