Intracellular distribution of two rna binding proteins irp1 and irp2

Iron Regulatory Proteins fIRPs) are family RNA binding proteins and involved in regulation of intracellular iron homeostasis. Two of IRPs have been identified in human, rodent and other species. IRPl and IRP2 are known to be expressed in all cells but IRPl is predominant in most cells. High expression of IRP2 was obsprvfd in brain and muscle tissues. Both proteins recognize and bind to a specific RNA structure termed Iron Regulatory Elements (IREs). A single IRE is found in 5ö DTR of ferritin and mitochondria aconitase mRNAs while five IRKs are in 3' UTR of transferrin receptor mRNA. Both IRPs bind equally to those IRE's. In this study, we focus on the intracellular distribution of IRPl and IRP2 in human astrocytoma cells (SW1088) and mouse fibroblast cells (NIH 3T3). Immunohistochemistry study with polyclonal antibodies to IRPl or IRP2 shows that there is a difference in intracellular distribution. IRPl is found in cytoplasm and predominantly in peririuclei while IRP2 is found in both nuclei and cytoplasm. The intracellular distribution of c-myc tagged IRPs is similar to that seen in histochemistry study. To establish a system for studying intracellular distribution and translocation of IRPs in live cells, four IRP-GFP (Green Fluenrence Protein) constructs have been transfected into SW1088 and NTH 3T3 cells. The transient expression of IRP GFP fusion proteins have been examined in either live or fixed cells. In addition, the binding activity of IRPs and its target ferritin mRNA have been determined in each fraction collected on sucrose gradient. The data reported here will help us to further understand the functional difference between IRPl and IRP2.