Characterization of Nuclear Localization and SUMOylation of the ATBF1 Transcription Factor in Epithelial Cells

Abstract

ATBF1/ZFHX3 is a large transcription factor that functions in development, tumorigenesis and other biological processes. ATBF1 is normally localized in the nucleus, but is often mislocalized in the cytoplasm in cancer cells. The mechanism underlying the mislocalization of ATBF1 is unknown. In this study, we analyzed the nuclear localization of ATBF1, and found that ectopically expressed ATBF1 formed nuclear body (NB)-like dots in the nucleus, some of which indeed physically associated with promyelocytic leukemia (PML) NBs. We also defined a 3-amino acid motif, KRK2615-2617, as the nuclear localization signal (NLS) for ATBF1. Interestingly, diffusely distributed nuclear SUMO1 proteins were sequestered into ATBF1 dots, which could be related to ATBF1's physical association with PML NBs, known SUMOylation hotspots. Furthermore, ATBF1 itself was SUMOylated. ATBF1 SUMOylation occurred at more than 3 lysine residues including K2349, K2806 and K3258 and was nuclear specific. Finally, the PIAS3 SUMO1 E3 ligase, which interacts with ATBF1 directly, diminished rather than enhanced ATBF1 SUMOylation, preventing the co-localization of ATBF1 with SUMO1 in the nucleus. These findings suggest that nuclear localization and SUMOylation are important for the transcription factor function of ATBF1, and that ATBF1 could cooperate with PML NBs to regulate protein SUMOylation in different biological processes.

MicroRNA‐133 suppresses ZFHX3‐dependent atrial remodelling and arrhythmia

“…STAT3–induced increases in oxidative stress have been implicated in AF pathogenesis . Moreover, ZFHX3 mediates the SUMOylation of misfolded and damaged proteins, which contribute to AF . Dysregulation of the ZFHX3‐Wnt‐miRNA signalling may alter calcium homeostasis .…”

“…20,21 Moreover, ZFHX3 mediates the SUMOylation of misfolded and damaged proteins, which contribute to AF. 22 Dysregulation of the ZFHX3-Wnt-miRNA signalling may alter calcium homeostasis. 23,24 ZFHX3 has been shown to cooperate with myocyte enhancer factor 2 to promote miRNA expression.…”

SUMOylation of the transcription factor ZFHX3 at Lys-2806 requires SAE1, UBC9, and PIAS2 and enhances its stability and function in cell proliferation

“…Zinc finger homeobox 3 (ZFHX3), originally named ATBF1 for AT motif binding factor 1, is a 404-kD transcription factor that comprises of 4 homeodomains, 23 zinc finger motifs, and several other domains. ZFHX3 was originally discovered as a negative transcriptional regulator of the alpha fetoprotein (AFP) gene in a hepatocellular carcinoma cell line (20), but it also plays roles in multiple pathophysiological processes such as atrial fibrillation (21), myogenic differentiation (22), embryonic development (23), circadian regulation (24), and carcinogenesis (25,26). For example, ZFHX3 is frequently mutated in advanced prostate cancer (27), deletion of Zfhx3 in mouse prostates induces and promotes neoplastic lesions, and ZFHX3 is essential for ERβ to inhibit cell proliferation via the downregulation of MYC and cyclin D1 in prostate cancer cells (28).…”

“…Biochemically, ZFHX3 can be degraded by the ubiquitin proteasome pathway; and EFP, an estrogen-responsive RING finger ubiquitin E3 ligase, mediates the ubiquitination and degradation of ZFHX3 in breast cancer cells (29). In addition, ZFHX3 can be SUMOylated endogenously (36), and expression of ZFHX3 makes diffusely distributed nuclear SUMO1 proteins form nuclear body-like structures that are associated with PML NBs (26). While SUMOylation of ZFHX3 occurs at multiple lysine residues and is nuclear specific, the PIAS3 SUMO E3 ligase, which interacts with ZFHX3 directly, diminishes rather than enhances ZFHX3 SUMOylation (26).…”

The Key Regulator for Language and Speech Development, FOXP2, is a Novel Substrate for SUMOylation

“…Currently, there are more than 20 types of PTMs. Among them, the small ubiquitin‐related modifier (SUMO) family, which is highly conserved from yeast to humans and a widely used reversible modification system, has emerged as an molecular switch for regulating various cellular pathways and biochemical processes, including cancer development and metastasis [Bellail et al, ], cell cycle regulation [Schimmel et al, ], nucleocytoplasmic translocalization [Sun et al, ], protein targeting and stability [Belaguli et al, ], signal transduction, and transcriptional regulation [Wang et al, ]. In mammalian cells, four SUMO paralogs (SUMO1 to ‐4, approximately 11 kDa proteins in size) are encoded by four distinct SUMO genes.…”