Heme-induced Trypanosoma cruzi proliferation is mediated by CaM kinase II.

Trypanosoma cruzi, the etiologic agent of Chagas disease, is transmitted through triatomine vectors during their blood-meal on vertebrate hosts. These hematophagous insects usually ingest approximately 10mM of heme bound to hemoglobin in a single meal. Blood forms of the parasite are transformed into epimastigotes in the crop which initiates a few hours after parasite ingestion. In a previous work, we investigated the role of heme in parasite cell proliferation and showed that the addition of heme significantly increased parasite proliferation in a dose-dependent manner [1]. To investigate whether the heme effect is mediated by protein kinase signalling pathways, parasite proliferation was evaluated in the presence of several protein kinase (PK) inhibitors. We found that only KN-93, a classical inhibitor of calcium-calmodulin-dependent kinases (CaMKs), blocked heme-induced cell proliferation. KN-92, an inactive analogue of KN-93, was not able to block this effect. A T. cruzi CaMKII homologue is most likely the main enzyme involved in this process since parasite proliferation was also blocked when Myr-AIP, an inhibitory peptide for mammalian CaMKII, was included in the cell proliferation assay. Moreover, CaMK activity increased in parasite cells with the addition of heme as shown by immunological and biochemical assays. In conclusion, the present results are the first strong indications that CaMKII is involved in the heme-induced cell signalling pathway that mediates parasite proliferation.

Induction of acid phosphatase activity during germination of maize (Zea mays) seeds.

Acid phosphatase activity (orthophosphoric-monoester phosphohydrolase, EC 3.1.3.2) increased during the first 24 h of maize (Zea mays) seed germination. The enzyme displayed a pH optimum of 4.5-5.5. Catalytic activity in vitro displayed a linear time course (60 min) and reached its half maximum value at 0.47 mM p-nitrophenyl phosphate (pNPP). Phosphatase activity towards phosphoamino acids was greatest for phosphotyrosine. The phosphatase activity was strongly inhibited by ammonium molybdate, vanadate and NaF and did not require divalent cations for the catalysis. The temperature optimum for pNPP hydrolysis was 37 degrees C. Under the same conditions, no enzyme activity was detected with phytic acid as substrate. Western blotting of total homogenates during seed germination revealed proteins/polypeptides that were phosphorylated on tyrosine residues; a protein of approximately 14 kDa is potentially a major biological substrate for the phosphatase activity. The results presented in this study suggest that the acid phosphatase characterized under the tested conditions is a member of the phosphotyrosine phosphatase family.

Exploring the sialome of the blood-sucking bug Rhodnius prolixus.

Rhodnius prolixus is a Hemiptera that feeds exclusively on vertebrate blood in all life stages. Its salivary glands produce potent pharmacological substances that counteract host hemostasis, including anti-clotting, anti-platelet, and vasodilatory substances. To obtain a further insight into the salivary biochemical and pharmacological complexity of this insect, a cDNA library was randomly sequenced, and salivary gland homogenates were fractionated by HPLC to obtain aminoterminal sequences of abundantly expressed proteins. Results indicate a remarkable expansion of the lipocalin family in Rhodnius salivary glands, among other protein sequences described. A summary of 31 new full length proteins deducted from their mRNA sequence is described, including several new members of the nitrophorin, triabin, and pallidipin families. The electronic version of the complete tables is available at http://www.ncbi.nlm.nih.gov/projects/vectors/rhodnius_prolixus.