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. 2023 Feb 17;14(2):512.
doi: 10.3390/genes14020512.

Effect of 11-Deoxycorticosterone in the Transcriptomic Response to Stress in Rainbow Trout Skeletal Muscle

Rodrigo Zuloaga  1   2 , Daniela Aravena-Canales  1   2 , Jorge Eduardo Aedo  3 , Cesar Osorio-Fuentealba  4 , Alfredo Molina  1   2 , Juan Antonio Valdés  1   2
Affiliations

Effect of 11-Deoxycorticosterone in the Transcriptomic Response to Stress in Rainbow Trout Skeletal Muscle

Rodrigo Zuloaga et al. Genes (Basel). .

Abstract

In aquaculture, many stressors can negatively affect growth in teleosts. It is believed that cortisol performs glucocorticoid and mineralocorticoid functions because teleosts do not synthesize aldosterone. However, recent data suggest that 11-deoxycorticosterone (DOC) released during stress events may be relevant to modulate the compensatory response. To understand how DOC modifies the skeletal muscle molecular response, we carried out a transcriptomic analysis. Rainbow trout (Oncorhynchus mykiss) were intraperitoneally treated with physiological doses of DOC in individuals pretreated with mifepristone (glucocorticoid receptor antagonist) or eplerenone (mineralocorticoid receptor antagonist). RNA was extracted from the skeletal muscles, and cDNA libraries were constructed from vehicle, DOC, mifepristone, mifepristone plus DOC, eplerenone, and eplerenone plus DOC groups. The RNA-seq analysis revealed 131 differentially expressed transcripts (DETs) induced by DOC with respect to the vehicle group, mainly associated with muscle contraction, sarcomere organization, and cell adhesion. In addition, a DOC versus mifepristone plus DOC analysis revealed 122 DETs related to muscle contraction, sarcomere organization, and skeletal muscle cell differentiation. In a DOC versus eplerenone plus DOC analysis, 133 DETs were associated with autophagosome assembly, circadian regulation of gene expression, and regulation of transcription from RNA pol II promoter. These analyses indicate that DOC has a relevant function in the stress response of skeletal muscles, whose action is differentially modulated by GR and MR and is complementary to cortisol.

Keywords: DOC; RNA-seq; glucocorticoid receptor; mineralocorticoid receptor; skeletal muscle.

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Conflict of interest statement

The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.

Figures

Figure 1
Figure 1
DOC, cortisol, and glucose quantification in plasma. DOC (a), cortisol (b), and glucose (c) in plasma were quantified in juvenile fish treated with vehicle, 11-deoxycorticosterone (DOC), mifepristone, eplerenone, mifepristone plus DOC, and eplerenone plus DOC. The results are expressed as means ± SEM (n = 4 per treatment). The ** symbol (p < 0.01) represents significant differences.
Figure 2
Figure 2
Gene enrichment analysis of biological processes in vehicle and DOC DETs. The graph indicates the −log10(p-value) enriched BPs of differentially expressed transcripts between the vehicle and DOC groups with p-values < 0.05.
Figure 3
Figure 3
Gene enrichment analysis of biological processes in DOC and mifepristone plus DOC DETs. The graph indicates the −log10(p-values) of enriched BPs of differentially expressed transcripts between the DOC and mifepristone plus DOC groups with p-values < 0.05.
Figure 4
Figure 4
Gene enrichment analysis of biological processes in DOC and eplerenone plus DOC DETs. The graph indicates the −log10(p-values) of enriched BPs of differentially expressed transcripts from the DOC vs. eplerenone plus DOC groups with p-values < 0.05.
Figure 5
Figure 5
Venn diagram indicating the numbers of differentially expressed transcripts under the vehicle vs. DOC, DOC vs. mifepristone plus DOC, and DOC vs. eplerenone plus DOC treatments. Underlined transcripts were selected for RNA-seq validation by RT-qPCR.
Figure 6
Figure 6
RT-qPCR validation of differentially expressed transcripts. Transcripts selected for the RT-PCR validation of RNA-seq were mybpc1, septin10, strada, ppp3cc, myom2, tsc1, robo2, insr, rab9a, fos, and bach2. (a) Validation between vehicle and DOC; (b) validation between DOC and DOC plus mifepristone; (c) validation between DOC and DOC plus eplerenone. For RNA-seq, in purple, “#” indicates a log2 fold change ≥2.0 and FDR <0.05. For RT-qPCR, in blue, relative expression was normalized against fau and actβ, and “*” indicates significant differences in fold change from vehicle or DOC groups (mean ± SEM, n = 4, p < 0.05). Abbreviations: mybpc1, myosin-binding protein C1; strada, STE20-related kinase adapter protein α; ppp3cc, calcineurin; myom2, myomesin 2; tsc1, TSC complex subunit 1a; robo2, roundabout homolog 2; insr, insulin receptor; rab9a, ras-related protein Rab-9A; fos, proto-oncogene c-Fos; bach2, BTB domain and CNC homolog 2; fau, 40S ribosomal protein S30; actβ, β actin.
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Grants and funding

This research was funded by the Fondo Nacional de Desarrollo Científico y Tecnológico (FONDECYT) under grant numbers 1201498 (to Juan Antonio Valdés) and 3210050 (to Jorge Eduardo Aedo); the Fondo de Financiamiento de Centros de Investigación en Áreas Prioritarias (FONDAP) INCAR 15110027 grant; and the Concurso de Apoyo a Centros de Excelencia en Investigación FONDAP 2022 1522A0004 grant. Daniela Aravena-Canales acknowledges support received from ANID (Ph.D. scholarship 21220218). Rodrigo Zuloaga acknowledges support received from ANID (Ph.D. scholarship 21230070).

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