The role of PLZF in the TLR-mediated inflammatory response and the oncogenic characteristics of renal cell carcinoma

The Promyelocytic Leukemia Zinc Finger protein (PLZF) is a transcription factor that belongs to the POK (POZ and Krüppel) family of transcriptional repressors. It functions by forming multi-protein nuclear co-repressor complexes that silence promoter activation of many tissue-specific genes. PLZF directs many major developmental processes including musculoskeletal-limb formation spermatogenesis and cellular proliferation. Previously, our group identified PLZF as a key player that is crucial for the activation of a specific subset of IFN-stimulated genes (ISGs). Here, we describe PLZF as a negative regulator of the MyD88-dependent Toll-like receptor (TLR) signaling pathway. Bone marrow-derived macrophages from wild-type and PLZF knockout mice were used as an in vitro cell model to study the effect of PLZF after stimulation with various TLR ligands. TLRs trigger production of inflammatory cytokines, thereby shaping innate and adaptive immunity to pathogens. PLZF repressed TLR-induced, NF-κB-dependent transcription by stabilizing a repressor complex encompassing the NF-κB p50 subunit and HDAC3. As a consequence, TLR-activated PLZF-null macrophages express higher levels of certain NF-κB-dependent transcripts, including those encoding the potent inflammatory cytokines TNFα, IL-12 and IL-6. Accordingly, PLZF-deficient animals are hypersensitive to septic shock and mount an exaggerated inflammatory response to Salmonella infection. These results provide new insights into mechanisms regulating selectivity in the NF-κB transcriptional program and suggest a novel strategy for therapeutic targeting of inflammatory diseases. PLZF has been identified as a tumor suppressor gene in leukemia but has not been implicated in solid tumors. Clear cell renal carcinoma is the most common type of kidney tumors in adults and is responsible for the highest mortality rate in that category. Using an in vitro system to study the effect of PLZF in two renal carcinoma cell lines (ACHN, RCC1) I found that PLZF inhibits their proliferation in culture. PLZF also reduces cell viability by inducing apoptosis and diminishes motility and cytoskeleton integrity. To investigate the mechanism used by PLZF to regulate the oncogenic features of these cells, I utilized short hairpin RNA to knockdown PLZF expression in RCC1 cells. I demonstrated that E-Cadherin expression is lost, after PLZF knockdown and the expression of both Vimentin and Snail were unregulated leading to an oncogenic phenotype. I also showed the B-Catenin localizes to the nucleus and drive the expression of different oncogenes. Furthermore, I generated a PLZF-regulated expression system in ACHN cells, where inducing PLZF by Doxycycline treatment caused the up regulation of E-Cadherin expression. Injection of RCC1 cells in NOD/SCID mice also showed that PLZF-knockdown cells formed tumors after 32 days of injection compared to the control cells which failed to form a tumor. Thus, I show a novel link between PLZF and E-Cadherin that can be further investigated to understand the role of PLZF in shaping oncogenic features of renal and other carcinomas. Thus, PLZF represents a promising therapeutic target where the invasive, aggressive and metastatic nature of tumorigenesis can be regulated.