This project investigates molecular mechanisms that allow fish to tolerate changes in water salinity. Salinity changes are pertinent in coastal fish habitats and desert lakes. Because water salinization represents a main environmental problem caused by climate change understanding how fish cope with salinity stress is critical. Some fish species, called euryhaline, such as tilapia have evolved extreme capacity for coping with salinity stress. This project utilizes molecular and biochemical approaches to investigate salinity stress response mechanisms in tilapia. It will reveal mechanisms underlying salinity tolerance and molecular cross-talk of stress and immune responses in fish. The focus is on gill tissue. Knowledge generated in this project allows targeting specific stress response mechanisms during fish management. A prerequisite for management of commercially important fish is knowledge of molecular pathways and physiological responses that should be targeted to alleviate environmental stress. Because many molecular responses to different types of stress are highly conserved the project has broad implications for understanding general stress response mechanisms in vertebrates (including humans). Moreover, the human kidney contains a region (the inner medulla) that experiences salinity fluctuations just like euryhaline fish. Understanding molecular coping mechanisms during salinity stress could benefit diagnosis and treatment of kidney diseases associated with failure of the renal urinary concentrating mechanism. Training and education in fish and general vertebrate stress biology and molecular and biochemical approaches is provided to graduate, undergraduate, and K-12 students. Outreach partnerships include the EnvironMentors and John Muir Institute of the Environment programs, aquaculture producers, state agencies, and the general public. Emphasis is placed on participation of underrepresented groups. The results of this project will be disseminated broadly via publications in peer-reviewed scientific journals, seminars, at scientific conferences and K-12 schools, web-based dissemination, posters, and during targeted hands-on outreach activities.
The key outcomes of this project include scientific and professional development achievements. The project has provided a wealth of resources and opportunities for professional training and development and this aspect has taken a significant amount of the overall time and resources spent for this proejct. Outreach via the Environmentors/ AggieMentors program and other venues to undergraduate and K12 students, aquaculture farmers, and state agencies has been provided. Graduate and undergraduate students enrolled at UC Dvais were trained and prepared for furthering their professional career goals. Such training was also extended to several international students and scholars that visited the PIs lab for periods ranging from 6 months to 1 year on fellowships awarded by their countries. Although these international researchers were not funded by this grant they profited greatly from the scientific resources established via this project in the PIs lab (see below). The professional development activities included training in state-of-the-art proteomics, bioinformatics, and other biochemical and physiological methods and approaches. They also included development of diagnostic and presentation skills as well as an awereness of the global and overarching nature of science for addressing global world-wide societal challenges and opportunities. Major scientific outcomes of this project included the generation of new basic knowledge that helps us better understand how animals cope with environmental change, Specifically, the project revealed novel underlying mechanisms that fish utilize for tolerating climate-induced salinity stress. For example the cytokine TNFalpha was cloned from tilapia, purified, recombinently expressed, and its function tested by exposing it to tilapia cell lines. The involvemnent of this protein in osmotic stress responses was shown to be relatively low. In contrast to TNFalpha, both enzymes of the myo-inositol biosynthesis pathway are extremely strongly regulated by salinity stress in adult tilapia, tilapia larvae, and tilapia cell lines at multiple levels of organization. Thus, the corresponding biochemical pathway is extremely important for for salinty stress responses of fish, in particular tilapia, which are a major aquaculture species, especially in developing countries that are impacted by climate change. The project also demonstrated that the myo-inositiol biosynthesis pathway represents a unique and extremely robust model system that is now available for functional mechanistic studies of osmotic stress sensing and signaling networks in fish. In addition, the project succeeded in the development, establishment, and validation of a novel workflow for quantitative molecular phenotyping based on two-tiered gel-free and label-free quantitative proteomics. This is a major accomplishment that required an extraordinary amount of intellectual resources and time commitment. Together with the generation and immortalization of tilapia cell lines accomplished during this project, the novel molceular phenotyping workflow breaks new ground for in-depth analysis of the biochemical mechanisms by which animals respond to environmental and climate stress. This project has already utilized these tools to identify many key proteins involved in the biochemical network that confers high salinity stress tolerance in fish. Knowledge of these proteins and their context-dependent functions provides deep mechanistic insight into the evolution and physiology of environmental stress responses of fish.
