Small Ubiquitin-like Modifier (SUMO) Modification Inhibits GLI2 Protein Transcriptional Activity in Vitro and in Vivo

Lizhang Han,Yong Pan,Baolin Wang

doi:10.1074/jbc.m112.359299

Lizhang Han, Yong Pan + Show 1 more

Open Access

https://doi.org/10.1074/jbc.m112.359299

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Abstract

The Gli transcription factors are key downstream mediators of the Hedgehog (Hh) signaling pathway. How the activities of Gli transcription factors are regulated by upstream Hh signaling events and protein modifications are not fully understood. Here we show that GLI2 is conjugated by small ubiquitin-like modifier (SUMO) at lysine residues 630 and 716 in the cell. The level of GLI2 sumoylation is reduced by either mutations in six serine residues that are normally phosphorylated by protein kinase A (PKA) or stimulation by HH. This suggests that PKA phosphorylation enhances GLI2 sumoylation, whereas HH signaling inhibits it. In addition, mutation of these two lysines into arginine residues significantly increases GLI2 transcriptional activity in a cell-based reporter assay. The same mutations in the GLI2 locus also result in an increase in GLI2 activity in the mouse. Interestingly, GLI2 can interact with HDAC5 (histone deacetylase 5), but the GLI2 mutant cannot. Taken together, our results suggest that SUMO modification inhibits GLI2 transcriptional activity by recruiting HDAC5.

Highlights

How the activity of GLI2 transcriptional factor is regulated is not well understood
GLI2 Is Modified by small ubiquitin-like modifier (SUMO)—To understand the molecular mechanism by which GLI2 activity is regulated, we sought to identify the proteins that interact with the GLI2 protein
Mutations in the two-sumoylation sites result in an increase in GLI2 activity both in cell-based reporter assay and in vivo, to a lesser extent

Summary

Background

How the activity of GLI2 transcriptional factor is regulated is not well understood. Results: Loss of SUMO modification in GLI2 results in an increase in GLI2 transcriptional activity in cultured cells and in vivo. The level of GLI2 sumoylation is reduced by either mutations in six serine residues that are normally phosphorylated by protein kinase A (PKA) or stimulation by HH. This suggests that PKA phosphorylation enhances GLI2 sumoylation, whereas HH signaling inhibits it. Our results suggest that SUMO modification inhibits GLI2 transcriptional activity by recruiting HDAC5. The molecular mechanisms of inhibition of GLI2 and GLI3 processing and activation of the full-length GLI2 and GLI3 proteins are unknown Once they are activated, GLI2 and GLI3 activate the downstream transcriptional targets. Our results indicate that SUMO modification inhibits GLI2 transcriptional activity through the recruitment of HDAC protein(s)

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION