Regulation of Stress-Inducible Phosphoprotein 1 Nuclear Retention by Protein Inhibitor of Activated STAT PIAS1

Abstract

Stress-inducible phosphoprotein 1 (STI1), a cochaperone for Hsp90, has been shown to regulate multiple pathways in astrocytes, but its contributions to cellular stress responses are not fully understood. We show that in response to irradiation-mediated DNA damage stress STI1 accumulates in the nucleus of astrocytes. Also, STI1 haploinsufficiency decreases astrocyte survival after irradiation. Using yeast two-hybrid screenings we identified several nuclear proteins as STI1 interactors. Overexpression of one of these interactors, PIAS1, seems to be specifically involved in STI1 nuclear retention and in directing STI1 and Hsp90 to specific sub-nuclear regions. PIAS1 and STI1 co-immunoprecipitate and PIAS1 can function as an E3 SUMO ligase for STI. Using mass spectrometry we identified five SUMOylation sites in STI1. A STI1 mutant lacking these five sites is not SUMOylated, but still accumulates in the nucleus in response to increased expression of PIAS1, suggesting the possibility that a direct interaction with PIAS1 could be responsible for STI1 nuclear retention. To test this possibility, we mapped the interaction sites between PIAS1 and STI1 using yeast-two hybrid assays and surface plasmon resonance and found that a large domain in the N-terminal region of STI1 interacts with high affinity with amino acids 450-480 of PIAS1. Knockdown of PIAS1 in astrocytes impairs the accumulation of nuclear STI1 in response to irradiation. Moreover, a PIAS1 mutant lacking the STI1 binding site is unable to increase STI1 nuclear retention. Interestingly, in human glioblastoma multiforme PIAS1 expression is increased and we found a significant correlation between increased PIAS1 expression and STI1 nuclear localization. These experiments provide evidence that direct interaction between STI1 and PIAS1 is involved in the accumulation of nuclear STI1. This retention mechanism could facilitate nuclear chaperone activity.

The Hsp90 ensemble: coordinated Hsp90–cochaperone complexes regulate diverse cellular processes

“…Given the critical role of cell migration in metastasis, the idea of HOP as an attractive therapeutic target in cancer was discussed. Marco Prado (University of Western Ontario, London, Ontario, Canada) reported on HOP requirement during development by using HOP-knockout and BAC-transgenic mice, showing that HOP interacts with several nuclear proteins including PIAS1, and this promotes the retention of HOP and Hsp90 at specific subnuclear regions 10 . Whether this mechanism could facilitate nuclear-chaperone activity in cancer cells was discussed.…”

Posttranslational modifications of lysine and evolving role in heart pathologies—Recent developments

“…Five sumoylation sites were identified on stress‐inducible phosphoprotein 1 (STI1) in vitro (K123, K210, K312, K395, and K486) by using LC‐MS/MS (LTQ Orbitrap Velos; Thermo Fisher Scientific) and it was found that a direct interaction between STI1 and PIAS1 (N‐terminal region of STI1 and amino acid 450–480 sequence of PIAS1) was responsible for STI1 nuclear accumulation and retention in astrocytes rather than a modification of STI1 by sumoylation on PIAS1 overexpression. The STI1 is known to mediate the association of the molecular chaperones, HSPs Hsp70 and Hsp90 , and it was demonstrated that STI1 plays essential role in neuroprotection as it prevents ischemia‐mediated neuronal death via the prion protein PrP c .…”

Papel do complexo PrP^c-HOP e vesículas extracelulares em câncer colorretal

“…Nosso grupo mostrou que a interação específica da PrP C com proteínas de matriz extracelular, laminina e vitronectina, está envolvida em vias de sinalização levando ao desenvolvimento e plasticidade neural (figura 6) , Hajj, Lopes et al 2007. Por sua vez a interação com a co-chaperona STI1/HOP é capaz de promover proteção neuronal e plasticidade a partir da ligação com complexo PrP C -STI1/HOP com o receptor alfa-7 de acetilcolina (Martins, Graner et al 1997, Zanata, Lopes et al 2002, Beraldo, Soares et al 2013. Trabalhos envolvendo camundongos mutantes para PrP C , mostraram que mutações em sítios específicos levam a desmielinização e perda axonal severa tanto na medula espinhal como na substância branca cerebelar, ataxia grave e apoptose no cerebelo (Shmerling, Hegyi et al 1998, Radovanovic, Braun et al 2005, Caetano, Beraldo et al 2011.…”

“…HOP was initially described in yeast due to its role organizing and modulating the activity of macromolecular complexes such as heat shock protein 70 (HSP70) and HSP90 [20,21]. HOP is predominantly localized in the cytoplasm, but its presence in the nucleus has also been described [22]. Furthermore, HOP can be secreted in a soluble form or can be found associated with extracellular vesicles [23][24][25][26].…”