Host blood proteins, represented mainly by hemoglobin and serum albumin, serve as the ultimate source of amino acids needed for de novo protein synthesis during tick development and reproduction. While uptake and processing of hemoglobin by tick gut cells have been studied in detail, molecular mechanisms of host serum albumin degradation remain unknown. In this work, we have used artificial membrane feeding of Ixodes ricinus females on a hemoglobin-free diet in order to characterize the proteolytic machinery involved in albuminolysis. Morphological comparisons of ticks fed on whole blood (BF) and serum (SF) at microscopic and ultrastructural levels showed that albumin and hemoglobin have different trafficking routes in tick gut cells. Analysis in vitro with selective inhibitors demonstrated that albumin is degraded at an acidic pH by a network of cysteine and aspartic peptidases with predominant involvement of cysteine cathepsins having endo- and exopeptidase activities. The cleavage map of albumin and the roles of individual peptidases in albumin degradation were determined. These results indicate that the albuminolytic pathway is controlled by the same proteolytic system that is responsible for hemoglobinolysis. This was further supported by the overall similarity of gut peptidase profiles in SF and BF ticks at the transcriptional and enzymatic activity levels. In conclusion, our work provides evidence that although hemoglobin and albumin are transported differentially during heterophagy they are digested by a common multienzyme proteolytic network. This central digestive system, critical for successful blood feeding in tick females, thus represents a valuable target for novel anti-tick interventions.

Institute of Chemistry and Biochemistry Czech Academy of Sciences Flemingovo nam 2 166 10 Prague Czech Republic

Institute of Parasitology Biology Centre of the Czech Academy of Sciences Branisovska 31 373 11 Ceske Budejovice Czech Republic

Institute of Parasitology Biology Centre of the Czech Academy of Sciences Branisovska 31 373 11 Ceske Budejovice Czech Republic; Faculty of Science University of South Bohemia Branisovska 31 373 11 Ceske Budejovice Czech Republic

References provided by Crossref.org

Insight Into the Dynamics of the Ixodes ricinus Nymphal Midgut Proteome

Haem-responsive gene transporter enables mobilization of host haem in ticks

Mialostatin, a Novel Midgut Cystatin from Ixodes ricinus Ticks: Crystal Structure and Regulation of Host Blood Digestion

Comparison of the hemolysis machinery in two evolutionarily distant blood-feeding arthropod vectors of human diseases

Insights into the Role of Tick Salivary Protease Inhibitors during Ectoparasite-Host Crosstalk

RNA-seq analyses of the midgut from blood- and serum-fed Ixodes ricinus ticks