Transcriptomic Response of Huanglongbing-Infected Citrus sinensis Following Field Application of a Microbial Fermentation Product.

Huanglongbing (HLB) is considered the most destructive disease in Citrus production and threatens the future of the industry. Microbial-derived defense elicitors have gained recognition for their role in plant defense priming. This work assessed a 5% (V/V) microbial fermentation application (MFA) and its role in the elicitation of defense responses in HLB-infected Citrus sinensis trees following a foliar application with a pump sprayer. Using a PCR detection method, HLB infection levels were monitored in healthy and infected trees for 20months. Nutrient analysis assessed N, P, K, Ca, Mg, Mn, Zn, Fe, B, and Cu concentrations in the trees. MFA significantly increased Cu concentrations in treated trees and resulted in the stabilization of disease index (DI) in infected trees. Initial real-time qPCR analysis of defense-associated genes showed a significant increase in pathogenesis-related protein 2 (PR2) and phenylalanine ammonia lyase (PAL) gene expression in healthy and HLB-infected trees in response to MFA. Gene expression of PR2 and PAL peaked 6h post-microbial fermentation application during an 8-h sampling period. A transcriptomic assessment using GeneChip microarray of the hour 6 samples revealed differential expression of 565 genes when MFA was applied to healthy trees and 909 genes when applied infected citrus trees when compared to their respective controls. There were 403 uniquely differentially expressed genes in response to MFA following an intersectional analysis of both healthy and infected citrus trees. The transcriptomic analysis revealed that several genes associated with plant development, growth, and defense were upregulated in response to MFA, including multiple PR genes, lignin formation genes, ROS-related genes, hormone synthases, and hormone regulators. This study provides further evidence that MFA may play an important role as a plant elicitor in an integrated pest management strategy in citrus and other agronomically important crops.

Oral administration of NPC43 counters hyperglycemia and activates insulin receptor in streptozotocin-induced type 1 diabetic mice.

INTRODUCTION: Adenosine, 5'-Se-methyl-5'-seleno-,2',3'-diacetate (NPC43) is a recently identified small, non-peptidyl molecule which restores normal insulin signaling in a mouse model of type 2 diabetes (Lan et al). The present study investigated the ability of NPC43 as an oral and injectable insulin-replacing agent to activate insulin receptor (INSR) and counter hyperglycemia in streptozotocin (STZ)-induced type 1 diabetic (T1D) mice. RESEARCH DESIGN AND METHODS: In this study, STZ was intraperitoneally injected into wild-type mice to induce hyperglycemia and hypoinsulinemia, the main features of T1D. These STZ-induced T1D mice were given NPC43 orally or intraperitoneally and blood glucose levels were measured using a glucometer. Protein levels of phosphorylated and total Insrß, protein kinase B (Akt) and AS160 (critical for glucose uptake) in the skeletal muscle and liver of STZ-induced T1D mice following oral NPC43 treatment were determined by western blot analysis. In addition, hepatic expression of activated Insr in STZ-induced T1D mice after intraperitoneal NPC43 treatment was measured by ELISA. Student's t-test was used for statistical analysis. RESULTS: Oral administration of NPC43 at a dose of 5.4 or 10.8 mg/kg body weight (mpk) effectively lowered blood glucose levels in STZ-induced T1D mice at ≥1 hour post-treatment and the glucose-lowering activity of oral NPC43 persisted for 5 hours. Blood glucose levels were also reduced in STZ-induced T1D mice following intraperitoneal NPC43 (5.4 mpk) treatment. Protein levels of phosphorylated Insrß, Akt and AS160 were significantly increased in the skeletal muscle and liver of STZ-induced T1D mice after oral NPC43 (5.4 mpk) treatment. In addition, activation of hepatic Insr was observed in STZ-induced T1D mice following intraperitoneal NPC43 (5.4 mpk) treatment. CONCLUSIONS: We conclude that NPC43 is a de facto fast-acting oral and injectable insulin mimetic which activates Insr and mitigates hyperglycemia in a mouse model of T1D.

Non-peptidyl small molecule, adenosine, 5'-Se-methyl-5'-seleno-, 2',3'-diacetate, activates insulin receptor and attenuates hyperglycemia in type 2 diabetic Leprdb/db mice.

The pathophysiology of type 2 diabetes mellitus (T2D) is characterized by reduced or absent insulin receptor (INSR) responsiveness to its ligand, elevated hepatic glucose output and impaired glucose uptake in peripheral tissues, particularly skeletal muscle. Treatments to reduce hyperglycemia and reestablish normal insulin signaling are much sought after. Any agent which could be orally administered to restore INSR function, in an insulin-independent manner, would have major implications for the management of this global disease. We have discovered a non-peptidyl small molecule, adenosine, 5'-Se-methyl-5'-seleno-, 2',3'-diacetate [referred to as non-peptidyl compound #43 (NPC43)], which restores INSR signaling in the complete absence of insulin. Initial screening of numerous compounds in human HepG2 liver cells revealed that NPC43 significantly inhibited glucose production. The compound was potently anti-hyperglycemic and anti-hyperinsulinemic in vivo, in insulin-resistant T2D Leprdb/db mice, following either acute or chronic treatment by oral gavage and intraperitoneal injection, respectively. The compound acted at the level of INSR and activated it in both liver and skeletal muscle of Leprdb/db mice. In cell culture, the compound activated INSR in both liver and skeletal muscle cells; furthermore, it cooperated with insulin to depress glucose-6-phosphatase catalytic subunit (G6pc) expression and stimulate glucose uptake, respectively. Our results indicated that the compound directly interacted with INSRα, triggering appropriate phosphorylation and activation of the receptor and its downstream targets. Unlike insulin, NPC43 did not activate insulin-like growth factor 1 receptor in either liver or skeletal muscle. We believe this compound represents a potential oral and/or injectable insulin replacement therapy for diabetes and diseases associated with insulin resistance.