Behind the Curtain: In silico and in vitro Experiments Brought to Light New Insights Into the Anticryptococcal Action of Synthetic Peptides

Abstract

Cryptococcus neoformans threaten the health, causing cryptococcal meningitis and pneumonia, especially in immunosuppressed patients, which can be fatal. Recently, our research group evaluated and studied the mechanisms of action of four synthetic peptides (SP) against C. neoformans. Here, in silico and in vitro analyses help deepen understanding of peptides' mechanisms of action. The interaction of the peptides with a membrane receptor was analyzed by docking analysis, in addition to ROS overproduction and the modulation of redox metabolism, inhibition of ergosterol biosynthesis, and release of cytochrome c. Out of four, three peptides interacted with membrane receptor PHO36 altering its structure and function and leading to a higher accumulation of O₂- and H2O2. C. neoformans cells treated with SP presented a reduction in the activity of antioxidant enzymes, corroborating ROS accumulation. However, in the presence of the antioxidant ascorbic acid, some peptides could not induce this oxidative stress and have the activity against C. neoformans affected. Curiously, two of these SPs still maintained the activity against C. neoformans and even induced the membrane pore formation as revealed by propidium iodide uptake assay, revealing their mechanism of action is ROS-independent. Additionally, SPs inhibited the biosynthesis of ergosterol, which corroborates the pore formation on the membrane of C. neoformans cells, inhibited the lactate dehydrogenase activity affecting the cell metabolism, and induced the release of Cyt c from the mitochondria inducing death by apoptosis in the cryptococcal cells. Our findings strongly suggest that SPs act by multiple mechanisms, making it difficult for C. neoformans to acquire resistance highlighting the potential of SPs as alternative molecules in treating infections caused by C. neoformans.

No Chance to Survive: Mo-CBP3-PepII Synthetic Peptide Acts on <em>Cryptococcus neoformans</em> by Multiple Mechanisms of Action

“…These data suggest the activity and mechanism of action of Mo-CBP3-PepII are dependent on ROS accumulation. This result is different from others presented by peptides RcAlb-PepIII and PepKAA [21], where the ability of pore formation is independent of ROS.…”

“…Recently, it was reported that two synthetic peptides, RcAlb-PepII and PepGAT, lost the ability to induce pore formation in the C. neoformans membrane in the presence of an antioxidant agent where no H2O2 accumulation. This result suggests both peptides induce pore formation by ROSdependent pathway [21]. Besides damaging the membrane, higher levels of H2O2 could also result in a higher level of oxidized protein and loss of activity.…”

“…Then The experiments to evaluate the anticryptococcal activity in the presence of the antioxidant agent ascorbic were done following the methodology described by Neto et al [27]. The experiments to perform the H2O2 overaccumulation and pore formation in ascorbic acid at 10 mM were performed as described by Aguiar et al [21].…”