title: Glucocorticoid-induced activation of caspase-8 protects the glucocorticoid-induced protein Gilz from proteasomal degradation and induces its binding to SUMO-1 in murine thymocytes.
abstract: In this study, we evaluated the possible cross-talk between glucocorticoid (GC)-induced leucine zipper (Gilz) and caspase-8 in dexamethasone (Dex)-treated thymocytes. We determined that expression of Dex-induced Gilz protein was reduced when caspase-8 activity was inhibited, and this effect was not partially due to altered Gilz mRNA expression. Inhibition of the proteasome abrogated this reduction in Gilz expression, suggesting that Dex-induced caspase-8 activation protects Gilz from degradation. We hypothesized that the caspase-8-dependent protection of Gilz could be due to caspase-8-driven sumoylation. As a putative small ubiquitin-like modifier (SUMO)-binding site was identified in the Gilz sequence, we assessed whether SUMO-1 interacted with Gilz. We identified a 30-kDa protein that was compatible with the size of a Gilz-SUMO-1 complex and was recognized by the anti-SUMO-1 and anti-Gilz antibodies. In addition, Gilz bound to SUMO ubiquitin-conjugating (E2)-conjugating enzyme Ube21 (Ubc9), the specific SUMO-1 E2-conjugating enzyme, in vitro and coimmunoprecipitated with Ubc9 in vivo. Furthermore, Gilz coimmunoprecipitated with SUMO-1 both in vitro and in vivo, and this interaction depended on caspase-8 activation. This requirement for caspase-8 was further evaluated in caspase-8-deficient thymocytes and lymphocytes in which Gilz expression was reduced. In summary, our results suggest that caspase-8 activation protects Gilz from proteasomal degradation and induces its binding to SUMO-1 in GC-treated thymocytes.
authors.affiliation: Section of Pharmacology Toxicology and Chemotherapy, Department of Clinical and Experimental Medicine, University of Perugia, Perugia, Italy. midelfin@unipg.it
authors.forename: D V
authors.initials: DV
authors.lastname: Delfino
authors.forename: S
authors.initials: S
authors.lastname: Spinicelli
authors.forename: N
authors.initials: N
authors.lastname: Pozzesi
authors.forename: S
authors.initials: S
authors.lastname: Pierangeli
authors.forename: E
authors.initials: E
authors.lastname: Velardi
authors.forename: S
authors.initials: S
authors.lastname: Bruscoli
authors.forename: M P
authors.initials: MP
authors.lastname: Martelli
authors.forename: V
authors.initials: V
authors.lastname: Pettirossi
authors.forename: L
authors.initials: L
authors.lastname: Falchi
authors.forename: T-b
authors.initials: TB
authors.lastname: Kang
authors.forename: C
authors.initials: C
authors.lastname: Riccardi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology; D001665:Binding Sites; D053181:Caspase 8 / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D002478:Cells, Cultured; D003907:Dexamethasone / Q000494:pharmacology*; D005938:Glucocorticoids / Q000494:pharmacology*; D051379:Mice; D008969:Molecular Sequence Data; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D011485:Protein Binding; D025842:SUMO-1 Protein / Q000378:metabolism*; D058207:Sumoylation; D013961:Thyroid Gland / Q000166:cytology / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D044763:Ubiquitin-Conjugating Enzymes / Q000378:metabolism; D000098843:Ubiquitin-Conjugating Enzyme UBC9
chemical_list: D000906:Antibodies; C498608:Dsip1 protein, mouse; D005938:Glucocorticoids; D025842:SUMO-1 Protein; D014157:Transcription Factors; D003907:Dexamethasone; D044763:Ubiquitin-Conjugating Enzymes; D053181:Caspase 8; D046988:Proteasome Endopeptidase Complex; D000098843:Ubiquitin-Conjugating Enzyme UBC9