Modulation of the Transactivation Function and Stability of Krüppel-like Zinc Finger Protein Gli-similar 3 (Glis3) by Suppressor of Fused

Abstract

Glis3 is a member of the Glis subfamily of Krüppel-like zinc finger transcription factors. Recently, Glis3 has been linked to both type I and type II diabetes and shown to positively regulate insulin gene expression. In this study, we have identified a region within the N terminus of Glis3 that shares high levels of homology with the Cubitus interruptus (Ci)/Gli family of proteins. We demonstrated that Glis3 interacts with Suppressor of Fused (SUFU), which involves a VYGHF motif located within this conserved region. We further showed that SUFU is able to inhibit the activation of the insulin promoter by Glis3 but not the activation by a Glis3 mutant deficient in its ability to bind SUFU, suggesting that the inhibitory effect is dependent on the interaction between the two proteins. Exogenous SUFU did not affect the nuclear localization of Glis3; however, Glis3 promoted the nuclear accumulation of SUFU. Additionally, we demonstrated that SUFU stabilizes Glis3 in part by antagonizing the Glis3 association with a Cullin 3-based E3 ubiquitin ligase that promotes the ubiquitination and degradation of Glis3. This is the first reported instance of Glis3 interacting with SUFU and suggests a novel role for SUFU in the modulation of Glis3 signaling. Given the critical role of Glis3 in pancreatic ␤-cell generation and maintenance, the elevated Glis3 expression in several cancers, and the established role of SUFU as a tumor suppressor, these data provide further insight into Glis3 regulation and its function in development and disease.

GLIS3 is indispensable for TSH/TSHR-dependent thyroid hormone biosynthesis and follicular cell proliferation

“…Several kinase pathways activated by TSH/TSHR have been implicated in mediating the downstream effects of TSH on gene expression, proliferation, and TH biosynthesis. We previously reported that GLIS3 undergoes a number of posttranslational modifications, including phosphorylation and ubiquitination, that can regulate GLIS3 transcriptional activity (54,55). This raised the possibility of a connection between TSH/TSHR-activated kinase pathways and the regulation of GLIS3 activity.…”

“…The expression vectors p-3′FLAG-CMV-Glis3-HA, p-3′FLAG-CMV-Glis3ΔN496-HA, p-3′FLAG-CMV-Glis3ΔC480-HA, and pCMV-SUFU-Myc were described previously (54,55). pcDNA3.1-Pax8 was obtained from GenScript, and pcDNA3.1-TTF1 was a gift from David Mu (Pennsylvania State University, Hershey, PA, USA) (Addgene plasmid, catalog 49989).…”

Emerging roles of GLIS3 in neonatal diabetes, type 1 and type 2 diabetes

“…Subsequently, co-immunoprecipitation assay confirmed that the SUFU was able to interact with GLIS3 through the conserved VYGHF motif at N-terminus of GLIS3. Functional study showed that the SUFU repressed the activation of the insulin promoter mediated by GLIS3 (ZeRuth et al 2011). In addition, the E3 ubiquitin ligase scaffolding protein Cullin 3 (CUL3) promotes GLIS3 polyubiquitination and degradation via the 26S proteasomal pathway.…”

“…SUFU was shown to inhibit the association of CUL3 with GLIS3, thereby protecting GLIS3 protein from proteolytic degradation and enhancing the stability of GLIS3 protein (Fig. 1A) (ZeRuth et al 2011).…”

GLIS1–3 transcription factors: critical roles in the regulation of multiple physiological processes and diseases

“…The ZFDs of GLIS1 and GLIS3 exhibit the highest homology suggesting that they are evolutionary the most closely related. Outside their ZFD, GLIS1–3 exhibit little homology with each other or GLI proteins with the exception of an N-terminal, conserved region (NCR) of about 60 aa that GLIS3 has in common with GLI1–3 [9]. …”