Engrailed-2 promotes a malignant phenotype of esophageal squamous cell carcinoma through upregulating the expression of pro-oncogenic genes

Abstract

Background A number of homeobox genes have been implicated in the development of various cancers. However, the role of engrailed 2 (EN2), a member of the homeobox gene superfamily, in esophageal squamous cell carcinoma (ESCC) remains unknown. Methods The expression of EN2 was examined using quantitative real-time PCR and immunohistochemistry. A stable cell line was established to express exogenous EN2 using a lentivirus system. The malignant phenotype was analyzed with proliferation, clonogenicity, wound-healing and invasion assays. The CRISPR/Cas9 system was adopted to deplete endogenous EN2. RNA profiling was performed using gene expression microarray. The ShRNA-mediated method was used to knock down the expression of SPARC. The structure-function relationship was determined using site-directed mutagenesis. Results EN2 is highly expressed in ESCC. The malignant phenotype of the ESCC cell line was amplified by an overexpression of EN2 but was attenuated by a disruption of EN2. RNA profiling analysis revealed that distinct sets of genes were modulated by the expression of EN2 in various ESCC cell lines and oncogenes were among these. EN2 greatly increased the expression of SPARC in Eca109. Site-directed mutagenesis revealed that the induction of SPARC was closely correlated with the protumor function of EN2. ShRNA-mediated knockdown of SPARC attenuated the malignant phenotype of EN2-infected cells. These data suggest that SPARC is crucial for mediating the protumor function of EN2. Discussion EN2 has an oncogenic function in ESCC that is mediated by upregulating the expression of pro-oncogenic genes downstream. EN2 may potentially act as a diagnostic marker or therapeutic target for ESCC treatment in the future.

MiR-33a targets FOSL1 and EN2 as a clinical prognostic marker for sarcopenia by glioma

“…In addition, disorders of EN2 regulation can lead to abnormal cell proliferation, leading to tumorigenesis. Studies show that EN2 plays an important role in the proliferation, migration, and invasion of various tumor cells, including prostate cancer, colorectal cancer, esophageal squamous cell carcinoma, lung cancer, and bladder cancer, and is closely related to the poor prognosis of tumor patients ( Zhou et al, 2017 ; Lin et al, 2018 ; Cao et al, 2020 ; Li et al, 2020 ; Li et al, 2021 ). EN2 promotes the proliferation and invasion of colorectal cancer cells by regulating CCL20, thus promoting the progression of colorectal cancer.…”

“…EN2 also promotes the invasion and metastasis of esophageal squamous cell carcinoma by upregulating SPARC expression. It promotes the proliferation, invasion, and metastasis of bladder cancer cells by activating the PI3K/Akt pathway and inhibiting the PTEN gene ( Cao et al, 2020 ; Li et al, 2020 ) ( Li et al, 2021 ). In addition, it was found that EN2 expression levels are also correlated with the degree of malignancy of gliomas and promoted the malignant progression of glioma ( Zeng et al, 2020 ).…”

A new allele of <i>engrailed</i>, <i>en^NK14</i>, causes supernumerary spermathecae in <i>Drosophila melanogaster</i>

“…The pleiotropy of En (McGrath et al, 2013;Wizenmann et al, 2015;Punia et al, 2019;Cao et al, 2020) gives rise to a wide variety of mutation phenotypes of en, such as scutellar cleft, malformation of the wing, an extra sex comb and lethality in D. melanogaster (Lindsley and Zimm, 1992). Two spermathecae arise from the anterior compartment of female genital primordia of the A8 segment in the wild type female genital disc during the pupal stage.…”

“…En is well known as a homeodomaincontaining transcription factor that plays important roles in boundary formation (Kornberg, 1981;Kornberg et al, 1985). En family proteins were recently revealed to be considerably pleiotropic, having three primary functions in many metazoans: transcription regulation, translation control and action in extracellular signaling pathways (reviewed by McGrath et al, 2013;Wizenmann et al, 2015;Punia et al, 2019;Cao et al, 2020). However, the molecular basis of numerical variation in spermathecae is yet to be elucidated.…”

Engrailed 2 serves as a master regulator of the super‐enhancer in the TNC gene locus in non‐small cell lung cancer

“…We have previously reported 27 that EN2 has an oncogenic role in esophageal squamous cell carcinoma. In the present work, we showed that the expression of EN2 was increased in tissue samples derived from patients with non-small cell lung cancer (NSCLC).…”