Carvajal-Gamez Bertha Isabel, Quintas-Granados Laura Itzel, Arroyo Rossana, Vázquez-Carrillo Laura Isabel, Ramón-Luing Lucero De los Angeles, Carrillo-Tapia Eduardo, Alvarez-Sánchez María Elizbeth
Genomic Sciences Postgraduate, Autonomous University of Mexico City (UACM), Mexico City, Mexico.
Department of Infectomics and Molecular Pathogenesis, Center for Research and Advanced Studies, IPN, Mexico City, Mexico.
PLoS One. 2014 Sep 24;9(9):e107293. doi: 10.1371/journal.pone.0107293. eCollection 2014.
Polyamines are involved in the regulation of some Trichomonas vaginalis virulence factors such as the transcript, proteolytic activity, and cytotoxicity of TvCP65, a cysteine proteinase (CP) involved in the trichomonal cytotoxicity. In this work, we reported the putrescine effect on TvCP39, other CP that also participate in the trichomonal cytotoxicity. Parasites treated with 1,4-diamino-2-butanone (DAB) (an inhibitor of putrescine biosynthesis), diminished the amount and proteolytic activity of TvCP39 as compared with untreated parasites. Inhibition of putrescine biosynthesis also reduced ∼ 80% the tvcp39 mRNA levels according to RT-PCR and qRT-PCR assays. Additionally, actinomycin D-treatment showed that the tvcp39 mRNA half-life decreased in the absence of putrescine. However, this reduction was restored by exogenous putrescine addition, suggesting that putrescine is necessary for tvcp39 mRNA stability. TvCP39 was localized in the cytoplasm but, in DAB treated parasites transferred into exogenous putrescine culture media, TvCP39 was re-localized to the nucleus and nuclear periphery of trichomonads. Interestingly, the amount and proteolytic activity of TvCP39 was recovered as well as the tvcp39 mRNA levels were restored when putrescine exogenous was added to the DAB-treated parasites. In conclusion, our data show that putrescine regulate the TvCP39 expression, protein amount, proteolytic activity, and cellular localization.
多胺参与某些阴道毛滴虫毒力因子的调控，如TvCP65（一种参与滴虫细胞毒性的半胱氨酸蛋白酶）的转录、蛋白水解活性和细胞毒性。在本研究中，我们报道了腐胺对TvCP39的影响，TvCP39是另一种也参与滴虫细胞毒性的半胱氨酸蛋白酶。用1,4-二氨基-2-丁酮（DAB，腐胺生物合成抑制剂）处理的寄生虫，与未处理的寄生虫相比，TvCP39的量和蛋白水解活性降低。根据逆转录-聚合酶链反应（RT-PCR）和定量逆转录-聚合酶链反应（qRT-PCR）分析，腐胺生物合成的抑制也使tvcp39 mRNA水平降低了约80%。此外，放线菌素D处理表明，在没有腐胺的情况下，tvcp39 mRNA的半衰期缩短。然而，通过添加外源性腐胺可恢复这种降低，这表明腐胺对于tvcp39 mRNA的稳定性是必需的。TvCP39定位于细胞质中，但在转移到外源性腐胺培养基中的经DAB处理的寄生虫中，TvCP39重新定位于滴虫的细胞核和核周边。有趣的是，当将外源性腐胺添加到经DAB处理的寄生虫中时，TvCP39的量和蛋白水解活性得以恢复，tvcp39 mRNA水平也得以恢复。总之，我们的数据表明腐胺调节TvCP39的表达、蛋白量、蛋白水解活性和细胞定位。
