مقالات پذیرفته شده در هفتمین کنگره بین المللی زیست پزشکی
Expression change of Multidrug Resistance Protein (MRP1) modulates chronic salinity tolerances of sea urchin Echinometra mathaei
Expression change of Multidrug Resistance Protein (MRP1) modulates chronic salinity tolerances of sea urchin Echinometra mathaei
Rezvan Mousavi-Nadushan,1,*Mojgan Chitsaz,2
1. Department of Marine Science, Faculty of Natural Resources and Environment, Tehran North Branch, Islamic Azad University, Tehran, Iran. 2. Department of Marine Science, Faculty of Natural Resources and Environment, Science and Research Branch, Islamic Azad University Tehran, Iran.
Introduction: The attainment of any organism relies on environmental plasticity and also on its aptitude to survive to ecological variation (for instance preserving homeostasis in stress conditions). The multidrug resistance protein (MRP) family encrypts a diverse repertoire of ATP-binding cassette (ABC) transporters with multiple roles in development, disease, and homeostasis. Understanding MRP evolution is significant to separating their roles in their miscellaneous actions. Sea urchins occupy an essential phylogenetic point for considering the evolution of vertebrate proteins and have been an imperative invertebrate model organism for study of ABC transporters. However, under non pathological conditions in mammals/sea urchins, MRP1 has been confirmed to perform several roles that comprise control of inflammatory response of mast cells. These tasks are apparently associated with the capacity of this protein to carry endogenous biomolecules that perform straightly or secondarily as signaling molecules.
In general, sea urchins are considered a worthy taxa for biomonitoring researches because of its benthonic adult long-lasting life, hence straight exposed to anthropogenic pollutants. The coelomic fluid of the sea urchin comprises free immune effector cells, among them, red sphere cells, showed an excessive upsurge in quantity in individuals collected from contaminated environments. These cells activate their response strategy against diverse types of physical and chemical disturbances/pollutants/stressors. All these actions yield an increase in the content of the classical stress protein, such as heat shock protein 70 and transporters. Furthermore, their upregulation throughout cellular stress supports organism resistance during stressful situations and make them main effector of both defense and immune systems.
Methods: Individuals of E. mathaei were collected from the intertidal zone during the low tide from desalination plant discharge outlets, then coelomic fluid of each specimen (up to 2 mL) was withdrawn. For protein isolation, the coelomic fluid was centrifuged at 12,000g, for 20 min at 4 °C, and 15 µg of the supernatant from control and salt stressed sea urchins was separated in a 12% SDS-PAGE gel. The 45 kDa band were selected for MALDI-TOF analyses. After tryptic digestion, the resultant peptide fragments were injected to a MALDI TOF/TOF Mass Spectrometer and the role of them was evaluated using homology analyses in reference to annotation of their homologs.
Results: In this study, SDS gel electrophoresis and matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS) technology was used to examine differentially expressed proteins in coelomic fluid from control and salt stressed individuals of E. mathaei. SDS-PAGE proteins profile of CF extracts showed intense bands at 45 kDa. After the BLASTp searches against the NR database, the transcript sequences were further clustered according to the top hits found (E-value, 8E-15 to 1E 10). Consequently isoforms gave an identical primary structure in the protein matching and were classified as ATP-binding cassette transporters. Finally, BLASTp comparisons revealed the closest homologous sequence as multidrug resistance protein (MRP) with 87 % Query cover (EV, 1e-14).
Conclusion: The multidrug resistance protein (MRP) are a group of extremely conserved proteins, deciphered in vertebrates and invertebrates while they are exposed to cellular stress. The transporters are vital osmoregulation elements, responsible for the uptake and efflux of main materials such as mineral ions, sugars and amino acids. These proteins also play an important role in detoxification and chemo protection by transporting a wide range of compounds, especially conjugates of lipophilic substances with glutathione, glucuronate and sulfate. This protein was shown to have a molecular weight of 190 kDa, but he sea urchin MRP appears to have additional protein components in the molecular weight range of 14-55 kDa that might correspond to a cargo. Moreover, researchers have proved that sea star P-gp-like transporter are different from vertebrate P-gp transporters. They demonstrated that reactive MRP migrated at 45 kDa rather than in the 150–200 kDa range of vertebrate P-gp transporters in SDSPAGE.
This study has shown that multidrug resistance protein expression level was higher in salt stressed sea urchins E. mathaei, and this indicating MRP transporters perform a pivotal role in responding to stress. In the white leg shrimp Litopenaeus vannamei, ABC transporters showed an essential task in the physiological fluctuations associated with uptake, tolerance, and cell detoxification when they were exposed to stress.