Protein expression patterns in zebrafish skeletal muscle: initial characterization and the effects of hypoxic exposure

Abstract

Patterns of protein expression were examined in white skeletal muscle from adult zebrafish (Danio rerio). High resolution two-dimensional gel electrophoresis resolved between 300 and 400 spots with molecular masses between 20 and 120 kDa and isoelectric points between about 5 and 8. Forty spots, representing a range of protein size, charge, and abundance were excised, digested with trypsin, and subjected to matrix-assisted laser-desorption/ionisation-time of flight mass spectrometry for protein identification. Twenty-nine spots were identified, including enzymes of energy metabolism, contractile proteins, an iron transport protein, and a heat shock protein. In addition, several spots matched theoretical proteins predicted from genome sequencing. These theoretical proteins were tentatively identified by similarity to known proteins. Patterns of muscle protein expression were then measured after zebrafish were exposed to low oxygen (16 torr) for 48 h, an exposure previously shown to increase the survival of zebrafish at more severe reductions in oxygen. Exposure to low oxygen (hypoxia) did not change the general pattern of protein expression but did affect the amounts of six low abundance proteins. The relatively subtle effects of hypoxia on patterns of muscle protein expression contrasts the widespread changes previously documented in mRNA levels in this and other species of fish during hypoxic stress. The difference between protein and mRNA expression illustrates the need to integrate both measures for a more complete understanding of gene expression in fish during hypoxic exposure.

Global cooling: Cold acclimation and the expression of soluble proteins in carp skeletal muscle

“…From the MALDI-TOF mass spectra obtained from these four spots, we identified the M2 and M3 CK isoforms. However, additional CK spots covering a mass range of 10-30 kDa were also identified across the charge dimension, a finding consistent with the skeletal muscle proteome of zebrafish [20].…”

“…The muscle samples were also analysed by 2-DE, immediately following homogenisation. Bosworth et al [20] observed Figure 6. Peptide coverage maps for the isoforms of CK isoforms and fragments.…”

Proteomic responses to environmentally induced oxidative stress

“…While these results point to an important role of GST during hypoxia, given the design of the experiment, it is not clear whether aerial exposure or subsequent re-immersion (re-oxygenation) caused the increase in antioxidant proteins. Studies on hypoxia-induced proteins in brain tissue of male medaka (Oryzias latipes) also showed an increase in GST (Oehlers et al, 2007), but brain and skeletal muscle in carp (Carasssius carassius) and zebrafish (Danio rerio), respectively, did not identify GST among proteins changing in response to anoxia/hypoxia (Bosworth et al, 2005;Smith et al, 2009). …”

Fundulus as the premier teleost model in environmental biology: Opportunities for new insights using genomics

“…In all three studies, hypoxia resulted in changes in the levels of numerous transcripts encoding proteins involved carbohydrate metabolism, protein synthesis, cell proliferation, and motility. In contrast, a proteomic study of zebrafish skeletal muscle revealed limited changes in protein expression after hypoxia (Bosworth et al, 2005). The difference between results of microarray and proteomic studies may be due to experimental design (levels and lengths of hypoxia) or biological differences (species, tissues, or developmental stages), or could be the result of uncoupling of mRNA and protein expression under hypoxia.…”