Regulation of the immediate-early genes of white spot syndrome virus by Litopenaeus vannamei kruppel-like factor (LvKLF)

• A kruppel-like factor was cloned from Litopenaeus vannamei and characterized.
• LvKLF upregulates transcription of key WSSV immediate-early genes IE1 and WSSV304.
• LvKLF binds to KLF response elements in the IE1 and WSSV304 promoters.
• Knockdown of LvKLF in WSSV-infected shrimp reduced IE1 and WSSV304 mRNA levels.
• LvKLF silencing was found to inhibit the viral replication of WSSV.

Kruppel-like factors (KLFs) belong to a subclass of Cys2/His2 zinc-finger DNA-binding proteins, and act as important regulators with diverse roles in cell growth, proliferation, differentiation, apoptosis and tumorigenesis. Our previous research showed that PmKLF from Penaeus monodon is crucial for white spot syndrome virus (WSSV) infection, yet the mechanisms by which PmKLF influences WSSV infection remain unclear. This study cloned KLF from Litopenaeus vannamei (LvKLF), which had 93% similarity with PmKLF. LvKLF formed a dimer via the C-terminal zinc-finger motif. Knockdown of LvKLF expression by dsRNA injection in WSSV-challenged shrimps was found to significantly inhibit the transcription of two important immediate-early (IE) genes, IE1 and WSSV304, and also reduced WSSV copy numbers. Moreover, reporter assays revealed that the promoter activities of these two WSSV IE genes were substantially enhanced by LvKLF. Mutations introduced in the promoter sequences of IE1 and WSSV304 were shown to abolish LvKLF activation of promoter activities; and an electrophoretic mobility shift assay demonstrated that LvKLF binds to putative KLF-response elements (KRE) in the promoters. Taken together, these results indicate that LvKLF transcriptional regulation of key IE genes is critical to WSSV replication.