The MAZ transcription factor is a downstream target of the oncoprotein Cyr61/CCN1 and promotes pancreatic cancer cell invasion via CRAF–ERK signaling

Abstract

Myc-associated zinc-finger protein (MAZ)is a transcription factor with dual roles in transcription initiation and termination. Deregulation of MAZ expression is associated with the progression of pancreatic ductal adenocarcinoma (PDAC). However, the mechanism of action of MAZ in PDAC progression is largely unknown. Here, we present evidence that MAZ mRNA expression and protein levels are increased in human PDAC cell lines, tissue samples, a subcutaneous tumor xenograft in a nude mouse model, and spontaneous cancer in the genetically engineered PDAC mouse model. We also found that MAZ is predominantly expressed in pancreatic cancer stem cells. Functional analysis indicated that MAZ depletion in PDAC cells inhibits invasive phenotypes such as the epithelial-tomesenchymal transition, migration, invasion, and the sphere-forming ability of PDAC cells. Mechanistically, we detected no direct effects of MAZ on the expression of K-Ras mutants, but MAZ increased the activity of CRAF-ERK signaling, a downstream signaling target of K-Ras. The MAZ-induced activation of CRAF-ERK signaling was mediated via p21-activated protein kinase (PAK) and protein kinase B 2(AKT/PKB) signaling cascades and promoted PDAC cell invasiveness. Moreover, we found that the matricellular oncoprotein cysteine-rich angiogenic inducer 61 (Cyr61/CCN1) regulates MAZ expression via Notch-1-sonic hedgehog signaling in PDAC cells. We propose that Cyr61/CCN1-induced expression of MAZ promotes invasive phenotypes of PDAC cells not through direct K-Ras activation, but instead through the activation of CRAF-ERK signaling. Collectively, these results highlight key molecular players in PDAC invasiveness and may help inform therapeutic strategies to improve clinical management and outcomes of PDAC.Pancreatic ductal adenocarcinoma (PDAC) is the fourth most common cancer in the United States and the eighth worldwide (1). PDAC is associated with a shockingly poor outcome as PDAC is rarely detected in its early stages, and the aggressive phenotypes are resistant to standard therapies. Despite extensive progress in our understanding of this disease, there is still a significant unmet need for exploration and understanding the novel molecular pathways associated with PDAC progression. Elucidation of these pathways may eventually help in detection and prevention/cure of this disease by developing new therapeutic options.Recent studies found a new clue in the genesis of PDAC, and there appear to be an oncogenic role of MAZ (Myc-associated zinc finger protein and also known as SAF-1) in the pathogenesis of PDAC (2). MAZ is an inducible transcription factor that has six potential zinc fingers and a proline-rich region, which function as DNA-binding and transcriptional activation domains, respectively (3,4). MAZ is ubiquitously expressed as a transcription factor with a dual role in the initiation and termination of transcription in most tissues. These include human heart, brain, placenta, lung, liver, skeletal muscle, and pancreas (3,4). MAZ activates the expre...

CYR61/CCN1 Regulates dCK and CTGF and Causes Gemcitabine-resistant Phenotype in Pancreatic Ductal Adenocarcinoma

“…CYR61/CCN1 promotes growth and invasive phenotypes of PDAC cells via integrin-dependent noncanonical pathways. Moreover, our studies also showed that CYR61/CCN1 reprograms the epithelial-mesenchymal transition (EMT) and maintains cancer stemness/tumor-initiating properties that are associated with metastasis and drug resistance (18,21,22,(26)(27)(28)(29)(30). This provokes the hypothesis that PDAC cell-secreted CYR61/CCN1 signaling contributes to gemcitabine resistance.…”

“…We and others have previously shown that CYR61/CCN1, which belongs to the CCN family of growth factors, acts as an oncoprotein in PDAC (21)(22)(23)(24)(25). CYR61/CCN1 promotes growth and invasive phenotypes of PDAC cells via integrin-dependent noncanonical pathways.…”

“…Pancreatic cancer cell lines BxPC-3, Capan-1, AsPC-1, and Panc-1 were purchased from ATCC. The cell lines were maintained as described previously (22). Human pancreatic stellate cells (HPaS-teC) was purchased from ScienCell (ScienCell Research Laboratory) and maintained in stellate cell media and FBS.…”

“…To determine the effect of different concentrations of gemcitabine (0, 0.10, 0.25, 0.5, 1.0, and 5 mmol/L) for 72 hours on the viability of different pancreatic cancer cell lines, we performed crystal violet-based cell viability (Cell Biolabs) in a 96-well format as described earlier by us (22,31,32). The cell viability was measured by means of a Spectra Max 340 microplate reader (Molecular Devices) at a wavelength of 600 nm.…”

“…For protein analysis, cells were harvested and subjected to Western blotting as described previously (22).…”