Regulation of theRana sylvaticabrevinin-1SY antimicrobial peptide during development and in dorsal and ventral skin in response to freezing, anoxia, and dehydration

Abstract

Brevinin-1SY is the only described antimicrobial peptide (AMP) of Rana sylvatica. As AMPs are important innate immune molecules that inhibit microbes, this study examined brevinin-1SY regulation during development and in adult frogs in response to environmental stress. The brevinin-1SY nucleotide sequence was identified and used for protein modeling. Brevinin-1SY was predicted to be an amphipathic, hydrophobic, alpha helical peptide that inserts into a lipid bilayer. Brevinin-1SY transcripts were detected in tadpoles and were significantly increased during the later stages of development. Effects of environmental stress (24 h anoxia, 40% dehydration or 24 h frozen) on the mRNA levels of brevinin-1SY in the dorsal and ventral skin were examined. The brevinin-1SY mRNA levels were increased in dorsal and ventral skin of dehydrated frogs, and in ventral skin of anoxic frogs, compared with controls (non-stressed). Brevinin-1SY protein levels in peptide extracts of dorsal skin showed a similar, but not significant, trend to that of brevinin-1SY mRNA levels. Antimicrobial activity of skin extracts from control and stressed animals were assessed for Escherichia coli, Bacillus subtilis, Saccharomyces cerevisiae, Botrytis cinerea, Rhizopus stolonifer and Pythium sulcatum using disk diffusion assays. Peptide extracts of dorsal skin from anoxic, frozen and dehydrated animals showed significantly higher inhibition of E. coli and P. sulcatum than from control animals. In ventral skin peptide extracts, significant growth inhibition was observed in frozen animals for E. coli and P. sulcatum, and in anoxic animals for B. cinerea, compared with controls. Environmental regulation of brevinin-1SY may have important implications for defense against pathogens.

Using “Omics” and Integrated Multi-Omics Approaches to Guide Probiotic Selection to Mitigate Chytridiomycosis and Other Emerging Infectious Diseases

“…However, apparent gene duplications allow for gradual genetic changes that appear to be positively selected in response to pathogens ( Tennessen and Blouin, 2007 ). Defensive AMPs appear to be released constitutively into the mucus at low concentrations, but they can be increased significantly when the amphibian hosts are alarmed or injured ( Pask et al, 2012 ) and can be affected by environmental stressors ( Katzenback et al, 2014 ). However, some amphibians appear to lack the capacity to produce conventional cationic AMPs ( Conlon, 2011 ).…”

Life history linked to immune investment in developing amphibians

“…Some studies used transmission electron microscopy to examine discharge of granular glands (Delfino et al ., 2006). Other studies used northern blotting, in situ hybridization or RT-PCR to detect mRNAs encoding AMPs (Clark et al ., 1994; Reilly et al ., 1994; Iwamuro et al ., 2006; Katzenback et al ., 2014). As none of these studies demonstrated secretion of active peptides from living tadpoles, we chose to use direct MALDI-TOF MS to examine AMP expression on tadpoles (Chaurand et al ., 1999; Woodhams et al ., 2007).…”

Effects of anoxic exposure on the nuclear factor of activated T cell (NFAT) transcription factors in the stress‐tolerant wood frog

“…We have previously shown that wood frogs can tolerate up to 48 hours of anoxia exposure at 5°C with no mortality . In addition, previous studies have investigated the effects of exposure to 24‐hour anoxia and 4 hours of aerobic recovery on metabolic rate depression in different wood frog tissues . As such, the current study uses the same treatment conditions to build on previous studies that focus on understanding the regulation of metabolic rate depression over the anoxia/reoxygenation cycle.…”

“…5 In addition, previous studies have investigated the effects of exposure to 24-hour anoxia and 4 hours of aerobic recovery on metabolic rate depression in different wood frog tissues. 1,5,13,[53][54][55] As such, the current study uses the same treatment conditions to build on previous studies that focus on understanding the regulation of metabolic rate depression over the anoxia/reoxygenation cycle. Both liver and skeletal muscle contain glycogen that can be used to generate glucose to sustain glycolysis under anaerobic conditions.…”