Do rainfed production systems have lower environmental impact over irrigated production systems?: On -farm mitigation strategies.

The intensive agriculture practices improved the crop productivity but escalated energy inputs (EI) and carbon foot print (CF) which contributes to global warming. Hence designing productive, profitable crop management practices under different production systems with low environmental impact (EI and CF) is the need of the hour. To identify the practices, quantification of baseline emissions and the major sources of emissions are required. Indian agriculture has diversified crops and production systems but there is dearth of information on both EI and CF of these production systems and crops. Hence the present study was an attempt to find hot spots and identify suitable strategies with high productivity, energy use efficiency (EUE) and carbon use efficiency (CUE). Energy and carbon balance of castor, cotton, chickpea, groundnut, maize, rice (both rainfed and irrigated), wheat, sugarcane (only irrigated), pigeon pea, soybean, sorghum, pearl millet (only rainfed) in different production systems was assessed. Field specific data on different crop management practices as well as grain and biomass yields were considered. Rainfed production systems had lower EI and CF than irrigated system. The nonrenewable sources of energy like fertilizer (64 %), irrigation (78 %), diesel fuel (75 %) and electricity (67 %) are the major source of energy input. Rainfed crops recorded higher CUE over irrigated condition. Adoption of technologies like efficient irrigation strategies (micro irrigation), enhancing fertilizer use efficiency (site specific nutrient management or slow release fertilizer), conservation agriculture (conservation or reduced tillage) rice cultivation methods (SRI or Direct seeded rice) were the mitigation strategies. These results will help policy makers and stake holders in adoption of suitable strategies for sustainable intensification.

Crystal structure of IRF-3 in complex with CBP.

Transcriptional activation of interferon beta (IFN-beta), an antiviral cytokine, requires the assembly of IRF-3 and CBP/p300 at the promoter region of the IFN-beta gene. The crystal structure of IRF-3 in complex with CBP reveals that CBP interacts with a hydrophobic surface on IRF-3, which in latent IRF-3 is covered by its autoinhibitory elements. This structural organization suggests that virus-induced phosphoactivation of IRF-3 triggers unfolding of the autoinhibitory elements and exposes the same hydrophobic surface for CBP interaction. The structure also reveals that the interacting CBP segment can exist in drastically different conformations, depending on the identity of the associating transcription cofactor. The finding suggests a possible regulatory mechanism in CBP/p300, by which the interacting transcription factor can specify the coactivator's conformation and influence the transcriptional outcome.

Contribution of Ser386 and Ser396 to activation of interferon regulatory factor 3.

IRF-3, a member of the interferon regulatory factor (IRF) family of transcription factors, functions in innate immune defense against viral infection. Upon infection, host cell IRF-3 is activated by phosphorylation at its seven C-terminal Ser/Thr residues: (385)SSLENTVDLHISNSHPLSLTS(405). This phosphoactivation triggers IRF-3 to react with the coactivators, CREB-binding protein (CBP)/p300, to form a complex that activates target genes in the nucleus. However, the role of each phosphorylation site for IRF-3 phosphoactivation remains unresolved. To address this issue, all seven Ser/Thr potential phosphorylation sites were screened by mutational studies, size-exclusion chromatography, and isothermal titration calorimetry. Using purified proteins, we show that CBP (amino acid residues 2067-2112) interacts directly with IRF-3 (173-427) and six of its single-site mutants to form heterodimers, but when CBP interacts with IRF-3 S396D, oligomerization is evident. CBP also interacts in vitro with IRF-3 double-site mutants to form different levels of oligomerization. Among all the single-site mutants, IRF-3 S396D showed the strongest binding to CBP. Although IRF-3 S386D alone did not interact as strongly with CBP as did other mutants, it strengthened the interaction and oligomerization of IRF-3 S396D with CBP. In contrast, IRF-3 S385D weakened the interaction and oligomerization of IRF-3 S396D and S386/396D with CBP. Thus, it appears that Ser385 and Ser386 serve antagonistic functions in regulating IRF-3 phosphoactivation. These results indicate that Ser386 and Ser396 are critical for IRF-3 activation, and support a phosphorylation-oligomerization model for IRF-3 activation.

Insights into interferon regulatory factor activation from the crystal structure of dimeric IRF5.

Interferon regulatory factors (IRFs) are essential in the innate immune response and other physiological processes. Activation of these proteins in the cytoplasm is triggered by phosphorylation of serine and threonine residues in a C-terminal autoinhibitory region, which stimulates dimerization, transport into the nucleus, assembly with the coactivator CBP/p300 and initiation of transcription. The crystal structure of the transactivation domain of pseudophosphorylated human IRF5 strikingly reveals a dimer in which the bulk of intersubunit interactions involve a highly extended C-terminal region. The corresponding region has previously been shown to block CBP/p300 binding to unphosphorylated IRF3. Mutation of key interface residues supports the observed dimer as the physiologically activated state of IRF5 and IRF3. Thus, phosphorylation is likely to activate IRF5 and other family members by triggering conformational rearrangements that switch the C-terminal segment from an autoinihibitory to a dimerization role.

Competition between Ski and CREB-binding protein for binding to Smad proteins in transforming growth factor-beta signaling.

The family of Smad proteins mediates transforming growth factor-beta (TGF-beta) signaling in cell growth and differentiation. Smads repress or activate TGF-beta signaling by interacting with corepressors (e.g. Ski) or coactivators (e.g. CREB-binding protein (CBP)), respectively. Specifically, Ski has been shown to interfere with the interaction between Smad3 and CBP. However, it is unclear whether Ski competes with CBP for binding to Smads and whether they can interact with Smad3 at the same binding surface on Smad3. We investigated the interactions among purified constructs of Smad, Ski, and CBP in vitro by size-exclusion chromatography, isothermal titration calorimetry, and mutational studies. Here, we show that Ski-(16-192) interacted directly with a homotrimer of receptor-regulated Smad protein (R-Smad), e.g. Smad2 or Smad3, to form a hexamer; Ski-(16-192) interacted with an R-Smad.Smad4 heterotrimer to form a pentamer. CBP-(1941-1992) was also found to interact directly with an R-Smad homotrimer to form a hexamer and with an R-Smad.Smad4 heterotrimer to form a pentamer. Moreover, these domains of Ski and CBP competed with each other for binding to Smad3. Our mutational studies revealed that domains of Ski and CBP interacted with Smad3 at a portion of the binding surface of the Smad anchor for receptor activation. Our results suggest that Ski negatively regulates TGF-beta signaling by replacing CBP in R-Smad complexes. Our working model suggests that Smad protein activity is delicately balanced by Ski and CBP in the TGF-beta pathway.

Smad3 allostery links TGF-beta receptor kinase activation to transcriptional control.

Smad3 transduces the signals of TGF-betas, coupling transmembrane receptor kinase activation to transcriptional control. The membrane-associated molecule SARA (Smad Anchor for Receptor Activation) recruits Smad3 for phosphorylation by the receptor kinase. Upon phosphorylation, Smad3 dissociates from SARA and enters the nucleus, in which its transcriptional activity can be repressed by Ski. Here, we show that SARA and Ski recognize specifically the monomeric and trimeric forms of Smad3, respectively. Thus, trimerization of Smad3, induced by phosphorylation, simultaneously activates the TGF-beta signal by driving Smad3 dissociation from SARA and sets up the negative feedback mechanism by Ski. Structural models of the Smad3/SARA/receptor kinase complex and Smad3/Ski complex provide insights into the molecular basis of regulation.

Crystal structure of IRF-3 reveals mechanism of autoinhibition and virus-induced phosphoactivation.

IRF-3, a member of the interferon regulatory factor (IRF) family of transcription factors, functions as a molecular switch for antiviral activity. IRF-3 uses an autoinhibitory mechanism to suppress its transactivation potential in uninfected cells, and virus infection induces phosphorylation and activation of IRF-3 to initiate the antiviral responses. The crystal structure of the IRF-3 transactivation domain reveals a unique autoinhibitory mechanism, whereby the IRF association domain and the flanking autoinhibitory elements condense to form a hydrophobic core. The structure suggests that phosphorylation reorganizes the autoinhibitory elements, leading to unmasking of a hydrophobic active site and realignment of the DNA binding domain for transcriptional activation. IRF-3 exhibits marked structural and surface electrostatic potential similarity to the MH2 domain of the Smad protein family and the FHA domain, suggesting a common molecular mechanism of action among this superfamily of signaling mediators.