Characterization of a recombinant Cathepsin B-like cysteine peptidase from Diaphorina citri Kuwayama (Hemiptera: Liviidae): A putative target for control of citrus huanglongbing

Authors: FERRARA, TAISE - Universidade Federal De Sao Carlos; SCHNEIDER, VANESSA - Universidade Federal De Sao Carlos; KISHI, LUCIANO - Universidade Federal De Sao Carlos; CARMONA, ADRIANA - Universidade Nacional De Sao Paulo; ALVES, MARCIO - Universidade Nacional De Sao Paulo; BELASQUE-JUNIOR, JOSE - Universidade Nacional De Sao Paulo; ROSA, JOSE - Universidad De Sao Paulo; Hunter, Wayne; HENRIQUE-SILVA, FLAVIO - Universidade Federal De Sao Carlos; SOARES-COSTA, ANDREA - Universidade Federal De Sao Carlos

Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/6/2015
Publication Date: 12/30/2015
Citation: Ferrara, T., Schneider, V.K., Kishi, L.T., Carmona, A.K., Alves, M., Belasque-Junior, J., Rosa, J.C., Hunter, W.B., Henrique-Silva, F., Soares-Costa, A. 2015. Characterization of a recombinant Cathepsin B-Like cysteine peptidase from Diaphorina citri Kuwayama (Hemiptera: Liviidae): A putative target onctrol of citrus huanglongbing. PLoS One.

Interpretive Summary: Huanglongbing (HLB) is one of the most destructive disease affecting citrus. The causal agent is a phloem-limited bacterium, Liberibacter asiaticus spread by the Asian citrus psyllid (ACP). HLB control strategies include methods of disrupting the biology of Asian citrus psyllid (ACP) including methods of reducing ACP enzyme activity. We investigated cysteine peptidases as a target for ACP control. We report on the identification and characterization of a cathepsin B-like cysteine peptidase (DCcathB). Analyses revealed that the expression of the DCcathB in nymph and adult psyllids was approximately 9-fold greater than in eggs. Moreover, the expression of this enzyme was thousands of times greater than in the other body tissues, suggesting that DCcathB is important to feeding and may be a target for managing ACP and thus HLB.

Technical Abstract: Huanglongbing (HLB) is one of the most destructive disease affecting citrus plants. The causal agent is associated with the phloem-limited bacterium Candidatus Liberibacter asiaticus (CLas) spread by the Asian citrus psyllid, Diaphorina citri (Hemiptera: Liviidae). Among the control strategies for HLB disease, is reducing the enzymes of Diaphorina citri (D. citri). Hemipterans, have cysteine peptidases in the gut. The importance of this class of enzymes led us to search for enzymes in the D. citri transcriptome for the establishment of alternative strategies for HLB control. In this study, identification and characterization of a cathepsin B-like cysteine peptidase from D. citri (DCcathB) is provided. DCcathB was recombinantly expressed in Pichia pastoris, presenting a molecular mass of approximately 50 kilo dalton. The recombinant enzyme was inhibited by the cysteine protease inhibitors E64 and by the selective cathepsin B inhibitor CA-074. RT-quantitative polymerase chain reaction (qPCR) analysis revealed that the expression of the DCcathB in nymph and adult was approximately 9-fold greater than in egg. Moreover, the expression of this enzyme in the gut was 175-fold and 3333-fold higher than in the remaining tissues and in the head, respectively, suggesting that DCcathB may be a suitable target to reduce D. citri and stop the spread of HLB.