Development of a Gill Assay Library for Ecological Proteomics of Threespine Sticklebacks (Gasterosteus aculeatus)

Abstract

A data-independent acquisition (DIA) assay library for quantitative analyses of proteome dynamics has been developed for gills of threespine sticklebacks (Gasterosteus aculeatus). A raw spectral library was generated by data-dependent acquisition (DDA) and annotation of tryptic peptides to MSMS spectra and protein database identifiers. The assay library was constructed from the raw spectral library by removal of low-quality, ambiguous, and low-signal peptides. Only unique proteins represented by at least two peptides are included in the assay library, which consists of 1506 proteins, 5074 peptides, 5104 precursors, and 25,322 transitions. This assay library was used with DIA data to identify biochemical differences in gill proteomes of four populations representing different eco- and morpho-types of threespine sticklebacks. The assay library revealed unique and reproducible proteome signatures. Warm-adapted, low-plated, brackish-water fish from Laguna de la Bocana del Rosario (Mexico) show elevated HSP47, extracellular matrix, and innate immunity proteins whereas several immunoglobulins, interferon-induced proteins, ubiquitins, proteolytic enzymes, and nucleic acid remodeling proteins are reduced. Fully-plated, brackish-water fish from Westchester Lagoon (Alaska) display elevated ion regulation, GTPase signaling, and contractile cytoskeleton proteins, altered abundances of many ribosomal, calcium signaling and immunity proteins, and depleted transcriptional regulators and metabolic enzymes. Low-plated freshwater fish from Lake Solano (California) have elevated inflammasomes and proteolytic proteins whereas several iron containing and ion regulatory proteins are reduced. Gills of fully-plated, marine fish from Bodega Harbor (California) have elevated oxidative metabolism enzymes and reduced transglutaminase 2, collagens, and clathrin heavy chains. These distinct proteome signatures represent targets for testing ecological and evolutionary influences on molecular mechanisms of gill function in threespine sticklebacks. Furthermore, the gill assay library represents a model for other tissues and paves the way for accurate and reproducible network analyses of environmental context-dependent proteome dynamics in complex organisms.

Identification of key proteins involved in stickleback environmental adaption with system-level analysis

“…With the publication of the protein-abundance data 1 , the authors identified proteins with elevated and reduced abundance levels using univariate statistics and separately testing each protein for differences in abundance in one population against the others. This basic set of tests identified proteins that were present with significantly elevated or reduced abundance in a specific population compared to the other populations, and sets of these proteins that were enriched for biological functions specific for a fish environment.…”

“…Here, we present a system-level data-driven analysis to shed light on this question using a recent large-scale proteomics dataset from three-spine stickleback gills 1 . The authors conducted measurements on a total of 96 fish, consisting of groups of 24 fish sampled from four separate locations characterized by the following environmental parameters: cold, brackish water (BW) in Westchester Lagoon, Alaska (AK), cold salt water (SW) in Bodega Harbor, California (BH), cold freshwater (FW) in Lake Solano, California (LS) and warm brackish water in Laguna de la Bocana del Rosario, Mexico (RE).…”

Proteomic characterization and comparison of ram (Ovis aries) and buck (Capra hircus) spermatozoa proteome using a data independent acquisition mass spectometry (DIA-MS) approach

“…ALDH is an enzyme, which has responsibility for the removing 4-hydroxynonenal adducts from lipid membranes, and a strong correlation between ALDH2 expression and various motility parameters of stallion spermatozoa were reported [46]. A higher abundant of ALDH2 in spermatozoa were also observed with high reproductive efficiency of Meishan boar [19]. To date, literature on the relative of ALDH2 expression with sperm functionality in buck is paucity, which is worthy of further exploration.…”

“…Because of simultaneous high-throughput scanning and identifying all peptides within a given mass range, data-independent acquisition-mass spectrometry (DIA-MS) provides the possibility of overcoming limitations related to data dependent acquisition (DDA) [16]. As a highly promising MS-acquisition method, DIA has been successful used to investigate proteomic changes in plasma [17], cell [18], and tissue [19] in recent years.…”

Chronic temperature stress effects on the liver proteome of two threespine stickleback (Gasterosteus aculeatus) populations using a novel DIA assay library

“…The copyright holder for this preprint this version posted June 22, 2021. ; https://doi.org/10.1101/2021.06.21.449281 doi: bioRxiv preprint reproducible identification and quantification of peptides, including lower abundance precursor ions, and greatly increases the number of proteins that can be reproducibly quantified (19).…”