Abstract

Background:
Pseudorabies virus (PRV), a zoonotic swine alphaherpesvirus, poses a severe threat to global pig production and public health.

Aim:
This study aimed to investigate the regulatory role of the RASSF6 gene in PRV-infected porcine alveolar macrophages (3D4/21).

Methods:
We established stable cellular models of RASSF6 overexpression (verified by qRT-PCR and immunofluorescence using an empty vector control) and siRNA-mediated RASSF6 silencing (verified by qRT-PCR) to investigate the role of RASSF6. Using these models, we assessed viral replication (quantified by qRT-PCR/TCID50) and apoptosis (analyzed by flow cytometry with Annexin V/PI staining and ELISA).

Results:
RASSF6 expression was significantly upregulated after PRV infection. RASSF6 overexpression inhibited 3D4/21 cell proliferation and PRV replication and promoted apoptosis through Bax and Caspase-3/9 activation. RASSF6 silencing promoted cell proliferation and viral replication and inhibited apoptosis.

Conclusion:
RASSF6 is a critical host defense factor against PRV infection in porcine alveolar macrophages. RASSF6 upregulation restricts viral replication by inducing apoptosis via Bax/caspase-3/9 activation and simultaneously suppressing cell proliferation. Conversely, RASSF6 silencing creates a permissive environment for PRV propagation by inhibiting apoptosis and enhancing cell survival. These findings reveal RASSF6-mediated apoptosis as a novel antiviral mechanism and highlight its potential as a therapeutic target for controlling PRV pathogenesis.

Key words: Apoptosis; Porcine alveolar macrophages; Pseudorabies virus; RASSF6.