Synopsis The gill proteome of threespine sticklebacks (Gasterosteus aculeatus) differs greatly in populations that inhabit diverse environments characterized by different temperature, salinity, food availability, parasites, and other parameters. To assess the contribution of a specific environmental parameter to such differences it is necessary to isolate its effects from those of other parameters. In this study the effect of environmental salinity on the gill proteome of G. aculeatus was isolated in controlled mesocosm experiments. Salinity-dependent changes in the gill proteome were analyzed by Liquid chromatography/Tandem mass spectrometry data-independent acquisition (DIA) and Skyline. Relative abundances of 1691 proteins representing the molecular phenotype of stickleback gills were quantified using previously developed MSMS spectral and assay libraries in combination with DIA quantitative proteomics. Non-directional stress responses were distinguished from osmoregulatory protein abundance changes by their consistent occurrence during both hypo- and hyper-osmotic salinity stress in six separate mesocosm experiments. If the abundance of a protein was consistently regulated in opposite directions by hyper- versus hypo-osmotic salinity stress, then it was considered an osmoregulatory protein. In contrast, if protein abundance was consistently increased irrespective of whether salinity was increased or decreased, then it was considered a non-directional response protein. KEGG pathway analysis revealed that the salivary secretion, inositol phosphate metabolism, valine, leucine, and isoleucine degradation, citrate cycle, oxidative phosphorylation, and corresponding endocrine and extracellular signaling pathways contain most of the osmoregulatory gill proteins whose abundance is directly proportional to environmental salinity. Most proteins that were inversely correlated with salinity map to KEGG pathways that represent proteostasis, immunity, and related intracellular signaling processes. Non-directional stress response proteins represent fatty and amino acid degradation, purine metabolism, focal adhesion, mRNA surveillance, phagosome, endocytosis, and associated intracellular signaling KEGG pathways. These results demonstrate that G. aculeatus responds to salinity changes by adjusting osmoregulatory mechanisms that are distinct from transient non-directional stress responses to control compatible osmolyte synthesis, transepithelial ion transport, and oxidative energy metabolism. Furthermore, this study establishes salinity as a key factor for causing the regulation of numerous proteins and KEGG pathways with established functions in proteostasis, immunity, and tissue remodeling. We conclude that the corresponding osmoregulatory gill proteins and KEGG pathways represent molecular phenotypes that promote transepithelial ion transport, cellular osmoregulation, and gill epithelial remodeling to adjust gill function to environmental salinity.
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Effects of pejus and pessimum zone salinity stress on gill proteome networks and energy homeostasis in Oreochromis mossambicus
Salinity tolerance in fish involves a suite of physiological changes, but a cohesive theory leading to a mechanistic understanding at the organismal level is lacking. To examine the potential of adapting energy homeostasis theory in the context of salinity stress in teleost fish,
- NSF-PAR ID:
- 10441972
- Publisher / Repository:
- Wiley Blackwell (John Wiley & Sons)
- Date Published:
- Journal Name:
- PROTEOMICS
- Volume:
- 24
- Issue:
- 1-2
- ISSN:
- 1615-9853
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Abstract Interactions of organisms with their environment are complex and environmental regulation at different levels of biological organization is often nonlinear. Therefore, the genotype to phenotype continuum requires study at multiple levels of organization. While studies of transcriptome regulation are now common for many species, quantitative studies of environmental effects on proteomes are needed. Here we report the generation of a data‐independent acquisition (DIA) assay library that enables simultaneous targeted proteomics of thousands of
Oreochromis niloticus kidney proteins using a label‐ and gel‐free workflow that is well suited for ecologically relevant field samples. We demonstrate the usefulness of this DIA assay library by discerning environmental effects on the kidney proteome ofO. niloticus . Moreover, we demonstrate that the DIA assay library approach generates data that are complimentary rather than redundant to transcriptomic data. Transcript and protein abundance differences in kidneys of tilapia acclimated to freshwater and brackish water (25 g/kg) were correlated for 2114 unique genes. A high degree of non‐linearity in salinity‐dependent regulation of transcriptomes and proteomes was revealed suggesting that the regulation ofO .niloticus renal function by environmental salinity relies heavily on post‐transcriptional mechanisms. The application of functional enrichment analyses using STRING and KEGG to DIA assay data sets is demonstrated by identifyingmyo ‐inositol metabolism, antioxidant and xenobiotic functions, and signalling mechanisms as key elements controlled by salinity in tilapia kidneys. The DIA assay library resource presented here can be adopted for other tissues and other organisms to study proteome dynamics during changing ecological contexts. -
Organismal physiology, morphology, and behavior are based on the function of structural proteins and enzymes. Proteins represent the central regulatory plane in the genome to phenome continuum. The protein complement of cells and tissues (the proteome) is highly dynamic and mirrors environmental and developmental influences on organismal phenotypes. Therefore, dynamic proteomes are excellent bioindicators of environmental exposure. Comprehensive blueprints of environmental exposures are reflected in specific proteome states and capturing those states is achieved by quantitative proteomics. We have developed quantitative proteomics workflows to characterize environmental influences on proteome states and proteome dynamics of euryhaline and euryhthermal fish populations in coastal areas. These workflows utilize tissue- and cell-specific assay libraries for data-independent acquisition (DIA) or Sequentially Windowed Acquisition of all THeoretically possible MSMS spectra (SWATH) mass spectrometry. Qunatitative proteome datasets generated with these workflows are highly accurate and they consistently cover precisely defined sets of proteomes. This consistent coverage renders systematic and long-term network and topological data analysis (TDA) approaches feasible. These workflows and approaches are explained and their application to coastal fish biology is discussed using selected datasets as examples. The data presented illustrate that habitat differences such as salinity and temperature changes are readily captured in state changes of tissue-specific proteomes. The overall topology of proteome states is indicative of particular tissues, species, and environmental contexts and is therefore suitable for deducing functional and phenotypic consequences of environmental changes on coastal organisms.more » « less
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Summary Cu+‐chaperones are a diverse group of proteins that allocate Cu+ions to specific copper proteins, creating different copper pools targeted to specific physiological processes.
Symbiotic nitrogen fixation carried out in legume root nodules indirectly requires relatively large amounts of copper, for example for energy delivery via respiration, for which targeted copper deliver systems would be required.
MtNCC1 is a nodule‐specific Cu+‐chaperone encoded in the
Medicago truncatula genome, with a N‐terminus Atx1‐like domain that can bind Cu+with picomolar affinities. MtNCC1 is able to interact with nodule‐specific Cu+‐importer MtCOPT1.MtNCC1 is expressed primarily from the late infection zone to the early fixation zone and is located in the cytosol, associated with plasma and symbiosome membranes, and within nuclei. Consistent with its key role in nitrogen fixation,ncc1 mutants have a severe reduction in nitrogenase activity and a 50% reduction in copper‐dependent cytochromec oxidase activity.A subset of the copper proteome is also affected in the
ncc1 mutant nodules. Many of these proteins can be pulled down when using a Cu+‐loaded N‐terminal MtNCC1 moiety as a bait, indicating a role in nodule copper homeostasis and in copper‐dependent physiological processes. Overall, these data suggest a pleiotropic role of MtNCC1 in copper delivery for symbiotic nitrogen fixation.
