title: Pepsin is a positive regulator of Ac-cathB-2 involved in the rat gut penetration of Angiostrongylus cantonensis.
abstract: Angiostrongyliasis caused by the rat lungworm, Angiostrongylus cantonensis (A. cantonensis), has globally spread from the traditional epidemic areas. The small intestine is the site where the third-stage larvae (L3) of this worm entered the host body, and parasite proteases are involved in this process. Ac-cathB-2, a cathepsin B-like cysteine of A. cantonensis, was formerly isolated from the insoluble part of fragmentised Escherichia coli without activity. The unplanned low activity of prokaryotic expression proteins and difficulties in genetic modification hindered understanding the function of this protein.
  
  
  
  The recombinant Ac-cathB-2 was expressed and harvested from 293 T cells and the enzymatic property and the effects of processing on the activity of the recombinant protease were investigated in vitro. The expression of Ac-cathB-2 in response to external stimulation was assessed, and the function of this protease during host gut penetration was observed by using antiserum for inhibition.
  
  
  
  Of the life-cycle stages studied, L3 expressed the highest level of Ac-cathB-2 gene and released the corresponding gene product from the body. The expression of this gene was rapidly upregulated after incubating L3 in small intestine homogenate of rat. Recombinant Ac-cathB-2 was harvested from 293 T cell culture medium. This protease was activated by pepsin-HCl and the enabled Ac-cathB-2 could subsequently digest laminin and fibronectin readily. Moreover, the small intestine isolated from rat was disrupted after incubating with the activated Ac-cathB-2, resulting in the detachment of epithelial cells. Antiserum treatment inhibited the hydrolytic ability of recombinant Ac-cathB-2 by 82.7 %, and also reduced the tissue penetration of activated L3 by 41.2 %. Additionally, pre-incubation of L3 with artificial gastric acid increased the number of penetrating larvae by 53.2 %, and this alteration could be partly blocked by antiserum treatment.
  
  
  
  We believe that Ac-cathB-2 from A. cantonensis might help the worm to penetrate the rat gut, because the protease was able to degrade the tissue components of host. Nevertheless, our results further indicated that host pepsin played a beneficial role in this process by cleaving Ac-cathB-2 for activation. Thus, Ac-cathB-2 may probably represent an important target for the control of A. cantonensis infection.
authors.affiliation: School of Life Sciences, Xiamen University, Xiamen, Fujian, 361102, China.|Translational Medicine Center, Hunan Cancer Hospital, Hunan, 410006, China.|State Key Laboratory of Cellular Stress Biology, Xiamen University, Xiamen, Fujian, 361102, China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Long
authors.affiliation: School of Life Sciences, Xiamen University, Xiamen, Fujian, 361102, China.|State Key Laboratory of Cellular Stress Biology, Xiamen University, Xiamen, Fujian, 361102, China.
authors.forename: Binbin
authors.initials: B
authors.lastname: Cao
authors.affiliation: Medical College, Xiamen University, Xiamen, Fujian, 361102, China.
authors.forename: Yinan
authors.initials: Y
authors.lastname: Wang
authors.affiliation: School of Life Sciences, Xiamen University, Xiamen, Fujian, 361102, China. dmluo@xmu.edu.cn.|State Key Laboratory of Cellular Stress Biology, Xiamen University, Xiamen, Fujian, 361102, China. dmluo@xmu.edu.cn.
authors.forename: Damin
authors.initials: D
authors.lastname: Luo
mesh_terms: D017185:Angiostrongylus cantonensis / Q000201:enzymology / Q000254:growth & development / Q000502:physiology*; D000818:Animals; D002401:Cathepsin B / Q000378:metabolism*; D005260:Female; D015801:Helminth Proteins / Q000378:metabolism; D007421:Intestine, Small / Q000469:parasitology*; D007814:Larva; D008018:Life Cycle Stages; D008297:Male; D010434:Pepsin A / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins; D017206:Strongylida Infections / Q000469:parasitology*
chemical_list: D015801:Helminth Proteins; D011994:Recombinant Proteins; D002401:Cathepsin B; D010434:Pepsin A
keywords: Ac-cathB-2; Angiostrongylus cantonensis; Intestinal penetration; Pepsin; Protease