The zebrafish (Danio rerio) has become a prevailing vertebrate model for developmental biology studies due to its ease of care, rapid embryogenesis stages development and translucent embryos. In this studies, ATM Kinase and MRN complex role as DNA damage response proteins during embryogenesis was examined by using specific MRN complex (Mirin) and ATM Kinase inhibitors (Ku60019 and Ku55933). To create DNA lesions in zebrafish, embryos at mid-blastula transition (MBT) stage were exposed to inhibitors (Mirin, Ku60019 or Ku55933) and later exposed to UVC irradiation wavelength of between 100 to 280 nm. Hatching but with visible physical deformation was observed for embryos treated with Mirin, Ku60019 or Ku55933 and UVC exposure at concentration of 3μM, 1.5 nM and 3nM or lower, respectively up to 72 hours-post fertilisation (hpf). On the other hand, no deformities were observed for all control as well as mock treated embryos. This study confirmed that DNA damage response proteins are crucial during embryo development to prevent undesired abnormal biological development. Thus, it is proven that protein inhibitors are a cheaper alternative in valuating specific protein roles during embryogenesis compared to both genomic and transcription modification tools.
A previous study has shown that synthetic antimicrobial peptides (AMPs) derived from Anabas testudineus (ATMP1) could in-vitro inhibit the progression of breast cancer cell lines. In this study, we are interested in studying altered versions of previous synthetic AMPs to gain some insight into the peptides functions. The AMPs were altered and subjected to bioinformatics prediction using four databases (ADP3, CAMP-R3, AMPfun, and ANTICP) to select the highest anticancer activity. The bioinformatics in silico analysis led to the selection of two AMPs, which are ATMP5 (THPPTTTTTTTTTTTYTAAPATTT) and ATMP6 (THPPTTTTTTTTTTTTTAAPARTT). The in silico analysis predicted that ATMP5 and ATMP6 have anticancer activity and lead to cell death. The ATMP5 and ATMP6 were submitted to deep learning databases (ToxIBTL and ToxinPred2) to predict the toxicity of the peptides and to (AllerTOP & AllergenFP) check the allergenicity. The results of databases indicated that AMPs are non-toxic to normal human cells and allergic to human immunoglobulin. The bioinformatics findings led to select the highest active peptide ATMP5, which was synthesised and applied for in-vitro experiments using cytotoxicity assay MTT Assay, apoptosis detection using the Annexin V FTIC-A assay, and gene expression using Apoptosis PCR Array to evaluate the AMP's anticancer activity. The antimicrobial activity is approved by the disc diffusion method. The in-vitro experiments analysis showed that ATMP5 had the activity to inhibit the growth of the breast cancer cell line (MDA-MB-231) after 48 h and managed to arrest the cell cycle of the MDA-MB-231, apoptosis induction, and overexpression of the p53 by interaction with the related apoptotic genes. This research opened up new opportunities for developing potential and selective anticancer agents relying on antimicrobial peptide properties.