Arabidopsisinositol polyphosphate kinases regulate COP9 signalosome deneddylase functions in phosphate-homeostasis

Abstract

Targeted protein degradation is essential for physiological development and adaptation to stress. Mammalian INOSITOL PENTAKISPHOSPHATE KINASE (IP5K) and INOSITOL HEXAKISPHOSPHATE KINASE 1 (IP6K1) pair modulates functions of Cullin RING Ubiquitin E3 ligases (CRLs) that execute targeted degradation of substrates. Coordinated activities of these kinases protect CRLs on a COP9 signalosome (CSN) platform and stimulates deneddylation-dependent disassembly to maintain continuity of its functions. In plants, CRL regulations on CSN by inositol phosphate (InsP) kinases are not known. Here, we show interactions of Arabidopsis thaliana INOSITOL PENTAKISPHOSPHATE 2-KINASE 1 (IPK1) and INOSITOL 1,3,4-TRISPHOSPHATE 5/6-KINASE 1 (ITPK1), counterparts of the above InsP-kinase pair, with CSN subunits and its positive influences on the dynamics of cullin deneddylation. We identify neddylation enhancements on CRLs as an early response to phosphate-starvation and its orchestration by perturbed IPK1/ITPK1 activities. At a molecular level, specific kinetics of CSN5 deneddylase is affected by the above InsP-kinases. Overall, our data reveal conserved InsP-kinase involvements on CRL-CSN synergism in plants.

Arabidopsis inositol polyphosphate kinases IPK1 and ITPK1 modulate crosstalk between SA-dependent immunity and phosphate-starvation responses

“…In the respective complemented lines ipk1-1:Myc-IPK1 (Walia et al, 2020) or itpk1-2:ITPK1-GFP (Laha et al, 2020), growth defects were abolished and the transgenic plants were indistinguishable to Col-0 (Figure 1a; Figure S1). Further, constitutive expression of phosphate-starvation-inducible (PSI) genes and resulting elevated Pi accumulations previously known for ipk1-1 or itpk1-2 (Kuo et al, 2014;Kuo et al, 2018) were also restored to Col-0 levels in the complemented lines (Figure S2a) (Laha et al, 2020;Walia et al, 2020). Overall, these results validated the corresponding T-DNA mutational consequences on the abnormal growth phenotypes and constitutive PSR in ipk1-1 or itpk1-2.…”

Arabidopsis inositol polyphosphate kinases IPK1 and ITPK1 modulate crosstalks between SA-dependent immunity and phosphate-starvation responses

“…In the respective complemented lines ipk1-1:Myc-IPK1 (Walia et al ., 2020) or itpk1-2:ITPK1-GFP (Laha et al ., 2020), growth defects were abolished and the transgenic plants were indistinguishable to Col-0 (Figure 1a; Figure S1). Further, constitutive expression of phosphate-starvation-inducible (PSI) genes and resulting elevated Pi accumulations previously known for ipk1-1 or itpk1-2 (Kuo et al ., 2014; Kuo et al ., 2018) were also restored to Col-0 levels in the complemented lines (Figure S2a) (Laha et al ., 2020; Walia et al ., 2020). Overall, these results validated the corresponding T-DNA mutational consequences on the abnormal growth phenotypes and constitutive PSR in ipk1-1 or itpk1-2 .…”

“…Indeed, in phr1 plants this increase was barely detected suggesting that these responses are primarily PHR1-orchestrated. We recently reported that IPK1 and ITPK1 protein stabilities are increased during PSR perhaps signifying their intersection on diverse signaling pathways (Walia et al ., 2020). Likewise, upon PstDC3000 challenge on ipk1-1 : Myc-IPK1 or itpk1-2 : ITPK1-GFP plants we noted improvements in IPK1 and ITPK1 protein stabilities likely reflective of a similar phenomenon (Figure 6f).…”

“…We showed that CSN activities in turn maintain cellular Pi-homeostasis by regulating the Cullin RING Ligases (CRLs) functioning in targeted-degradation of substrates by the 26S proteasome complex. Functional deficiencies in CSN result in strong pleiotropic effects including auxin-insensitivity (Dohmann et al ., 2008), constitutive activation of PSR (Walia et al, 2020), and impaired JA-defenses accompanied by elevated PR1 transcripts (Hind et al ., 2011), thus revealing strong parallel with ipk1-1 or itpk1-2 defects. Rice SPX4, SNC1 or NPR1 stabilities are monitored by CRL functions placing molecular implications of IPK1/ITPK1 on defense outcomes via CSN-CRL dynamics (Gou et al ., 2012; Ruan et al ., 2019; Ried et al ., 2019; Shen et al ., 2020).…”

“…These phenotypic defects were more pronounced under long-day (LD) conditions of growth. In the respective complemented lines ipk1-1:Myc-IPK1 (Walia et al, 2020) or itpk1-2:ITPK1-GFP (Laha et al, 2020), growth defects were abolished and the transgenic plants were indistinguishable to Col-0 (Figure 1a; Figure S1). Further, constitutive expression of phosphate-starvation-inducible (PSI) genes and resulting elevated Pi accumulations previously known for ipk1-1 or itpk1-2 (Kuo et al, 2014;Kuo et al, 2018) were also restored to Col-0 levels in the complemented lines (Figure S2a) (Laha et al, 2020;Walia et al, 2020).…”