Impact of MAPK Cascade Pathway and P53 Pathway upon Liver Transplant

Summary： The change and the role of MAPK cascade pathway and P53 pathway after liver transplantation were explored. Thirty-four punctured donor liver specimens and 10 normal liver specimens were classified as group A （no rejection, n=10）, group B （mild/moderate acute rejection, n=10）, group C （serious acute rejection, n=8）, group D （chronic rejection/fibrosis, n=6） and group E （control, n= 10）. By using immunohistochemistry, the expression levels of mitogen activated protein kinase （MAPK）, Ras and P53 proteins, and by in situ hybridization, MAPK and ras mRNA expression levels were detected. The results showed that the expression levels of MAPK and Ras proteins were increased by turns in groups A, B and C, and decreased by turns in groups D and E. The protein expression of P53 was higher in the treated groups. The expression of Ras, HSP70 mRNA was identical as that of protein. It is suggested that the MAPK cascade pathway and P53 pathway can protect the hepatocytes by different mechanisms after liver transplantation. MAPKs cascade pathway repairs hepatocyte injury or accelerates hepatocytes into proliferation or differentiation. P53 pathway blocks cell cycle within G1 phase to make hepatocyte repair or apoptosis to reduce disorder differentiation.

Arabidopsis EXTRA-LARGE G PROTEIN 1(XLG1) functions together with XLG2 and XLG3 in PAMP-triggered MAPK activation and immunity

Pattern-triggered immunity(PTI) is an essential strategy used by plants to deploy broad-spectrum resistance against pathogen attacks. Heterotrimeric G proteins have been reported to contribute to PTI.Of the three non-canonical EXTRA-LARGE G PROTEINs(XLGs) in Arabidopsis thaliana, XLG2 and XLG3 were shown to positively regulate immunity,but XLG1 was not considered to function in defense,based on the analysis of a weak xlg1 allele.In this study, we characterized the xlg1 xlg2 xlg3 triple knockout mutants generated from an xlg1 knockout allele. The strong xlg1 xlg2 xlg3 triple mutants compromised pathogen-associated molecular pattern(PAMP)-triggered activation of mitogen-activated protein kinases(MAPKs) and resistance to pathogen infection. The three XLGs interacted with MAPK cascade proteins involved in defense signaling, including the MAPK kinase kinases MAPKKK3 and MAPKKK5, the MAPK kinases MKK4 and MKK5, and the MAPKs MPK3 and MPK6. Expressing a constitutively active form of MKK4 restored MAPK activation and partially recovered the compromised disease resistance seen in the strong xlg1 xlg2 xlg3 triple mutant. Furthermore, mutations of all three XLGs largely restored the phenotype of the autoimmunity mutant bak1-interacting receptor-like kinase 1. Our study reveals that all three XLGs function redundantly in PAMP-triggered MAPK activation and plant immunity.

OsWRKY78 regulates panicle exsertion via gibberellin signaling pathway in rice

Panicle exsertion is one of the crucial agronomic traits in rice(Oryza sativa).Shortening of panicle exsertion often leads to panicle enclosure and severely reduces seed production.Gibberellin(GA)plays important roles in regulating panicle exsertion.However,the underlying mechanism and the relative regulatory network remain elusive.Here,we characterized the oswrky78 mutant showing severe panicle enclosure,and found that the defect of oswrky78 is caused by decreased bioactive GA contents.Biochemical analysis demonstrates that OsWRKY78 can directly activate GA biosynthesis and indirectly suppress GA metabolism.Moreover,we found OsWRKY78 can interact with and be phosphorylated by mitogen-activated protein kinase(MAPK)kinase OsMAPK6,and this phosphorylation can enhance OsWRKY78 stability and is necessary for its biological function.Taken together,these results not only reveal the critical function of OsWRKY78,but also reveal its mechanism via mediating crosstalk between MAPK and the GA signaling pathway in regulating panicle exsertion.

Sporophytic control of tapetal development and pollen fertility by a mitogen-activated protein kinase cascade in rice

Tapetum,the innermost layer of the anther wall,provides essential nutrients and materials for pollen development.Timely degradation of anther tapetal cells is a prerequisite for normal pollen development in flowering plants.Tapetal cells facilitate male gametogenesis by providing cellular contents after highly coordinated programmed cell death(PCD).Tapetal development is regulated by a transcriptional network.However,the signaling pathway(s)involved in this process are poorly understood.In this study,we report that a mitogen-activated protein kinase(MAPK)cascade composed of OsYDA1/OsYDA2-OsMKK4-OsMPK6 plays an important role in tapetal development and male gametophyte fertility.Loss of function of this MAPK cascade leads to anther indehiscence,enlarged tapetum,and aborted pollen grains.Tapetal cells in osmkk4 and osmpk6 mutants exhibit an increased presence of lipid body-like structures within the cytoplasm,which is accompanied by a delayed occurrence of PCD.Expression of a constitutively active version of OsMPK6(CA-OsMPK6)can rescue the pollen defects in osmkk4 mutants,confirming that OsMPK6 functions downstream of OsMKK4 in this pathway.Genetic crosses also demonstrated that the MAPK cascade sporophyticly regulates pollen development.Our study reveals a novel function of rice MAPK cascade in plant male reproductive biology.

OsCERK1-Mediated Chitin Perception and Immune Signaling Requires Receptor-like Cytoplasmic Kinase 185 to Activate an MAPK Cascade in Rice

Conserved pathogen-associated molecular patterns （PAMPs）, such as chitin, are perceived by pattem recognition receptors （PRRs） located at the host cell surface and trigger rapid activation of mitogen- activated protein kinase （MAPK） cascades, which are required for plant resistance to pathogens. However, the direct links from PAMP perception to MAPK activation in plants remain largely unknown. In this study, we found that the PRR-associated receptor-like cytoplasmic kinase Oryza sativa RLCK185 transmits immune signaling from the PAMP receptor OsCERK1 to an MAPK signaling cascade through interaction with an MAPK kinase kinase, OsMAPKKKε, which is the initial kinase of the MAPK cascade. OsRLCK185 interacts with and phosphorylates the C-terminal regulatory domain of OsMAPKKKε. Coexpression of phosphomi- metic OsR LCK185 and OsMAPKKKε activates MAPK3/6 phosphorylation in Nicotiana benthamiana leaves. Moreover, OsMAPKKKε interacts with and phosphorylates OsMKK4, a key MAPK kinase that transduces the chitin signal. Overexpression of OsMAPKKKε increases chitin-induced MAPK3/6 activation, whereas OsMAPKKKε knockdown compromises chitin-induced MAPK3/6 activation and resistance to rice blast fungus. Taken together, our results suggest the existence of a phospho-signaling pathway from cell surface chitin perception to intraceilular activation of an MAPK cascade in rice.

The rice Raf-like MAPKKK OsILA1 confers broad-spectrum resistance to bacterial blight by suppressing the OsMAPKK4-OsMAPK6 cascade

Mitogen-activated protein kinase kinase kinase(MAPKKK)are the first components of MAPK cascades,which play pivotal roles in signaling during plant development and physiological processes.The genome of rice encodes 75 MAPKKKs,of which 43 are Raf-like MAPKKKs.The functions and action modes of most of the Raf-like MAPKKKs,whether they function as bona fide MAPKKKs and which are their downstream MAPKKs,are largely unknown.Here,we identified the osmapkkk43 mutant,which conferred broad-spectrum resistance to Xanthomonas oryzae pv.oryzae(Xoo),the destructive bacterial pathogen of rice.Oryza sativa(Os)MAPKKK43 encoding a Raf-like MAPKKK was previously known as Increased Leaf Angle 1(OsILA1).Genetic analysis indicated that OsILA1 functioned as a negative regulator and acted upstream of the OsMAPKK4-OsMAPK6 cascade in rice-Xoo interactions.Unlike classical MAPKKKs,OsILA1 mainly phosphorylated the threonine 34 site at the N-terminal domain of OsMAPKK4,which possibly influenced the stability of OsMAPKK4.The N-terminal domain of OsILA1 is required for its homodimer formation and its full phosphorylation capacity.Taken together,our findings reveal that OsILA1 acts as a negative regulator of the OsMAPKK4-OsMAPK6 cascade and is involved in rice-Xoo interactions.

Overlapping functions of YDA and MAPKKK3/MAPKKK5 upstream of MPK3/MPK6 in plant immunity and growth/development

Arabidopsis MITOGEN-ACTIVATED PROTEIN KINASE3(MAPK3 or MPK3)and MPK6 play important signaling roles in plant immunity and growth/development.MAPK KINASE4(MKK4)and MKK5 function redundantly upstream of MPK3 and MPK6 in these processes.YODA(YDA),also known as MAPK KINASE KINASE4(MAPKKK4),is upstream of MKK4/MKK5 and forms a complete MAPK cascade(YDA–MKK4/MKK5–MPK3/MPK6)in regulating plant growth and development.In plant immunity,MAPKKK3 and MAPKKK5 function redundantly upstream of the same MKK4/MKK5–MPK3/MPK6 module.However,the residual activation of MPK3/MPK6 in the mapkkk3 mapkkk5 double mutant in response to flg22 pathogen-associated molecular pattern(PAMP)treatment suggests the presence of additional MAPKKK(s)in this MAPK cascade in signaling plant immunity.To investigate whether YDA is also involved in plant immunity,we attempted to generate mapkkk3 mapkkk5 yda triple mutants.However,it was not possible to recover one of the double mutant combinations(mapkkk5 yda)or the triple mutant(mapkkk3 mapkkk5 yda)due to a failure of embryogenesis.Using the clustered regularly interspaced short palindromic repeats(CRISPR)–CRISPRassociated protein 9(Cas9)approach,we generated weak,N-terminal deletion alleles of YDA,yda-del,in a mapkkk3 mapkkk5 background.PAMP-triggered MPK3/MPK6 activation was further reduced in the mapkkk3 mapkkk5 yda-del mutant,and the triple mutant was more susceptible to pathogen infection,suggesting YDA also plays an important role in plant immune signaling.In addition,MAPKKK5 and,to a lesser extent,MAPKKK3 were found to contribute to gamete function and embryogenesis,together with YDA.While the double homozygous mapkkk3 yda mutant showed the same growth and development defects as the yda single mutant,mapkkk5 yda double mutant and mapkkk3 mapkkk5 yda triple mutants were embryo lethal,similar to the mpk3 mpk6 double mutants.These results demonstrate that YDA,MAPKKK3,and MAPKKK5 have overlapping functions upstream of the MKK4/MKK5–MPK3/MPK6 module in both plant immunity and growth/development.

The MEKK1-MKK1/2-MPK4 cascade phosphorylates and stabilizes STOP1 to confer aluminum resistance in Arabidopsis

Aluminum(Al)toxicity can seriously restrict crop production on acidic soils,which comprise 40%of the world’s potentially arable land.The zinc finger transcription factor STOP1 has a conserved and essential function in mediating plant Al resistance.Al stress induces STOP1 accumulation via post-transcriptional regulatory mechanisms.However,the upstream signaling pathway involved in Al-triggered STOP1 accumulation remains unclear.Here,we report that the MEKK1-MKK1/2-MPK4 cascade positively regulates STOP1 phosphorylation and stability.Mutations of MEKK1,MKK1/2,or MPK4 lead to decreased STOP1 stability and Al resistance.Al stress induces the kinase activity of MPK4,which interacts with and phosphorylates STOP1.The phosphorylation of STOP1 reduces its interaction with the F-box protein RAE1 that mediates STOP1 degradation,thereby leading to enhanced STOP1 stability and Al resistance.Taken together,our results suggest that the MEKK1-MKK1/2-MPK4 cascade is important for Al signaling and confers Al resistance through phosphorylation-mediated enhancement of STOP1 accumulation in Arabidopsis.

Role of c-Jun NH_(2)-terminal kinase-mediated mitogen-activated protein kinase pathway in response to pesticides in Apis cerana cerana

The mitogen-activated protein kinase(MAPK)cascade pathway plays an important role in regulating stress responses.The function of the c-Jun NH_(2)-terminal kinase(JNK),a component of the MAPK cascade pathway,in Apis cerana cerana(Acc)remains unclear.Here,JNK was isolated and identified from Acc.Bioinformatics analyses revealed there is a typical serine/threonine protein kinase catalytic domain in the AccJNK protein.An expression profile analysis showed that AccJNK was significantly induced by pesticide treatments.To further explore the functional mechanisms of AccJNK,a yeast 2-hybrid screen was performed,activator protein-1(AP-1)was screened as the interaction partner of AccJNK,and the interaction relationship was further verified by pull-down assay.Quantitative real-time polymerase chain reaction showed the expression pattern of AccAP-I was similar to that of AccJNK.After a knockdown of AccJNK or AccAP-I by RNA interference,the survival rate of Acc after pesticide treatments increased.Additionally,the expression levels of antioxidant-related genes and the activities of antioxidant enzymes increased,suggesting that the knockdown of AccJNK or AccAP-I increased the antioxidant capacity of bees.Our study revealed that the JNK-mediated MAPK pathway responds to pesticide stress by altering the antioxidant capacity of Acc.

Protein kinases in plant responses to drought, salt,and cold stress

Protein kinases are major players in various signal transduction pathways. Understanding the molecular mechanisms behind plant responses to biotic and abiotic stresses has become critical for developing and breeding climate-resilient crops. In this review,we summarize recent progress on understanding plant drought, salt, and cold stress responses, with a focus on signal perception and transduction by different protein kinases, especially sucrose nonfermenting1(SNF1)-related protein kinases(Sn RKs),mitogen-activated protein kinase(MAPK) cascades,calcium-dependent protein kinases(CDPKs/CPKs),and receptor-like kinases(RLKs). We also discuss future challenges in these research fields.

Arabidopsis E3 ligase KEG associates with and ubiquitinates MKK4 and MKK5 to regulate plant immunity

Mitogen-activated protein kinase(MAPK) cascades are highly conserved signaling modules that regulate plant immune responses. The Arabidopsis thaliana Raf-like MAPK kinase kinase ENHANCED DISEASE RESISTANCE1(EDR1) is a key negative regulator of plant immunity that affects the protein levels of MKK4 and MKK5, two important MAPK cascade members, but the underlying mechanism is poorly understood. Here, genome-wide phosphorylation analysis demonstrated that the E3 ligase KEEP ON GOING(KEG) is phosphorylated in the edr1 mutant but not the wild type, suggesting that EDR1 negatively affects KEG phosphorylation. The identified phosphorylation sites in KEG appear to be important for its accumulation. The keg-4 mutant, a previously identified edr1 suppressor, enhances susceptibility to the powdery mildew pathogen Golovinomyces cichoracearum. In addition, MKK4 and MKK5 protein levels are reduced in the keg-4 mutant. Furthermore,we demonstrate that MKK4 and MKK5 associate with full-length KEG, but not with truncated KEG-RK or KEG-RKA, and that KEG ubiquitinates and mediates the degradation of MKK4 and MKK5. Taken together, these results indicate that MKK4 and MKK5 protein levels are regulated by KEG via ubiquitination, uncovering a mechanism by which plants finetune immune responses by regulating the homeostasis of key MAPK cascade members via ubiquitination and degradation.

Expression of the inactive ZmMEK1 induces salicylic acid accumulation and salicylic acid-dependent leaf senescence

Leaf senescence is the final leaf developmental process that is regulated by both intracellular factors and environmental conditions. The mitogen-activated protein kinase（MAPK） signaling cascades have been shown to play important roles in regulating leaf senescence; however, the component（s） downstream of the MAPK cascades in regulating leaf senescence are not fully understood. Here we showed that the transcriptions of Zm MEK1, Zm SIMK1, and Zm MPK3 were induced during dark-induced maize leaf senescence.Furthermore, in-gel kinase analysis revealed the 42 k Da MAPK was activated. Zm MEK1 interacted with Zm SIMK1 in yeast and maize mesophyll protoplasts and Zm SIMK1 was activated by Zm MEK1 in vitro. Expression of a dominant negative mutant of Zm MEK1 in Arabidopsis transgenic plants induced salicylic acid（SA） accumulation and SA-dependent leaf senescence.Zm MEK1 interacted with Arabidopsis MPK4 in yeast and High-Im pact activated MPK4 in vitro. SA treatment accelerated darkinduced maize leaf senescence. Moreover, blockage of MAPK signaling increased endogenous SA accumulation in maize leaves. These findings suggest that Zm MEK1-Zm SIMK1 cascade and its modulating SA levels play important roles in regulating leaf senescence.

The warfare for plant highway:vascular plant–microbe interaction pinpoints lignin

Plant vascular pathogens are one kind of destructive pathogens in agricultural production.However,mechanisms behind the vascular pathogen-recognition and the subsequent defense responses of plants are not well known.A recent pioneering study on plant vascular immunity discovered a conserved MKP1-MPK-MYB signaling cascade that activates lignin biosynthesis in vascular tissues to confer vascular resistance in both monocot rice and the dicot Arabidopsis.The breakthrough provides a novel view on plant immunity to vascular pathogens and offers a potential strategy for the future breeding of disease-resistant crops.