An Essential Role of c-Fos in Notch1-mediated Promotion of Proliferation of KSHV-Infected SH-SY5Y Cells

Huiling Xu; Jinghong Huang; Lixia Yao; Wenyi Gu; Aynisahan Ruzi; Yufei Ding; Ying Li; Weihua Liang; Jinfang Jiang; Zemin Pan; Dongdong Cao; Naiming Zhou; Dongmei Li; Jinli Zhang

doi:10.2174/0118761429264583231106104202

An Essential Role of c-Fos in Notch1-mediated Promotion of Proliferation of KSHV-Infected SH-SY5Y Cells

Curr Mol Pharmacol. 2024 Jan 11. doi: 10.2174/0118761429264583231106104202. Online ahead of print.

Authors

Huiling Xu^{1

2}, Jinghong Huang¹, Lixia Yao¹, Wenyi Gu³, Aynisahan Ruzi⁴, Yufei Ding⁵, Ying Li⁶, Weihua Liang¹, Jinfang Jiang¹, Zemin Pan¹, Dongdong Cao¹, Naiming Zhou^{6

7}, Dongmei Li¹, Jinli Zhang¹

Affiliations

¹ Key Laboratory of Xinjiang Endemic and Ethnic Diseases, NHC Key Laboratory of Prevention and Treatment of Central Asia High Incidence Diseases, School of Medicine, Shihezi University, Shihezi City, Xinjiang 832002, China.
² Department of Pathology, Central People's Hospital of Zhanjiang, Zhanjiang, Guangdong 52400, China.
³ Australian Institute for Bioengineering and Nanotechnology (AIBN), University of Queensland (UQ), Corner College and Cooper Roads (Building 75), St Lucia, Brisbane QLD 4072, Australia.
⁴ Xinjiang Bazhou People's Hospital, Korla, Xinjiang 841000, China.
⁵ Department of Pathology, the Fourth Division Hospital of Xinjiang Production and Construction Corps, Yining 835100, China.
⁶ School of Medicine, Tarim University, Alaer 843300, Xinjiang, China.
⁷ Institute of Biochemistry, College of Life Sciences, Zijingang Campus, Zhejiang University, Hangzhou, Zhejiang 310058, China.

PMID: 38258595
DOI: 10.2174/0118761429264583231106104202

Abstract

Background: This study aimed to investigate the influence of Notch1 on c-Fos and the effect of c-Fos on the proliferation of Kaposi's sarcoma-associated herpesvirus (KSHV)-infected neuronal cells.

Methods: Real-time PCR and western blotting were used to determine c-Fos expression levels in KSHV-infected (SK-RG) and uninfected SH-SY5Y cells. C-Fos levels were measured again in SK-RG cells with or without Notch1 knockdown. Next, we measured c-Fos and p-c-Fos concentrations after treatment with the Notch1 γ-secretase inhibitor LY-411575 and the Notch1 activator Jagged-1. MTT and Ki-67 staining were used to evaluate the proliferation ability of cells after c-Fos levels downregulation. CyclinD1, CDK6, and CDK4 expression levels and cell cycle were analyzed by western blotting and flow cytometry, respectively. After the c-Fos intervention, the KSHV copy number and gene expression of RTA, LANA and K8.1 were analyzed by real-time TaqMan PCR.

Results: C-Fos was up-regulated in KSHV-infected SK-RG cells. However, the siRNA-mediated knockdown of Notch1 resulted in a significant decrease in the levels of c-Fos and p-c-Fos (P <0.01, P <0.001). Additionally, a decrease in Cyclin D1, CDK6, and CDK4 was also detected. The Notch1 inhibitor LY-411575 showed the potential to down-regulate the levels of c-Fos and p-c-Fos, which was consistent with Notch1 knockdown group (P <0.01), whereas the expression and phosphorylation of c-Fos were remarkably up-regulated by treatment of Notch1 activator Jagged-1 (P <0.05). In addition, our data obtained by MTT and Ki-67 staining revealed that the c-Fos down-regulation led to a significant reduction in cell viability and proliferation of the SK-RG cells (P <0.001). Moreover, FACS analysis showed that the cell cycle was arrested in the G0/G1 stage, and the expressions of Cyclin D1, CDK6, and CDK4 were down-regulated in the c-Fos-knockdown SK-RG cells (P <0.05). Reduction in total KSHV copy number and expressions of viral genes (RTA, LANA and K8.1) were also detected in c-Fos down-regulated SK-RG cells (P <0.05).

Conclusion: Our findings strongly indicate that c-Fos plays a crucial role in the promotion of cell proliferation through Notch1 signaling in KSHV-infected cells. Furthermore, our results suggest that the inhibition of expression of key viral pathogenic proteins is likely involved in this process.

Keywords: C-Fos; Kaposi's sarcoma-associated herpesvirus; Notch1; Proliferation; Therapeutic targets.; Virus.