THE PUTATIVE PHYSIOLOGICAL ROLES OF TWO RHIPICEPHALUS MICROPLUS CYSTATINS
Autor(es): Luís Fernando Parizi, Gabriela Alves Sabadin, María Fernanda Alzugaray, Adriana Seixas, Carlos Logullo, Satoru Konnai, Kazuhiko Ohashi, Aoi Masuda, Itabajara da Silva Vaz Jr
THE PUTATIVE PHYSIOLOGICAL ROLES OF TWO RHIPICEPHALUS MICROPLUS CYSTATINS
» Área de pesquisa: ACAROLOGIA
» Instituição: UFRGS
» Agência de fomento e patrocinadores: CNPq, CAPES, FAPERGS, FAPERJ and INCT-EM (Brazil); MEXT (Japan).
The cystatin family is composed by cysteine peptidase inhibitors. Regulation of tick endogenous metabolism as well as host immune system by cystatins is related during tick blood-feeding, digestion and development. However, cystatin physiological roles remain uncharacterized in Rhipicephalus microplus, a tick responsible for significant economic losses in livestock. Here we report the cysteine peptidase inhibition profiles and tissue localization of two R. microplus cystatins. Recombinant cystatins named rBrBmcys2b and rBrBmcys2c were expressed in Escherichia coli and purified by affinity chromatography. Apparent inhibition constants were measured to estimate the rBrBmcys2b and rBrBmcys2c inhibitory capacity to cathepsin B, C, and L. The presence of native cystatins in salivary glands, ovary, gut and fat body were analyzed by western blot. Cystatins showed distinct affinities for cathepsins: rBrBmcys2b inhibited all cathepsins tested, showing higher affinity for cathepsin B (Ki 0.6 nM), while rBrBmcys2c does not inhibited cathepsin B, showing higher affinity for cathepsin C (Ki 0.8 nM). By serological analysis, BrBmcys2b was detected in ovary, salivary glands and fat body of partial and fully engorged R. microplus. BrBmcys2c was detected in partially engorged R. microplus gut and in fully engorged R. microplus ovary, salivary glands and fat body. These results support the differential presence of BrBmcys2b and BrBmcys2c in R. microplus tick tissues, and demonstrate that cathepsins B, C and L are inhibited by these cystatins at different degrees. These results suggest distinct tick physiological roles of these inhibitors: BrBmcys2b could module enzymes involved in egg development or host immune system response, while BrBmcys2c could be involved in modulation of enzymes responsible for blood meal processing. Further interference RNA and immunomodulatory experiments can provide more details about the role of BrBmcys2b and BrBmcys2c in tick physiology.
