Proteomic profiling identifies nucleolin as a U3-interacting protein that inhibits avian leukosis virus replication.

The long terminal repeat (LTR) of the avian retroviruses has been extensively characterized as a model of a strong transcription regulatory unit. U3 gene of LTR contains many cis-acting elements and trans-acting factors that bind to a variety of host transcription factors to form a complex network regulating high levels of transcription of viral and host genes. However, the host proteins that bind to the U3 gene and their complex mechanisms are largely unknown. In the present study, coupling surface-plasmon resonance and mass spectrometry were performed to identify proteins binding to the U3 gene of avian leukosis virus (ALV). A total of 181 U3-binding proteins with score > 20 were identified. Six of the proteins with score > 120 were successfully knockdown using siRNA transfection and the transcription of ALV gp85 mRNA was significantly increased when the nucleolin (NCL) was down regulated. The effect of NCL interference was optimized by combining siRNAs and we found that down regulation of NCL significantly increased the transcription activity of LTR, the enhancer ability of U3, and furthermore enhanced the expression of viral protein. On the contrary, over expression of NCL significantly decreased the transcription activity of LTR, the enhancer ability of U3, and the expression level of viral protein. These data demonstrate that NCL is critical for inhibition ALV replication and laid a molecular basis for development of efficient strategies against retroviruses from a new perspective.