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Supplemental Information

Figure S1

Representative indents made during micromechanical testing for the (A) 400 µatm and (B) 2800 µatm treatment. The radius of a circle radiating from the center of the indent enclosing all visible cracks was used to calculate fracture toughness, a portion of which is shown for each treatment. Arrow denotes the longest crack found for each indent. Radius length is shown on the image in µm. Mean crack radius was similar between the 400 and 1000 µatm treatments.

DOI: 10.7287/peerj.preprints.388v1/supp-1

Figure S2

Glycogen content (µg glycogen per mg tissue) for oysters from the pCO2 treatments of 400, 800, and 2800 µatm. There is no difference in glycogen content among treatment groups.

DOI: 10.7287/peerj.preprints.388v1/supp-2

Figure S3

Representation of key metabolic pathways that are significantly affected by ocean acidification (A), mechanical stress at low pCO2 (B), and mechanical stress at high pCO2 (C). Red lines are proteins/pathways that are differentially expressed at higher levels in the stress treatments and blue lines are those that are differentially expressed at lower levels. In the key, the different colored lines represent different metabolic pathways that are affected by oyster exposure to ocean acidification and/or mechanical stimulation. Figures are also available on FigShare with input files for iPath2 to allow for interactive exploration of the data [88].

DOI: 10.7287/peerj.preprints.388v1/supp-3

Figure S4

Heat maps of differentially expressed proteins annotated with protein names. Protein expression values have been log-transformed. The dendrograms on the left of the heat maps represent the clustering of proteins according to expression profile.

DOI: 10.7287/peerj.preprints.388v1/supp-4

Table S1

Raw and normalized (proportion) fatty acid data for 8 oysters each from 3 treatments: 400, 800, and 2800 µatm.

DOI: 10.7287/peerj.preprints.388v1/supp-5

Table S2

ProteinProphet output for each technical replicate. Information for each protein includes percent coverage by sequenced peptides, total number unique peptides, total independent spectra (spectral count), and peptide sequences.

DOI: 10.7287/peerj.preprints.388v1/supp-6

Table S3

Protein expression values (NSAF) for each oyster for the 1 616 proteins identified. Also included are average expression values across treatments (i.e. 2800 avg NSAF is the average expression across all four high pCO2-exposed oysters); fold change for treatment/control oysters (i.e. Fold Diff OA is [2800 avg NSAF]/[400 avg NSAF]); columns for each of the three treatment comparisons with an asterisk indicating if the protein is >5-fold up- or down-regulated; SwissProt annotation, e-value, and gene description; proteins responsible for enrichment and the treatment comparisons in which they are enriched; a column indicating in which stress treatment proteins are differentially expressed (q-value < 0.1). In the fold difference columns “up” signifies that the protein was only expressed in oysters from the 2800 µatm treatment (versus the 400 µatm , Fold Diff OA), mechanical stress at 400 µatm treatment (versus 400 µatm, Fold Diff 400 MechS), or in the mechanical stress at 2800 µatm (versus 2800 µatm, Fold Diff 2800 MechS); “down” represents proteins that were only expressed in the other treatment for each comparison.

DOI: 10.7287/peerj.preprints.388v1/supp-7

Table S4

C. gigas proteins with associated SwissProt/UniProt-KB, Gene Ontology (GO), and GO Slim annotations.

DOI: 10.7287/peerj.preprints.388v1/supp-8

Table S5

Enriched biological processes for proteins >2-fold differentially expressed in the stress responses to elevated pCO2 of 2800 µatm (“OA”), mechanical stress after a one month exposure to 400 µatm (“Mech Stress 400 µatm”), and mechanical stress after a one month exposure to 2800 µtam (“Mech Stress 2800 µatm”). Table includes enriched GO term, number of proteins contributing to that GO term, p-value indicating degree of enrichment, the SwissProt accession numbers for those proteins, the fold enrichment for each GO term, and the false discovery rate (FDR).

DOI: 10.7287/peerj.preprints.388v1/supp-9

Additional Information

Competing Interests

The authors have no competing interests to declare.

Author Contributions

Emma Timmins-Schiffman conceived and designed the experiments, performed the experiments, analyzed the data, wrote the paper, prepared figures and/or tables, reviewed drafts of the paper.

William D Coffey performed the experiments, wrote the paper, reviewed drafts of the paper.

Wilber Hua performed the experiments, analyzed the data, reviewed drafts of the paper.

Brook L Nunn analyzed the data, contributed reagents/materials/analysis tools, wrote the paper, reviewed drafts of the paper.

Gary H Dickinson contributed reagents/materials/analysis tools, wrote the paper, prepared figures and/or tables, reviewed drafts of the paper.

Steven B Roberts conceived and designed the experiments, analyzed the data, contributed reagents/materials/analysis tools, wrote the paper, reviewed drafts of the paper.

Grant Disclosures

The following grant information was disclosed by the authors:

NOAA Saltonstall-Kennedy Program grant #NA09NMF4270093

Data Deposition

The following information was supplied regarding the deposition of related data:

ProteomeXchange #PXD000835


This research was funded in part by the National Oceanographic and Atmospheric Administration Saltonstall-Kennedy Program grant # NA09NMF4270093 to Dr. Steven Roberts, The College of New Jersey Mentored Undergraduate Summer Experience (MUSE) program, The University of Washington’s Proteomics Computer Resource Center (UWPR95794), as well as by contributions from 68 “fuelers” on RocketHub. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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