Biophysical Characterization of Zebrafish Transient Receptor Potential Cation Channel, Subfamily M, Member 7.

Abstract

TRPM7 is a non-selective divalent ion channel that belongs to the transient receptor potential superfamily; it is permeable to divalent ions such as calcium and magnesium as well as trace elements such as manganese and zinc. Although TRPM7 is permeable to many divalent ions, it is regulated by intracellular magnesium and magnesium adenosine triphosphate. Moreover, TRPM7 is thought to be involved in magnesium homeostasis of the cell. Magnesium is important for many processes within an organism. When serum levels of magnesium drop, a state of hypomagnesemia is reached. Hypomagnesemia is associated with many symptoms such as tremors, convulsions and even coma. TRPM7 has been implicated in hypomagnesemia, as well as breast cancer metastasis and delayed neuronal death post brain ischemia. Human TRPM7 has been characterized and studied, but limitations do exist within animal models to study TRPM7. Global knockout of TRPM7 in mice leads to death at embryonic day 7.5. This fact limits the developmental studies that can be performed on a mouse. A nonmammalian model, zebrafish, may be able to bridge the gap in studies of TRPM7. There exists zebrafish with a loss-of-function mutation in TRPM7 that survive; these mutants show skeletogenesis defects and kidney stone formation. Zebrafish TRPM7 has yet to be biophysically characterized. Here, it is investigated whether zebrafish TRPM7 is biophysically similar to human TRPM7. Zebrafish TRPM7 and three truncated mutants of zebrafish TRPM7 were overexpressed in human embryonic kidney cells. The truncated mutants of TRPM7 were utilized as a means to narrow down the magnesium and magnesium adenosine triphosphate binding sites within zebrafish TRPM7. This study shows that: 1- Zebrafish TRPM7 is biophysically similar to human TRPM7 in regards to regulation, divalent ion permeability and pH sensitivity, 2- Zebrafish TRPM7 is less sensitive to osmolarity than human TRPM7, but remains sensitive. 3- Common human TRPM7 inhibitors do not inhibit Zebrafish TRPM7. Additionally, investigation of the three truncated zebrafish TRPM7 mutants suggests that there is a third magnesium-binding site. Experimental designs with zebrafish as an animal model for TRPM7 pathologies and diseases should take into account the differences in zebrafish TRPM7 and human TRPM7.