System-Wide Changes to SUMO Modifications in Response to Heat Shock

Abstract

Covalent conjugation of the small ubiquitin-like modifier (SUMO) proteins to target proteins regulates many important eukaryotic cellular mechanisms. Although the molecular consequences of the conjugation of SUMO proteins are relatively well understood, little is known about the cellular signals that regulate the modification of their substrates. Here, we show that SUMO-2 and SUMO-3 are required for cells to survive heat shock. Through quantitative labeling techniques, stringent purification of SUMOylated proteins, advanced mass spectrometric technology, and novel techniques of data analysis, we quantified heat shock-induced changes in the SUMOylation state of 766 putative substrates. In response to heat shock, SUMO was polymerized into polySUMO chains and redistributed among a wide range of proteins involved in cell cycle regulation; apoptosis; the trafficking, folding, and degradation of proteins; transcription; translation; and DNA replication, recombination, and repair. This comprehensive proteomic analysis of the substrates of a ubiquitin-like modifier (Ubl) identifies a pervasive role for SUMO proteins in the biologic response to hyperthermic stress.

The serine/arginine-rich protein SF2/ASF regulates protein sumoylation

“…5A). Furthermore, SUMO2 and SUMO3 are required for cells to survive to hyperthermic stress (46). Depletion of SF2/ASF by siRNA greatly inhibits heat shock-induced sumoylation, pointing to SF2/ASF as a key factor in this regulatory phenomenon.…”

Quantitative Proteomics Reveals Factors Regulating RNA Biology as Dynamic Targets of Stress-induced SUMOylation in Arabidopsis

“…Important with regard to stress protection are observations that the abundance of SUMO conjugates rises dramatically and reversibly when yeast, animals, and plants are exposed to numerous environmental stresses (7,19,20). For example, a substantial increase in conjugates assembled with the SUMO1 and SUMO2 isoforms can be detected within minutes after treating Arabidopsis seedlings to a relatively mild heat shock (37°C), and this can be subsequently reversed upon a return to non-stress temperatures (7).…”

“…Such global quantification has recently become possible through advances in proteomic technologies, including the stable isotope labeling of amino acids in cell culture (SILAC) mass spectrometry (MS) approach (20,42). Although SILAC-MS has advantages for cell cultures that permit facile protein labeling via the addition of isotopically tagged amino acids to the growth medium, its use remains challenging for intact organisms generally, and for plants in particular, which naturally synthesize the full complement of amino acids (46,47).…”

“…Some of these activities are mediated through SUMO-interacting motifs (SIMs) present within binding partners (39,40). More recent proteomic studies on stress-induced SUMOylation found a strong emphasis on nuclear targets, with their collective functions implying that SUMO participates in stress protection by regulating chromatin architecture, the transcriptome, and protein quality control (15,20,(41)(42)(43)(44). In Arabidopsis, the list of over 350 SUMO1/2 targets includes histones H1 and H2B, complexes involved in DNA and histone modifications, various transcription factors and co-activators/co-repressors, RNA-binding proteins, and subunits of the nuclear pore complex, as well as various components of the SUMO pathway (41).…”

Large-scale analysis of lysine SUMOylation by SUMO remnant immunoaffinity profiling

“…Our group and others have used His-tagged SUMO to enrich potential targets using immobilized metal affinity chromatography (IMAC) under denaturing conditions [11][12][13] . Affinity purification using IMAC and/or immunoprecipitation have been successfully applied to the large-scale analysis of SUMOylated proteins from different model organisms, including Saccharomyces cerevisiae [14][15][16] , Drosophila melanogaster 17 , Arabidopsis thaliana 18 and mammalian cells 11,12,[19][20][21] . The combination of these affinity approaches with quantitative proteomics using label-free or metabolic labelling have also expanded the repertoire of protein substrates regulated by heat shock or stress conditions 18,19,[22][23][24] .…”