Arabidopsis DPB3-1, a DREB2A interactor, specifically enhances heat stress-induced gene expression by forming a heat stress-specific transcriptional complex with NF-Y subunits.

DEHYDRATION-RESPONSIVE ELEMENT BINDING PROTEIN2A (DREB2A) is a key transcription factor for drought and heat stress tolerance in Arabidopsis thaliana. DREB2A induces the expression of dehydration- and heat stress-inducible genes under the corresponding stress conditions. Target gene selectivity is assumed to require stress-specific posttranslational regulation, but the mechanisms of this process are not yet understood. Here, we identified DNA POLYMERASE II SUBUNIT B3-1 (DPB3-1), which was previously annotated as NUCLEAR FACTOR Y, SUBUNIT C10 (NF-YC10), as a DREB2A interactor, through a yeast two-hybrid screen. The overexpression of DPB3-1 in Arabidopsis enhanced the expression of a subset of heat stress-inducible DREB2A target genes but did not affect dehydration-inducible genes. Similarly, the depletion of DPB3-1 expression resulted in reduced expression of heat stress-inducible genes. Interaction and expression pattern analyses suggested the existence of a trimer comprising NF-YA2, NF-YB3, and DPB3-1 that could synergistically activate a promoter of the heat stress-inducible gene with DREB2A in protoplasts. These results suggest that DPB3-1 could form a transcriptional complex with NF-YA and NF-YB subunits and that the identified trimer enhances heat stress-inducible gene expression during heat stress responses in cooperation with DREB2A. We propose that the identified trimer contributes to the target gene selectivity of DREB2A under heat stress conditions.

Correlation of Solid State and Solution Coordination Numbers with Infrared Spectroscopy in Five-, Six-, and Eight-Coordinate Transition Metal Complexes of DOTAM.

Three new DOTAM (1,4,7,10-tetrakis(acetamido)-1,4,7,10-tetraazacyclododecane) complexes have been synthesized and characterized by X-ray crystallography: [Co(DOTAM)]Cl2•3H2O, [Ni(DOTAM)]Cl2•4H2O, and [Cu(DOTAM)](ClO4)2•H2O. Solid state and solution IR spectroscopic features for a series of [M(DOTAM)]2+ complexes (M=Mn, Co, Cu, Ni, Ca, Zn) correlate with solid state and solution coordination numbers. [Co(DOTAM)]2+, [Ni(DOTAM)]2+, and [Zn(DOTAM)]2+ are demonstrated to be six-coordinate in both the solid state and in solution, while [Mn(DOTAM)]2+ and [Ca(DOTAM)]2+ are eight-coordinate in the solid state and remain so in solution. [Cu(DOTAM)]2+, which is five-coordinate by X-ray crystallography, is shown to increase its coordination number in solution to six-coordinate.

Specific amino acids regulate Sestrin2 mRNA and protein levels in an ATF4-dependent manner in C2C12 myocytes.

BACKGROUND: Sestrin2 is a conserved protein in several species, and its expression is upregulated in cells under various environmental stresses. Sestrin2 content is involved in the function of mechanistic target of rapamycin complex 1 (mTORC1) in mouse embryonic fibroblasts and C2C12 cells. METHODS: C2C12 cells were treated with amino acid-free DMEM (AF-DMEM) for 5 h. The effects of the addition of specific amino acids to AF-DMEM on Sestrin2 mRNA and protein expression were examined using RT-qPCR and western blotting, respectively. The mechanism by which amino acids regulate Sestrin2 mRNA expression was examined using blocking and siRNA experiments. RESULTS: AF-DMEM increased the mRNA and protein levels of both Sestrin2 and activating transcription factor 4 (ATF4). The addition of a specific amino acid changed Sestrin2 mRNA and protein levels. The response pattern of Sestrin2 to specific amino acids was similar to that of ATF4. ATF4 siRNA reduced Sestrin2 mRNA levels. AF-DMEM increased eukaryotic initiation factor 2α (eIF2α) phosphorylation as early as 10 min after the treatment; however, ATF4 and Sestrin2 were increased 300 min after the treatment. Nuclear factor erythroid 2-related factor 2 and pancreatic and duodenal homeobox 1 siRNA did not affect ATF4 and Sestrin2 mRNA expression. CONCLUSIONS: Specific Amino acids regulate Sestrin2 levels in an ATF4-dependent manner in C2C12 cells. GENERAL SIGNIFICANCE: The results of the present study indicate that amino acids regulate levels of Sestrin2, which might cause phenotypic alterations, including mTORC1 activity, in C2C12 cells.