Hongwei Cao,Hao Deng,Yanjin Wang,Diqiu Liu,Lianfeng Li,Meilin Li,Dingkun Peng,Jingwen Dai,Jiaqi Li,Huaji Qiu,Su Li

Viruses.2024 Jun 30;16(7):1058.doi: 10.3390/v16071058.

Abstract

The B169L protein (pB169L) of African swine fever virus (ASFV) is a structural protein with an unidentified function during the virus replication. The sequences of the  B169L  gene and the downstream  B438L  gene are separated by short intergenic regions. However, the regulatory mode of the gene transcription remains unknown. Here, we identified two distinct promoter regions and two transcription start sites (TSSs) located upstream of the open reading frame (ORF) of  B438L . Using the promoter reporter system, we demonstrated that the cis activity of the ORF proximal promoter exhibited significantly higher levels compared with that of the distal promoter located in the  B169L  gene. Furthermore, transfection with the plasmids with two different promoters for  B438L  could initiate the transcription and expression of the  B438L  gene in HEK293T cells, and the cis activity of the ORF proximal promoter also displayed higher activities compared with the distal promoter. Interestingly, the  B438L  distal promoter also initiated the transcription of the alternatively spliced  B169L  mRNA ( B169L  mRNA2) encoding a truncated pB169L (tpB169L) (amino acids 92-169), and the gene transcription efficiency was increased upon mutation of the initiation codon located upstream of the alternatively spliced  B169L  gene. Taken together, we demonstrated that the distal promoter of  B438L  gene initiates the transcription of both the  B438L  mRNA and  B169L  mRNA2. Comprehensive analysis of the transcriptional regulatory mode of the  B438L  gene is beneficial for the understanding of the association of B438L protein and pB169L and the construction of the gene-deleted ASFV.

Keywords: African swine fever virus; B169L protein; B438L promoter; transcription initiation.