Figure 2.

Multiple regulatory signaling pathways of NADPH oxidases (NOXs/RBOHs) in plant immunity of Arabidopsis. Botrytis-induced kinase1 (BIK1), a protein of the RLCKVII subfamily member, is the central immune regulator for multiple signaling pathways from upstream to downstream to trigger an oxidative burst from RbohD. BIK1 can directly bind to multiple RLKs/PRRs in the resting state, such as flagellin sensing 2 (FLS2), elongation factor-Tu receptor (EFR), and PEP 1 receptor (PEPR1) [180,181,182]. These RLKs all associate with the regulatory LRR-receptor kinase BRI1-associated receptor kinase 1 (BAK1) (also known as SERK3) and form the immune receptor complexes upon bacterial flagellin (flg22), bacterial elongation factor-Tu (elf18 or elf26) or the endogenous AtPep1 (and related peptides) by interacting with them. When flg22 or elf18 are recognized by FLS2 or EFR respectively, the latter rapidly recruit the co-receptor BAK1 for trans-phosphorylation events between BIK1 and BAK1 [193,194]. Other RLK members, FER (FERONIA) and ANX1/2 (ANXURs), belonging to malectin-like receptor kinases, also known as Catharanthus roseus L. receptor-like kinase 1-like proteins (CrRLK1Ls), also participate in NOXs/RBOHs regulation during plant immune response [188]. FER acts as a Rapid Alkalinization Factors (RALF)-regulated scaffold that modulates receptor kinase complex assembly [189], while ANX1 and possibly ANX2 negatively regulate pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) by putatively competing with FLS2 for interaction with BAK1 [190]. Where, RALF23/33, the secreted peptides, play the negative role in repressing FER signaling during PTI responses [189]. In addition, a LORELEI-like GPI-anchored protein 1 (LLG1), as a cooperator of FER, facilitates FER-FLS2-BAK1 ligand-induced receptor complex formation to activate BIK1, and subsequently phosphorylates the ROS-producing RbohD [190]. Another important immune complex composes of CERK1 (the homologs of BAK1) and two LysM proteins, LYM1 and LYM3 (the homologs of LYP4 and LYP6 in rice), upon recognizing PGN (peptidoglycan) by directly binding to LYM1 and LYM3 [200], but this immune complex does not contribute to chitin signaling in Arabidopsis [201]. Meanwhile, a new LysM-containing receptor like kinase 5 was found that binds chitin at a much higher affinity than CERK and forms a chitin-dependent complex with CERK1 [202]. These immune complexes in which AtCERK1 is involved also activate BIK1 by phosphorylation directly. The activated BIK1 (GTP-BIK1) by immune complexes then directly phosphorylates and activates AtRbohD [182,195]. However, AtCERK1-associated PBL27 (an ortholog of OsRLCK185) is not involved in chitin-induced ROS production [203], but it regulates the activation of the MAPK cascade by phosphorylation of AtMAPKKK5 [204] and supervenes with tier phosphorylation events: between AtMAPKKK5 and AtMKK4/5 and AtMPK3/6 in PAMPs signaling [203]. Besides BIK1 and PBL27, there are other RLCKs, such as BSK1, PCRK1, and PBL1, that are also genetically required for a PAMP-triggered ROS burst. PBL1, a close homolog of receptor-like cytoplasmic kinases of BIK1, acting as the molecular chaperone of BIK1, is also important for PTI signaling potentially via interactions with FLS2 at rest state [187]. BIK1 and PBL1 play a positive role in the RbohD-dependent ROS production but are not required for MAPK activation [181,196]. Besides controlling RbohD, BIK1 and PBL1 are also required for the PAMP and/or DAMP-triggered cytosolic Ca²⁺ burst that precedes ROS production [99,178,197]. Just recently, SIK1, a mitogen-activated protein kinase kinase kinase kinase (MAP4K) family member, was found to directly interact with and phosphorylate RbohD to promote the extracellular ROS burst upon flagellin perception. Moreover, SIK1 interacts with and stabilizes BIK1 by direct phosphorylation at rest state [200]. In other words, SIK1 positively regulates immunity not only by binding to and activating RbohD directly, but also indirectly through the BIK1 mediated ways. In yeast and humans, MAP4Ks can directly activate MAPK cascades [205,206]. In addition, in parallel with ROS production, PTI also induces MAPK activation [191,207]. Two regulation pathways of NOX/RBOH activity mediated by MAPK might exist in plant immune response: elicitor-MAP4K (SIK1)-RbohD or/and elicitor-MAP4K (SIK1)-MAPK cascades-RBOH. The regulation imposed by BIK1 is a common regulatory mechanism for RbohD during immune response, and in which BIK1 is the central immune regulator for multiple signaling from upstream to downstream to trigger an oxidative burst from RbohD. Therefore, its protein stability is tightly regulated to ensure appropriate and robust immune activation. PUB25 and PUB26 (E3 ligases homologous) are the crucial factors which directly target BIK1 for degradation by the ubiquitin proteasome system, while the activity of PUB25/26 were negatively or positively regulated by the directly binding with G protein or the phosphorylation from CDPK28, respectively. Interestingly, PUB25/26 specifically target non-activated BIK1 (GDP-BIK1), suggesting that activated BIK1 (GTP-BIK1) is maintained for immune signaling [195]. The member of MAP4K, SIK1, can also phosphorylate directly and enhance BIK1 stability at a resting state by coupling with heterotrimeric G proteins [200]. In addition, ATP can also be released into the extracellular matrix and referred to as extracellular ATP (eATP), functioning in signaling. Its mediated stomata close is NOX-dependent under bacterial and fungal infection, and during the process, a L-type lectin receptor-like kinase (LecRKI.9) DORN1 acts as the receptor of eATP to directly phosphorylate downstream of AtRbohD [85]. BAK1, BRI1-associated receptor kinase 1; BIK1, botrytis-induced kinase1; BSK1, brassinolide-signaling kinase1; CDPK, calcium-dependent protein kinase; CERK, chitin-elicitor receptor kinase; DAMP, damage associated molecular pattern; DORN1, a L-type lectin receptor-like kinase; eATP, extracellular ATP; EFR, elongation factor-Tu receptor; elf18, a bacterial elongation factor-Tu; FLS2, flagellin sensing 2; flg22, a bacterial flagellin; G_pro, GTP-binding protein; LLG1, LORELEI-like GPI-anchored protein 1; LRR, leucine-rich repeat; LYK, LysM-containing receptor-like kinase; MAPK, mitogen-activated protein kinase; PAMP, pathogen-associated molecular pattern; PBL, PBS-like kinase; PCRK1, pattern-triggered immunity compromised receptor-like cytoplasmic kinase 1; Pep, plant elicitor peptide; PEPR1, phosphoenolpyruvic acid 1 receptor; PGN, peptidoglycan; PRR, pattern recognition receptor; PTI, PAMP-triggered immunity; RALF, rapid alkalinization factors; RLCK, receptor-like cytoplasmic kinases; RLK, receptor-like protein kinase; SIK1, MAP4K, salt inducible kinase 1.