Elsevier

Gene

Volume 676, 15 November 2018, Pages 164-170
Gene

Short communication
The novel cathelicidin of naked mole rats, Hg-CATH, showed potent antimicrobial activity and low cytotoxicity

https://doi.org/10.1016/j.gene.2018.07.005Get rights and content

Highlights

  • Hg-CATH was the single cathelicidin identified in the genome of naked mole rats.

  • ΔHg-CATH exhibited bactericidal activity against gram-negative bacteria.

  • ΔHg-CATH showed low toxicity to chicken erythrocyte and mammalian cells.

  • Peptides like Hg-CATH may protect against multi-drug resistance bacteria.

Abstract

We performed the in silico genome-wide identification of antimicrobial peptides against the available genome sequence of the naked mole rat Heterocephalus glaber (H. glaber). Our results showed the presence of Hg-CATH, the single cathelicidin containing the antimicrobial domain in H. glaber. We chemically synthesized a 25 amino-acid peptide (ΔHg-CATH) corresponding to the predicted antimicrobial-active core region of Hg-CATH, and evaluated its antibacterial activity against seven bacterial strains. The ΔHg-CATH peptide exhibited strong bactericidal activity against gram-negative bacteria, including a multi-drug resistant strain, while showing low toxicity towards mammalian cells, including erythrocytes. Scanning electron microscopy images of bacterial cells treated with ΔHg-CATH showed disruption of their membranes due to the formation of toroidal pores. Identifying novel antimicrobial peptides, such as Hg-CATH, may be important for identifying candidate peptides for the control of multi-drug resistant bacteria.

Introduction

The identification of endogenous antimicrobial peptides (AMPs) has been difficult, and thus limited to only a few species. However, newly emerging genome-sequences of diverse species has greatly expedited the discovery of diverse AMPs, and through reverse genetics demonstrate the importance of these molecules as a part of the innate immune system (Tessera et al., 2012; Liu et al., 2015; Kim et al., 2017).

Cathelicidins are a family of AMPs that has been identified in most vertebrate animals (Hancock and Chapple, 1999). Cathelicidins consist of a signal peptide in the N-terminal of the protein, a conserved cathelin-like domain (CLD) in the middle, and an antimicrobial domain of variable length and biological activity in the C-terminal region (Zanetti et al., 1995). Previous studies have shown that cathelicidins are activated by proteolytic cleavage by specific enzymes, which releases the C-terminal antimicrobial domain producing the mature peptides (Scocchi et al., 1992). Cathelicidins show antimicrobial activities against bacteria, fungi, and enveloped viruses, along with other biological activities including angiogenesis, and wound healing (Zanetti, 2005).

The naked mole rat (Heterocephalus glaber) is an animal with an unusual life span, significantly longer than those of other rodents and shows delayed aging, high fecundity until death, and resistance to both spontaneous and experimentally-induced cancers (Seluanov et al., 2009; Liang et al., 2010). Although the genome sequence of H. glaber indicates the presence of AMPs, the biochemical and functional characterizations of these AMPs have not been clearly addressed.

In the present study, we performed a genome-wide in silico analysis of naked mole rats, and determined the AMP repertoire. Subsequently, we investigated the functional characteristics of Hg-CATH, the only cathelicidin containing the antimicrobial domain in naked mole rats. Understanding the characteristics of novel AMPs, like Hg-CATH, would expand our knowledge of the functions of these molecules and contribute to the rational development of antibiotic alternatives and their biomedical application.

Section snippets

In silico identification of AMPs from the genome of Heterocephalus glaber

A search using the query, “antimicrobial peptide AND reviewed: yes” against UniProtKB/Swiss-Prot (http://www.uniprot.org/uniprot/), and manual examination resulted in 2477 matches (data not shown). After removing sequences from non-vertebrate origins, 1166 sequences consisting of 36 protein families with antimicrobial activity remained (Table S1), and were searched against the genome sequence (AHKG00000000) of H. glaber (taxid: 10181) using the National Center for Biotechnology Information

In silico identification of putative AMPs from analysis of the Heterocephalus glaber genome

BLAST analysis of 1166 vertebrate AMPs (Table S1) consisting of 36 gene families against the genome sequence of H. glaber resulted in the identification of 27 AMPs, including one alpha-defensin, 22 beta-defensins, two cathelicidins, one LEAP-2, and one hepcidin (e-value < 0.001; Table S2). The remaining hits were proteins bearing partial sequence homology to AMPs, or included antimicrobial activity as part of their functions, indicating that the number of AMPs in the genome of H. glaber was

Discussion

Heterocephalus glaber are underground dwelling mammals, and therefore may more frequently face microorganisms than above-land dwelling animals. This hypothesis leads to the expectation of the presence of an improved innate immune system including components such as AMPs that would be active against microorganisms. However, in contrast to our expectation, the AMP repertoire of the naked mole rat genome was smaller than that found in other mammals, including mice and human (Table S3, (Belov et

Acknowledgements

This paper was supported by Konkuk University in 2016.

Conflict of interest

The authors declare competing financial interests. The peptide sequences ΔHg-CATH are the subject of domestic and foreign patent applications by Konkuk University.

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