Maintenance of energy homeostasis is a fundamental requirement for organismal fitness: defective glucose homeostasis underlies numerous metabolic diseases and cancer. At the cellular level, the ability to sense and adapt to changes in intracellular glucose levels is an essential component of this strategy. The basic helixloop-helix-leucine zipper (bHLHZip) transcription factor complex MondoA-Mlx plays a central role in the transcriptional response to intracellular glucose concentration. MondoA-Mlx complexes accumulate in the nucleus in response to high intracellular glucose concentrations and are required for 75% of glucose-induced transcription. We show here that, rather than simply controlling nuclear accumulation, glucose is required at two additional steps to stimulate the transcription activation function of MondoA-Mlx complexes. Following nuclear accumulation, glucose is required for MondoA-Mlx occupancy at target promoters. Next, glucose stimulates the recruitment of a histone H3 acetyltransferase to promoter-bound MondoA-Mlx to trigger activation of gene expression. Our experiments establish the mechanistic circuitry by which cells sense and respond transcriptionally to various intracellular glucose levels.

While basic mechanisms of several major molecular chaperones are well understood, this machinery has been known to be involved in folding of only limited number of proteins inside the cells. Here, we report a chaperone type of protein folding facilitated by interaction with RNA. When an RNA-binding module is placed at the N-terminus of aggregation-prone target proteins, this module, upon binding with RNA, further promotes the solubility of passenger proteins, potentially leading to enhancement of proper protein folding. Studies on in vitro refolding in the presence of RNA, coexpression of RNA molecules in vivo and the mutants with impaired RNA binding ability suggests that RNA can exert chaperoning effect on their bound proteins. The results suggest that RNA binding could affect the overall kinetic network of protein folding pathway in favor of productive folding over off-pathway aggregation. In addition, the RNA binding-mediated solubility enhancement is extremely robust for increasing soluble yield of passenger proteins and could be usefully implemented for high-throughput protein expression for functional and structural genomic research initiatives. The RNA-mediated chaperone type presented here would give new insights into de novo folding in vivo.

The fusion of soluble partner to the N terminus of aggregation-prone polypeptide has been popularly used to overcome the formation of inclusion bodies in the E. coli cytosol.The chaperone-like functions of the upstream fusion partner in the artificial multidomain proteins could occur in de novo folding of native multidomain proteins. Here, we show that the N-terminal domains of three E. coli multidomain proteins such as lysyl-tRNA synthetase, threonyl-tRNA synthetase, and aconitase are potent solubility enhancers for various C-terminal heterologous proteins. The results suggest that the N-terminal domains could act as solubility enhancers for the folding of their authentic C-terminal domains in vivo. Tandem repeat of N-terminal domain or insertion of aspartic residues at the C terminus of the N-terminal domain also increased the solubility of fusion proteins, suggesting that the solubilizing ability correlates with the size and charge of N-terminal domains. The solubilizing ability of N-terminal domains would contribute to the autonomous folding of multidomain proteins in vivo, and based on these results, we propose a model of how N-terminal domains solubilize their downstream domains.

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