Molecular and structural characterization of novel cystatins from the taiga tick Ixodes persulcatus.
Cystatins are cysteine peptidase inhibitors that in ticks mediate processes such as blood feeding and digestion. The ixodid tick Ixodes persulcatus is endemic to the Eurasia, where it is the principal vector of Lyme borreliosis. To date, no I. persulcatus cystatin has been characterized. In the present work, we describe three novel cystatins from I. persulcatus, named JpIpcys2a, JpIpcys2b and JpIpcys2c. In addition, the potential of tick cystatins as cross-protective antigens was evaluated by vaccination of hamsters using BrBmcys2c, a cystatin from Rhipicephalus microplus, against I. persulcatus infestation. Sequence analysis showed that motifs that are characteristic of cystatins type 2 are fully conserved in JpIpcys2b, while mutations are present in both JpIpcys2a and JpIpcys2c. Protein-protein docking simulations further revealed that JpIpcys2a, JpIpcys2b and JpIpcys2c showed conserved binding sites to human cathepsins L, all of them covering the active site cleft. Cystatin transcripts were detected in different I. persulcatus tissues and instars, showing their ubiquitous expression during I. persulcatus development. Serological analysis showed that although hamsters immunized with BrBmcys2c developed a humoral immune response, this response was not adequate to protect against a heterologous challenge with I. persulcatus adult ticks. The lack of cross-protection provided by BrBmcys2c immunization is perhaps linked to the fact that cystatins cluster into multigene protein families that are expressed differentially and exhibit functional redundancy. How to target such small proteins that are secreted in low quantities remains a challenge in the development of suitable anti-tick vaccine antigens.
Centro de Biotecnologia, Universidade Federal do Rio Grande do Sul, Avenida Bento Gonçalves, 9500, Prédio 43421, Porto Alegre 91501-970, RS, Brazil.
Carolina K
CK
Rangel
Centro de Biotecnologia, Universidade Federal do Rio Grande do Sul, Avenida Bento Gonçalves, 9500, Prédio 43421, Porto Alegre 91501-970, RS, Brazil; Laboratory of Infectious Diseases, Department of Disease Control, Graduate School of Veterinary Medicine, Hokkaido University, 060-0818, Sapporo, Hokkaido, Japan.
Luís F
LF
Parizi
Centro de Biotecnologia, Universidade Federal do Rio Grande do Sul, Avenida Bento Gonçalves, 9500, Prédio 43421, Porto Alegre 91501-970, RS, Brazil.
Gabriela A
GA
Sabadin
Unidade de Experimentação Animal, Universidade Estadual do Norte Fluminense Darcy Ribeiro, Avenida Alberto Lamego, 2000, Campos dos Goytacases, 28035-200, RJ, Brazil.
Evenilton P
EP
Costa
LICC-Laboratório Integrado de Computação Científica-Universidade Federal do Rio de Janeiro-Campus Macaé, Macaé, 27901-000, RJ, Brazil.
Nelilma C
NC
Romeiro
Laboratory of Infectious Diseases, Department of Disease Control, Graduate School of Veterinary Medicine, Hokkaido University, 060-0818, Sapporo, Hokkaido, Japan.
Masayoshi
M
Isezaki
Laboratory of Infectious Diseases, Department of Disease Control, Graduate School of Veterinary Medicine, Hokkaido University, 060-0818, Sapporo, Hokkaido, Japan.
Naftaly W
NW
Githaka
Departamento de Farmacociências, Universidade Federal de Ciências da Saúde de Porto Alegre, Rua Sarmento Leite, 245, Porto Alegre 90050-170, RS, Brazil; Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular, Brazil.
Adriana
A
Seixas
Unidade de Experimentação Animal, Universidade Estadual do Norte Fluminense Darcy Ribeiro, Avenida Alberto Lamego, 2000, Campos dos Goytacases, 28035-200, RJ, Brazil; Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular, Brazil.
Carlos
C
Logullo
Laboratory of Infectious Diseases, Department of Disease Control, Graduate School of Veterinary Medicine, Hokkaido University, 060-0818, Sapporo, Hokkaido, Japan.
Satoru
S
Konnai
Laboratory of Infectious Diseases, Department of Disease Control, Graduate School of Veterinary Medicine, Hokkaido University, 060-0818, Sapporo, Hokkaido, Japan.
Kazuhiko
K
Ohashi
Centro de Biotecnologia, Universidade Federal do Rio Grande do Sul, Avenida Bento Gonçalves, 9500, Prédio 43421, Porto Alegre 91501-970, RS, Brazil; Faculdade de Veterinária, Universidade Federal do Rio Grande do Sul, Avenida Bento Gonçalves, 9090, Porto Alegre 91540-000, RS, Brazil; Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular, Brazil. Electronic address: itabajara.vaz@ufrgs.br.
Itabajara
I
da Silva Vaz
D000818:Animals; D000906:Antibodies / Q000097:blood / Q000276:immunology; D060829:Arthropod Proteins / Q000737:chemistry* / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D001665:Binding Sites; D056668:Cathepsin L / Q000737:chemistry; D006224:Cricetinae; D015891:Cystatins / Q000737:chemistry* / Q000235:genetics*; D006801:Humans; D056724:Immunity, Humoral; D018884:Ixodes / Q000276:immunology / Q000378:metabolism*; D008958:Models, Molecular; D062105:Molecular Docking Simulation; D005810:Multigene Family; D010802:Phylogeny; D060888:Real-Time Polymerase Chain Reaction; D048494:Rhipicephalus / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013984:Tick Infestations / Q000517:prevention & control*
D000906:Antibodies; D060829:Arthropod Proteins; D015891:Cystatins; D056668:Cathepsin L
Cystatin; Ixodes persulcatus; Rhipicephalus microplus; Tick; Vaccine